Monthly Archives: February 2017

Retrograde Polymorphism during Sublethal Environmental Stress

By Jean Guex (ISTE, University of Lausanne, Switzerland)

A detailed discussion of the effect of sublethal stress on the evolution of invertebrates is given in the book of Guex (2016; review in Guex and Verkhratsky, this issue). In this paper we will present a brief description of the effect of major stress on the development and evolution of some foraminifera and ammonoids. In the two cases discussed below, we show that retrogradation does’nt generate only retrograde evolution but also retrograde polymorphism.

One notable case of polymorphisms was named “Buckman’s law of covariation”. Buckman first discovered that some extreme morphotypes of atavistic habitus with simple and archaic morphologies show a perfectly continuous morphological spectrum towards the most advanced forms in the same sedimentary beds (synchroneity of the ammonite population). One example given by Buckman concerns the beautiful and famous ammonite genus Amaltheus of the gibbosus group which first occur during the Upper Pliensbachian located at the transition between the Gibbosus and Spinatum zones in the Late Pliensbachian (see Guex 2016 for references). The evolute spinose zonal index Amaltheus gibbosus is an atavistic form generated from involute A. margaritatus which strongly resembles the ancestral morphology of the Sinemurian Eoderoceras.

We consider as fascinating the fact that some planctonic foraminifera (e.g. the lineage Ticinella Thalmanninella) display a covariation which is similar to that observed in some ammonoids. Foraminifera are well known to be extremely sensitive to environmental stress and several classical studies have shown that the variability of these organisms was polarized in function of thermal and chemical stress and by depth.

The evolutionary trend observed in the Ticinella Thalmaninella lineage illustrates the frequent evolutionary trend occurring in coiled shells (Fig.1): in the T. greenhornensisT. multiloculata plexus a strong keel is developed at the maximum lateral curvature of the chamber, demonstrating the relation between morphogen concentration and strong curvature of the membrane secreting the shell. During episodes of environmental stress such as anoxic events, a retrograde polymorphism is observed in these protists, which is perfectly similar to the case of Amaltheus described above, as illustrated in Fig.1.

The fact that Buckman’s First Law of Covariation applies to both unicellulars and metazoans proves that similar biochemical signals are at work in both types of organisms, the membrane of foraminifera functions in a way similar to the ammonoids’ mantle.

Fig.1 (a) Evolutionary lineage going from the simple and evolute ancestral planktonic foraminifera Ticinella (right) towards the involute and carinated Thalmaninella. The variability of a single synchronous population illustrated in Fig.1-a mirrors the evolutionary sequence observed in this group.

(b) Highly evolute “Eoderoceras-looking” (right) Amaltheus gibbosus showing a similar trend from evolute towards involute coiling (A.margaritatus) during a Pliensbachian environmental stress episode.

Technical Remark

Covariation depends on the internal shell geometry, namely the lateral and ventral curvature of the shell which controls the amount of morphogens present in the more or less curved mantle, the most salient ornamentation being present where the whorls are most curved, shells with slight angular bulges often being spinose or carinate and flat ones being almost smooth. Our empirical conclusion was that the covariation phenomenon could be explained within the framework of Gierer-Meinhardt’s reaction diffusion models. To prove that conclusion, we simulated the distribution of “morphogens” (in the physical sense) in a quadrangular body chamber and demonstrated that morphogens maxima are located, as expected, in the part of the mantle located in the angular parts of the shell. For this, we calculated a numerical solution of the Gierer-Meinhardt equations for a cross section through an ammonite shell, orthogonal to the growth axis. In the computation, the units of distance, time and concentration are arbitrary. The boundaries of the domains are supposed to be impervious for the inhibitor. The outer boundary (arc of circle) is unaffected by the activator whereas the other boundaries are susceptible to this factor. The choice of these boundary conditions is motivated by the following arguments: The activator is supposed to diffuse freely outside the mantle’s cells (or the membrane of foraminifera) into the environment (intercellular medium and sea water). The reaction-diffusion equations are solved numerically on a hexagonal mesh containing 1500 nodes corresponding to a hexagon radius of 0.23 units. The concentrations a(x,t) and h(x,t) are determined at each node of the mesh. The initial values of the concentrations at t = 0 correspond approximatively to the values taken from the (unstable!) homogenous stationary solution. We add small random deviations ε(x,0) to the concentrations of the activator to allow the system to leave the initially homogenous state. The stationary inhomogeneous solution is found using a standard iterative procedure (details in Guex 2016 with references). Similar conclusions were obtained by Newell et al. (2008) in their general study of phyllotaxy: “… buckling leads to a template for primordia, it is growth that leads to the visible primordial bumps and phylla. This growth is postulated to be a biochemical response, perhaps through chemical agents such as auxin, to the local stress or curvature inhomogeneities of the buckled surface …”. In our carbonate shelly invertebrates, the morphogens have obviously nothing to do with auxin but could simply be Ca2+ ions. The output of our calculation shows that the distribution of the activator and inhibitor in a bended shell displays very low concentration in the smooth part and very high concentration in the curved part, both in foraminifera and ammonoids. More recently, an alternative model leading to the same kind of conclusions has been proposed by Mercker et al. 2013 where the authors write that “biomechanical forces may replace the elusive long-range inhibitor and lead to formation of stable spatially heterogeneous structures without existence of chemical prepatterns. We propose new experimental approaches to decisively test our central hypothesis that tissue curvature and morphogen expression are coupled in a positive feedback loop”. It might be useful to note that a multitude of mathematical models can simulate the pattern formation in shelly organisms. To illustrate this we can mention the fact that phyllotaxy, a domain studied by botanists, has been formally described by models developed in three totally different fields of mathematics: pure geometry by Van Iterson (1907), pure physics by Douady and Couder (1992) and reaction-diffusion by Meinhardt et al. (1998). In other words it is clear that our model of spine formation in ammonites is obviously not the only possible (all refs. in Guex 2016).


Jean Guex (2016). Retrograde evolution during major extinction crises. Springerbriefs in Evolutionary Biology. (Personal copies: write to

How Deep is the Neuron?

By Tam Hunt

A conversation with Anirban Bandyopadhyay about new advances in neuroscience.

The study of consciousness is becoming more serious. From being a rather fringe field during the 20th Century it has in the new century become a more mainstream endeavor with substantial funding and many careers now being built on our effort to understand the mind, brain and their interactions.

Somewhat surprisingly, many large questions about the functioning of our psyches are still unanswered, including how memory works, or even where memories reside, why we sleep, why we dream, how free will works (or doesn’t) and many more.

And the biggest of our unsolved mysteries: what the heck is consciousness itself and how does it relate to matter and the brain?

We are on our way to answering these questions and key to good answers to these longstanding mysteries is a better understanding of how neurons work and how they interact. Neurons are the building blocks of the brain and our understanding of these specialized cells has improved immeasurably in recent decades.

As we’ll see below, however, there is much that we don’t know, including exactly how much computation goes on in each neuron. Some researchers, including Hameroff and Penrose, have highlighted the role of microtubules in adding many layers of complexity and computation to our current brain models. Others, like the researcher interviewed here, think that might just be the tip of the computational iceberg.

Anirban Bandyopadhyay is a senior researcher at the National Institute of Materials Science in Tsukuba, Japan, and an adjunct professor at the Michigan Technological University. He studies the brain and is creating a detailed unconventional model of the brain he calls the frequency fractal model. Anirban is on the cutting edge of research into the neuroscience of consciousness.

I met Anirban originally at the annual Science of Consciousness conference hosted by the University of Arizona, where he is a regular speaker. I interviewed Anirban by email for my upcoming book further examining the intersection of philosophy, science and spirituality.

My own thoughts on the nature of consciousness are detailed in a technical manner in a 2011 paper in the Journal of Consciousness and a book length treatment called Eco, Ego, Eros: Essays in Philosophy, Science and Spirituality.


  1. What are the most interesting questions in neuroscience today? 

The most interesting questions in neuroscience today are questions like what is information, what is memory and where is it located? The synaptic junction is widely believed to be the locus of memory storage, but it may not be true.

Neuroscience needs more effective eyes at the atomic scale to see what happens inside the neuron. The days of Hodgkin-Huxley and Sakman Neher, which led to the idea that information in a neuron is stored in the synaptic junctions only, are over. We are seeing more and more evidence that supports the idea that it is not the firing above threshold but below the synaptic threshold that plays the major role in information processing in the brain.

To reveal what is happening within neurons, we need to find elements that live inside the neuron and regulate sub-threshold firing in a sophisticated manner. So neuroscience is awaiting a major invention in the form of an “atomic eye” that would enable us to take the next quantum leap in understanding information processing in the brain at a deeper level.


  1. Do you find philosophical questions like the “hard problem” of consciousness intriguing or useful? Where do you fall on the spectrum from idealism to panpsychism to materialism?

Whether we tag this the “awesome problem” or “hard problem” or “jackass problem,” the fact remains that unless we better define what consciousness is, mere adjective tags don’t serve much good.

I don’t consider the various philosophical positions on the nature of consciousness that important when we consider that the universe and human minds are frequency fractals that generate frequency wheels. Thus, inside the giant frequency wheel of the universe, our human minds are a simple subset. This position may fairly be labeled panpsychism because mind exists at levels in this model of the universe.


  1. Do you have a preferred theory of consciousness, such as Hameroff and Penrose’s Orchestrated Objective Reduction (Orch OR) theory? Or do you have your own theory of consciousness? 

I am trying to derive a better definition of consciousness, and I prefer to experimentally verify sensible models of consciousness. I like Orch OR but I feel that it is too early to have a well-developed theory of consciousness because there still isn’t enough information to create one.


  1. You stated in a 2014 interview: “Evolution is just a march to capture material or matter from nature and enrich that resonance chain.” Could you flesh out what this means?

We have an experimental laboratory where we build organic supramolecular structures. Our structures grow differently than a normal crystal. They change their symmetry over time and at various stages look very different. We have seen that when material growth is mostly incomplete, we find some of the resonance chains or vibrations are un-occupied, they need oscillators to complete this growth. What interaction enables the structure to evolve their geometry and generate new ones? We found that coupled resonance of elementary components may drive a very complex growth process. Elementary component seed A forms B, then B forms C, with each becoming more complex. The resonant oscillations of A creates a coupled electric field, which enables a new arrangement constituting B, and so on.

More generally, the drive to become more conscious is the primary drive in the universe, and this drive to become more conscious thus drives evolution. To be more conscious or more profoundly resonating with the universe is the objective of a living system. This drive results in structural modifications and we call this process evolution.


  1. What are the most promising “atomic eyes” that are being developed for looking more deeply into neurons?

What skin is to our body, cell membranes are to a neuron. The membrane’s ionic signals are strong and cellular electromagnetic signals appear as noise in a patch clamp, for example. Most of the noise elimination is done in the hardware and software using extensive filters. Above threshold, due to strong ionic bursts, the subtle changes in the potential are not visible. Patch clamp techniques can capture ions and neutralize them to estimate the membrane potential. This is a slow process, however, so it fails to map faster signals. The technology is limited in this key way.

Atom probes can, however, capture faster electromagnetic signals and we are thus able to track and record in far more detail the signal produced in the cells with these new tools. The atom probe is our new invention. It is a needle whose front part is 0.1 nanometers sharp. We cover it with an insulated area. Then, on the top we place another metal layer, creating a three-layered nano structure. It is an advancement of co-axial cable at the nano-scale.


  1. Orch OR focuses on microtubules as the key locus for quantum processing that leads to consciousness. Do you agree with this idea or are there other structures in the neuron that also play a heretofore unknown role in information processing and thus consciousness?

Microtubule and tubulin are extremely important for the information processing in the sub-neuronal scale, there is no doubt about this. There are, however, many other protein complexes involved in computation in neurons. For example, clathrin-SNARE complex, ribosome, proteosome, NOS (n+e), apoptosome and spliceosome etc. These five proteins complexes are as rich as microtubules and generate equally complex electromagnetic resonance bands similar to microtubules. In a fractal structure, there are various elements that play a role in filling up the major gaps in the resonance chain of the human brain. The resonance chain connects the smallest atomic structure to the largest sub-organs of the brain.


  1. In terms of the information processing ability of the human brain, if Orch OR or a similar model that focuses on sub-synaptic information processing is accurate, how many orders of magnitude of computational ability do we need to add in terms of creating viable human brain models? 

We need a new information theory to explain the decision making of the biological system. If we consider the brain to be a computer and then calculate how many bits, and how fast is it, we may miss some key details about how the brain really works. We have proposed that the human brain does not follow a traditional Turing tape model of computation; rather, it follows a fractal tape computational model. Transitioning conceptually from a Turing tape to a Fractal tape is easy: just consider that every single cell of a Turing tape also has a Turing tape inside it.

In a fractal tape based machine, we have a single bit, and then there are bits inside of that bit. At any time when you count the number of bits, it is always one. In a fractal system, the counting is never done the way we see it in a von Neumann type computer. The ability to build a brain-like machine depends on how many frequencies make a wheel. How complex is the wheel required to include all layers of computation present in the human brain? We add layers of fractals one above the other in our model and for human brain there are 12 layers. We need at least 12 layers of wheels to model the complexity of the human brain.

All memories are geometric in nature; here is a very brief summary of twelve kinds of memories that we found in the brain and how to edit them:

  1. Periodic memory: a chain of guitar string we see mostly in DNA, proteins, period of periods in the nodes of the string. You can edit the memory by changing the periodic length of elementary point oscillators.
    2. Spiral of spiral memory, in DNA and proteins, also in microtubules. Edit the memory by changing the twists and the number of hierarchical periods by inducing more and more twists.
    3. Vector memory, orientation of helices to make cavities inside protein. 3D orientation makes it a multipolar complex 3D vibrational element. Edit the memory by changing the orientation of column oscillators.
    4. Lattice memory, microtubule, actin-beta spectrin network, protein complex. Change the lattice parameters to edit the memory.
    5. Chemico-electric memory, chemical transmission cycles. There are nested time cycles highly interconnected, to edit the memory, change the diameter, phase, delay or starting points.
    6. Leak density of Cavity memory, ion diffusion holes in the cellular membrane. To edit the memory change the leak density on the cavity surface.
    7. Spiral geometric memory, assembly of neurons etc. To edit the memory, change the pitch-diameter-length of the spiral.
    8. Vortex or fractal memory, memory stored in the geometric parameters of vortex. A vortex has divergence parameters and scale repeat unit, these two parameters are changed to edit memory.
    9.Nodal & polar memory: Nodes in the spinal cord brain network (nodes in a tear drop). Geometry of the shapes are changed among 8 different choices from teardrop to ellipsoid to edit nodes and polarity memory.
    10. Electromechanical phase memory: organs sync and desync like a giant molecule. This memory is edited by changing the bond length or wiring length and distribution.
    11. Multipolar loop in phase space loop memory constitutes a “hyperspace memory.” This memory is non-physical and hence not editable.
    12. Phase of phase duality generator memory: hierarchical assembly of reality sphere. This memory is also non-physical and hence not editable.


7a. Ok, so if we view the human brain as a fractal tape type of computer rather than a Turing tape computer how many orders of magnitude of computation do we need to add to account for these 12 fractal layers of computation? Hameroff 2013 estimated about 1016 additional operations per neuron per second, for a total of 1021 operations per neuron per second, and a massive 1037 total operations per second for the entire brain, which accounts for the additional sub-synaptic microtubular computational operations. Do you agree? Or are there even more levels of computation going on inside neurons than Hameroff accounts for?

When we consider the fractal tape model, we cannot count the number of bits involved. Rather, what we count is always one rhythm. Instead of bits, or 0s and 1s, you have time cycles. We proposed the frequency fractal model of the human brain and introduced a very new kind of computing that rejects the Turing machine model of computation as well as Bertrand Russell’s way of thinking (Ghosh, S.; Aswani, K.; Singh, S.; Sahu, S.; Fujita, D.; Bandyopadhyay, A. Design and Construction of a Brain-Like Computer: A New Class of Frequency-Fractal Computing Using Wireless Communication in a Supramolecular Organic, Inorganic System. Information 2014, 5, 28-100). In our approach, you have only one time cycle or rhythm, but when you go deeper inside any point you find another time-cycle. So, there is no total number of operations per second. Those kinds of terms are applicable to those who believe in the Turing model of computing. For us, it is just one.

We argue that a user can see only the one clock in any given system and this clock constitutes a triplet of information: a seconds tick, a minutes tick and an hour’s tick. We have triplet clocks everywhere in all the layers of the human brain. We have already identified 350 different classes of cavities in the brain distributed over 12 layers nested one inside another. If each cavity resonator is an octave musical flute, then nearly 2,800 frequencies and time cycles compose one nested rhythm and that constitutes our total brain power. The current idea that more number of operations per second is an accurate approach to brain modeling is not right. That is the Turing way of thinking.

Information for us is a time cycle that can be modeled in particular geometric shapes. It is all about one rhythmic vibration that arises through integration of geometries using 2,800 frequencies over 10^20 Hz. Each vibration includes a “bing” moment (Hameroff’s metaphor for the moment of conscious awareness) and also a “silence” period. The “silence” contains the “phase”, and this holds the true information about the geometric shape of the particular time cycle. A band of frequencies always make a circular strip as it always vibrates periodically. Several such concentric circles make a frequency wheel that is an integration of the included time cycles. Such a fractal like integration of information (FIT) is fundamentally different than other theories of consciousness such as Tononi’s Integration Information Theory (IIT). Just one bit is enough for us to model the brain’s operations.

It takes only the product of 12 primes (1X3X5X7X11X13X17X19X23X29X31X37=10^11) oscillators to generate all possible patterns of wheels at each layer. This is the mathematically largest number of components we need to generate our brain dynamics. Our brain dynamics model is also relatively simple, with only eight steps to convert a tear-drop shape into an ellipsoid. These eight steps are enough to replicate all cavity shapes and their dynamics in the brain. The simplicity of our approach is a major benefit when compared to other computational approaches.


  1. Can you flesh out what you mean by resonance chains with respect to human consciousness or consciousness more generally? 

Every single element in the brain is a cavity resonator and has an electromagnetic resonance band. When several materials come together their resonance bands overlap to form a chain. Since the number of cavities in a larger cavity is finite, the chain is finite in length and as the guest cavities process only the geometric information, the total phase lag is always two π. Hence there is a time cycle. So, we convert a resonance chain into a wheel layer. Many nested layers make a frequency wheel and that constitutes our model of this type of system.


8a. What do you mean by brain cavity resonators and the geometric shapes used to model them?

For example, a cortical column is a cavity resonator. A neuron is also a cavity resonator. Inside the neuron the microtubule is like a flute and that’s also a cavity resonator. Tubulin proteins are dumbbell-shaped cavity resonators, and the alpha helices inside surrounded by beta sheets form yet another cavity resonator. A column of alpha helices is itself a flute-like cavity resonator.  Similarly, we can consider self-assembly or neural columns forming vital components of the brain, like nucleus or even hippocampus. Therefore, the whole brain is a giant leaky cavity resonator and then if we open it up we find a large number of cavity resonators inside. Our journey to open up and find the new cavities at each level continues until we reach the molecular scale.


  1. You mention missing resonance chains in our current models. What level of physical structure are you referring to?

We consider that every single biological element is a cavity resonator or a flute. You can integrate the flutes in two different ways. First, you place them side-by-side. This is called an iterative function system type fractal. Or you can place them “one inside another. That is called an escape time fractal. We can generate an analogue of any biological structure using the elementary cavities like flutes and integrating them side by side and/or “above and within”. We proposed a fusion of these two kinds of fractals as a generic approach of nature to construct an analogue of a biological systems.

As described above, a fusion of escape time fractal and iterative function system type fractals made of cavity resonators are generated to create an analogue of a biological system. This analogue structure vibrates in sets of resonance frequencies. Resonance can happen in various ways, like, for example, a tuning fork vibrates resonantly following a mechanical vibration. We have identified twelve different kinds of resonance frequencies that might be operating simultaneously in the human brain to make rhythms or cyclic vibrations, as follows:

  1. Electromagnetic rhythms [carrier is photon or electromagnetic wave is trapped in a cavity to generate beating or rhythm].
  2. Magnetic rhythm [spiral flow of electrons or ions, they are the carriers editing the magnetic flux, geometry of path forms the periodicity].
  3. Electrical potential rhythm [change in the arrangement of dipoles editing the electric field, fractal distribution of local resonators generate a time function of potential]
  4. Solitonic & quasi particle rhythm [carriers are solitons, defect in the order flows in a ordered structure, the ordered structure is edited to make a loop].
  5. Ionic diffusion rhythm, [ions are carriers, tube like cavities are formed in a circular shape or continuous path to generate a loop]
  6. Molecular chemical rhythm [molecules like proteins, enzymes, etc., are carriers, tube like cavities are formed and sensory systems make sure a circular signaling pathway]
  7. Quantum beating [spin is the carrier, wavefunctions interfere in a squeezed excited photonic, electromagnetic or spin state]
  8. Density of states rhythm [orbitals coupling, wave function modulation, virtual carrier, a virtual continuous loop is made]
  9. van-der Waal rhythm [atomic thermal vibration is looped in a spiral pathway].
  10. electro-mechanical rhythm [classical beating with a mechanical beating like tuning fork]
  11. Quasi-charge rhythm polaron, polariton [topological fractured band based continuous loops]
  12. Mechanical rhythm, sound wave [similarly elastic pathways to make a circuit of sound waves].

Now, we can put the resonance frequencies side by side, say, from 1 micro Hz to 1 peta Hz to create a chain. To understand resonance chains, just imagine that the chain of vibrations that starts from 1 micro Hz ends at 1 peta Hz. The group of frequencies arranged in a sequence for a system is the resonance chain and when vibration in the chain is repeated many times, it is a rhythm. A loop is represented by a circle. For twelve kinds of rhythms, several such circles can be put together and we get integration of that information, or what we call a frequency wheel.


  1. How does resonance relate specifically to the nature of consciousness? Do all resonating structures have some associated consciousness?

Our mathematical model shows that when the resonance chains in the form of a frequency wheel are drawn for a system, as described above, we get a composition of rhythms. A rhythm means, say, a sinusoidal wave that propagates continuously for an infinite time. We represent it using a single frequency value, which is the inverse of the time period of the waveform. Now, how could we combine frequencies? Let’s do some simple math here. A rhythm should have at least two frequencies. Or it could have three frequencies. If it has four frequencies they can be played as a pair of two. In this way we can integrate frequencies to make various kinds of rhythms.

What are the elementary compositions? For basic rhythms, which cannot be divided into smaller loops, we need prime numbers of frequencies to couple. For example, take a pair of frequencies and combine with that seven-frequency rhythm. This is a unit now and no one could find a sub-rhythm. This is why groups of prime numbers of different frequencies link together to form the architecture of information in the biological system.

Now, let’s do some more math. Try to find how many ways one could deconstruct a rhythm made of 12 frequencies? 2X (3X2) or 3X (2X2)? Both solutions are equally possible. This is a remarkable situation: we have a giant cavity resonator, say, our brain and we have two sets of rhythms, which exist in parallel. This occurs when one hardware system generates two replicas of the particular information structure and both can edit each other. This is called the mathematical criteria for consciousness.


  1. How many years are we from having a good working model of the human brain and mind?

We are creating the model now. We have already built one, which will be perfected over time. We feel strongly that redefining “information” and creating a new type of “information theory” is the key to move forward. We will have a good working model by the end of this year. We are writing a manuscript, and of course we will upload the entire database freely in the website, where we regularly update our research activities.


  1. Once we have a good working model of the human brain how many years do you think it will be before we’ll be able to upload human consciousness into a computational substrate?

We are already building small machines to demonstrate the duality of frequency fractals as an essential feature of consciousness and we have achieved some good progress. Once we generate the two distinct information structures from a singular hardware setup it will improve rapidly. We project that within five years we will patent the most primitive conscious machine. Note that the definition of consciousness is only “self-aware” here, it means a hardware that analyzes the environment and continuously edits it’s intelligence, knowledge and learning rules.

We will never, however, be able to upload human consciousness because artificial structures cannot be a replica. It will be a new consciousness of its own, but not a replica of human consciousness. The principle that we use to build the conscious machine suggests superposition of various features one on top of another. Such an integration has a simultaneity prerequisite and that cannot be replicated in any construction process at the molecular scale. In the future, if human race masters the  regulation of simultaneity, then it may be possible. Otherwise, however, we will be able to make humanlike machines, but never a true replica of any particular human

Pre-Biotic Evolution: Part IV. The Development of Electrochemically-Generated Energy Linkage, Extraction and Storage in Protocells

by Joseph H. Guth*

Published by the

Society for the Advancement of Metadarwinism, Volume 4 


One Scientist’s Overview and Perspectives


The first three parts of this series1, 2, 3 have described the time-dependent processes that  provide a plausibly likely pathway from the production and evolution of various atomic and molecular species through the formation of huge collections of varying complex chemical mixtures under early earth conditions.  The story continued with the general application of common physical activities and phenomena that would have commonly led to their packaging within simple membrane-enclosed volumes of such mixtures and their dispersal into new extracellular aqueous media.  This current chapter in pre-biotic evolution looks more closely at that and begins to mate these energy-requiring functions with energy sources and the growing complexities of a fully competent, self-sustaining version of protocell.

Uncountable numbers of protocell-like vesicular structures containing complex combinations of molecules and macro molecules collected over millenia.  They were derived in the primary laboratory, namely primordial earth, from the most diverse range of combinatorial chemical reactions.  Without living organisms yet present seeking out a food supply, those molecular libraries could have lasted for long periods in some environments.  Adsorbed onto mineral surfaces deep within the water-filled interstices of submerged clays and sediments, they could have remained relatively protected for greatly extended periods.   There, structures that contained macro-molecular assemblies that had reaction sequences within them, became the macro-molecular platforms for future super-complex branched predecessors of our current, well-regulated metabolic pathways made of various enzyme sequences.  Closed, lipid-based membranes spontaneously formed around many kinds of collected macro-molecule assemblages.  The new physicochemical

rules of lipids in water became the method and means for future combinations of these sequential functions to extend their evolutionary story through.  Simple fusion of all possible combinations then could have created brand new levels of complexity in which multiple end-products of simple pathway operation would have been generated in proximity with one another.  Such concentration of multiple new metabolic end-products  within single protocells now allowed larger scale leaps in the evolution of complex cell structure, design, operation and functioning.  At that point the rate at which evolution of inanimate matter into a truly self-sustaining state must have greatly accelerated.  With multiple occurrences of such breakthroughs at each of those critical points, this pre-biotic epoch appeared unchanging to the naked eye while major microscopic and chemical changes in growing complexity sprung forth on earth.

Looking back in retrospect at different pathways within modern life’s biochemical pathways and sub-cellular structures, we might very well be seeing a journal of the most successful types of ancient protocells that ultimately produced the first self-sustaining “living” early cells.  For example, the glycolytic pathway, an important pathway in most anaerobic as well as aerobic procaryotes and eucaryotes, regenerates adenosine triphosphate (ATP) from adenosine diphosphate (ADP) and inorganic phosphate in modern organisms as it breaks down six carbon carbohydrates down to 3 carbon forms in the initial stages of their complex energy-extracting metabolism.  This part of the modern cell’s overall metabolic pathway complexity, composed of a small number of sequentially-operating chemical catalysts (i. e., what are now called enzymes) could very well have developed through combinatorial chemistry followed by self-selection.  If then captured within liposomes as a “starter set” within a relatively few early pre-biotic protocells, this would have given them certain advantages over the many other kinds of protocells they were mixed in with in various niches around the early earth.  But some of those other types of protocells must have had their own fragmentary reaction sequences operating that, if combined within some future hybrid already possessing the glycolytic reaction sequence, then provides that evolutionary quantum leap to a new set of survival capabilities.  See Figure 1.

Figure 1.  Protein-rich protocells with differing sets of contained unitary functions produced in uncountable numbers must have been a common occurrence in many niches on early earth.  Examples might include an ocean foam collecting at the water’s edge on a beach or a kind of bubbling, floating pond scum.  Mixing of such collections in a somewhat random fashion would have been accompanied by all manner of fusion hybridizations between these different parental lines of biochemical and biophysical function-carrying units.
It is not so difficult to imagine that when a few of these functional pathways combined into a single protocellular package it could have generated a major enhancement in its survival properties and thus ability to persist.  And as those more successful hybrids out-performed and multiplied by simple physical scission in flowing rivulets and crashing waves, each resultant metabolizing vesicle regenerated more of its unique collection of molecules.  Their overall numbers then could preferentially increase over the less functional previous forms.  Once this new hybrid became dominant, further mixing and fusions with other functionally distinct protocells would allow subsequent quantum leaps of development to become the newest dominant forms.  At some distant step in this fusion recycling, the independently free-living protocell would have come into existence.

No one should ever expect the formative steps of life on earth to not have been hap-hazardous, chaotic but bordering on random, irrational or inelegant.  Nature is really messy that way.  The march to a living state had to be a very crooked one at best.  Though not the current prevailing view, this piecemeal building up of cellular complexity derived through a long series of steps involving protocellular fusions could also provide an even more important level of understanding.  This step-wise combining of already self-assembled and self-selected functioning protocell units presents a natural explanation for how the membrane-bound nucleus evolved in eucaryotes and even how multiple individual chromosomes came about in more complex asexually-reproducing modern uni-cellular types of free-living cells.  This evolution-controlled formation process could also be extended to explain how and from where other types of subcellular structures were derived.  It would explain the earliest origins of functionally separate sub-cellular structures such as lysosomes, peroxisomes, Golgi apparatus, ribosomes, microtubules, actin and myosin-related dynamic structures, endoplasmic reticulum membrane vesicles, and various kinds of storage, excretory and secretory vacuoles.  More elaborate and convoluted histories must have been followed however for the ultimate incorporation into early evolving protocells for the modern day, double-membrane chloroplast, mitochondrion and membrane-enclosed “super-control room”, the eucaryotic nucleus.  These subjects will be individually explored in future additions to these articles.

Such an incomplete picture then provides a somewhat mechanistic but still far-removed protocellular structure because no continuing energy-related processes are present or linked to the static metabolic schema that would have existed at that point.  Ultimately we need to now focus our attention on the energetics of what has developed in this picture to convert the finite existence of such protocells to a continuing, dynamic, persistent form we have come to know as organic-based terrestrial life.  This may not be the only such pathway but is argued to be a logical, consistent and realistic one that merits serious consideration.

In a previous part of this series, the author described the characteristics of a living system and alluded to the structural and mechanistic features that imbued that system with what we all must ultimately agree is the essence of the living state.  Life can be recognized as a persisting active, thermodynamically “uphill” set of processes contrary to what would happen if allowed to become depleted of energy sources and undergo normal spontaneous changes.  This has often been referred to as negatively entropic and focuses on the fact that instead of living cells slowly dissipating like a cube of sugar dissolving in a glass of room temperature water, they continuously work and function to hold together, repair and replace their losses in structural components and their non-random organization.  And here we are referring to the persistence of cells between their reproductive cycles.  That is an energy-requiring process.  Such behavior requires a continuing resupply of useful forms of energy and raw building materials to draw upon for this self repair behavior to out-perform Nature’s ever-present deteriorative influences.   So we might best be served in seeking a more detailed definition of the living state and how it is connected to energy sources.  Let us approach this by first defining the non-living, or dead state.

The most basic unit of life, an individual, free-living cell, can be in a living state one moment and dead in the next.  The difference is that the living version will persist indefinitely if continued to be supplied with all it needs to maintain its structure, composition and internal dynamic operations.  The dead cell will be irreversibly changed to a state where the structure and all organized and reorganizing processes cease.  It will no longer persist as that meta-stable processor of matter and energy.  In life, the cell behaved as a homeostatically-operating, steady-state flowing, chemical processing unit.   It is both a delicate and robust thing, the living cell.  And it has a continuing need for energy and raw matter input from which it maintains its overall structure.

To find the thing that physically defines that living state, one must closely observe, probe and analyze a living cell and follow the changes in it when a lethal injury is applied.  The simplest type of injury that can cause cell death is a rupture or uncontrolled leakage across the outer membrane of the cell.  Once such a breach of the plasma membrane occurs, extracellular fluid and its dissolved substances flows in at a much greater rate than it would if that membrane were intact.  At the same time, intracellular contents also uncontrollably leak out.

At the end of this injury scenario, the cell plasma membrane, like an empty burst balloon, floats away from the discharged dissolved cytosolic substances, the gelled cytosolic proteome, and sub-cellular debris.  The dead cell debris that used to be the nucleus, mitochondria, lysosomes, peroxisomes, endoplasmic reticulum, as well as any other specialized structures characteristic of the type of cell last only a short time before they also degrade.

In the living state, the semi-permeable plasma membrane completely enclosed and originally maintained a highly asymmetric set of concentration gradients, both inwardly and outwardly directed, in a permanent non-equilibrium condition.  It takes energy to maintain these gradients and part of the cell’s energy design is devoted to continuing to rebuild those gradient processes.

Thus the one thing that is always a marker for cell death is a loss of the ability of the cell to continue to maintain its internal milieu within very narrow and vital chemically-defined limits.  Internal dissolved calcium ion concentration in the cytosol compartment of modern eucaryotic cells typically ranges from about 10-8 up to 10-5 molar during various aspects of the cell’s life.  Important internal increases and decreases in cytosol calcium ion concentration are controlling features of the animal cell division cycle.  It is a key indicator of the biochemical activity patterns in operation.  But when approaching 1 millimolar in concentration, the cytosolic calcium concentration becomes a harbinger as well as agent for the cell’s death.  It directly causes loss of viability and the cessation of all its previous life processes.

The modern eucaryotic cell possesses multiple types of membrane-enclosed compartments.  Of course the outer plasma membrane provides this function for the cell as a whole, allowing it to maintain a different chemical milieu from the surrounding fluid.  Each type of sub-cellular organelle that has an uninterrupted enclosed membrane-bound volume also contains its preferred optimum milieu compatible with its unique chemical reaction sequences housed within.  All such membrane-bounded compartments must forever maintain the integrity and limited permeability of those internal compartments for the cell to remain in a completely viable condition.

Within a modern eucaryotic cell, the primary chemical reactions housed in various sub-cellular organelles frequently have quite different and conflicted optima for pH and other chemical reaction requirements.  The enzymes located in one compartment could rapidly destroy the enzymes or intermediates if they had access to the contents of other compartments.  The limited permeability and macro-molecular barrier functions of most cell membranes also provides protection and isolation of incompatibly self-destructive portions of the cell’s metabolic operating parts.   The semi-permeability of all such isolative membrane systems is also a long term requirement for generating and storing energy and creating each of its functional molecular parts.  In that sense it furthers the maintenance of the living state as well.

These facts are important to appreciate when looking for the clues to how life on earth began and then continued to evolve and improve its ability to persist, if not grow.  For without semi-permeable membranes and the necessary bioenergetic apparatus presenting itself in a life-compatible condition during the time of first capture, our protocells might form but not be able to persist.  In a somewhat literary sense, we can infer that the “vital energy” possessed by all living things can be biophysically defined in terms of chemical and electrical properties of the cell membrane and chemical gradients as produced through the asymmetric transport and longer duration storage of a multitude of differing chemical species.

The Origins of Bioenergetics

When biophysicists characterize modern cell plasma membranes, whether from procaryotes or eucaryotes, virtually all normal nutrients, specific cell secretory products and waste products transfer through such membranes by way of specific or relatively non-specific transport molecules or channels in those membranes.  If we begin with more of substance A on one side of such a membrane than on the other, the natural or spontaneous tendency is for such a gradient of concentration of A to flow in a net fashion from the higher to lower concentration until it arrives at final equilibrium concentrations. Those concentrations, or more formally, the chemical activities, becomes equal on both sides.  Even at equilibrium, the transfer of molecules or atoms of A continue, but the transfer rate in one direction would be equal to the back transfer rate in the other direction in a non-energized, permeable.

It is here that a series of seminal events took place during the early origins of life on earth.  These events must have occurred over a long period and many variations must have been present simultaneously.  In a very real sense, development of trans-membrane transport mechanisms must have followed a similar script through the genesis engine of  combinatorial chemistry.  That continuously operating agency would have led to the highly diverse molecular species that became available as the early building blocks of the membranes that formed the basic protocells.  These events included the development of passive, assisted and active transporting systems that not only spanned those protocell membranes, but were themselves asymmetrically co-orientated in a group fashion across such membranes.   These allowed dissolved substances that could closely interact with them to essentially move in one direction and not in the other direction, very much like a check valve or back-flow preventer in a water pipe.

So how could a collection of transport molecules get embedded in a phospholipid bilayer membrane all in the same trans-membrane orientation?  If combining in a purely random fashion, equal numbers of a given trans-membrane transporter could become embedded in a membrane oriented in opposing directions.  That would not only be self-defeating, it would eliminate the ability for that membrane to be able to generate a chemical gradient across it.

Transport molecules are typically protein in composition.  They are uniquely composed of chains made from a combination of varying length sequences of more highly polar amino acids joined to internally-connected sequences of lower polarity, lipophilic amino acids.  The lower polarity sequences  generally anchor the middle of transport molecules within the phospholipid membrane interior.  They control the way in which the secondary protein structure forms through folding of the more polar portions of the chains.  Those more polar regions are hydrophilic and closely connected to the aqueous medium on either side of the membrane.  The more polar sequences and regions of the protein’s amino acid sequences are usually found either extending out from the membrane surface and into the highly polar aqueous phases on either side of the membrane.  This makes them more “visible” or accessible to the kinds of molecules that are seeking passage through the membrane.  Such polar amino acids can also be found in a hollowed out, tubular interior of the protein.  These interior trans-membrane pipelines with polar groups projecting inwards, create trans-membrane channels possessing more selective size, charge polarity and geometric constraints for what kinds of substances can pass through them.  They are similar to the same chemical criteria found within an enzyme’s active site.

Such membrane-compatible protein molecules when free can form quasi-crystallized, weakly-aggregated films at or near air-water interfaces.  If phospholipids are also present, these amphiphilic molecules spontaneously form mono-, bi- and multi-layered membranes that spread across the surface of any available water-air interfaces.  Their appearance is quite like that of the black and multi-colored sheen of water contaminated by petroleum oil.  They have a strong tendency to keep spreading thinner and thinner and if the surface area is large enough, black single bi-layer membranes form.  The co-presence of what we might call proto-transport molecules floating as a regularly-ordered, two-dimensional, quasi-crystalline aggregate just under the bi-layer allows them to approach the bi-layer from one direction and become embedded in it in a uniformly oriented fashion.

An alternative means of accomplishing this same goal could utilize a solid mineral surface that presented adsorption sites for the proto-transport molecules to first attach to.   Following attachment to the mineral surface, the floating phospholipid bi-layer could be juxtaposed to that surface through direct contact streaming or evaporation of the water phase with deposition of the bilayer onto the protein-coated surfaces.  The result is a transfer of the proto-transport protein molecules from the mineral surface to a single side of the bi-layer membrane sheet.  And once this formation of an asymmetrically distributed collection of protein molecules is completed, uni-directional semi-permeability and asymmetric chemical reactions can take place across the membrane after enclosed vesicles are produced from such larger sheets through different types of physical agitation.

Such a mechanism for the formation of asymmetric membrane functions would ultimately be highly inefficient and lack stability and reproducibility.  This set of events are simply offered as a short term means to an end.  That end is the final bringing together of all the necessary structures, activities and components that are finally capable of indefinite persistence, and packaging them within a membrane-bounded, single small vesicular volume.  In other words, the crucial point in which they have the basic starting capabilities of life.

Electrochemistry and Life

Up until now we have only looked at the chemical complexity, reaction conditions and ability for a protocell membrane vesicle to concentrate and maintain a gradient across  its membrane.  The generation of the gradient, requiring an external source of usable energy, is very much analogous to a rechargeable battery.  The external energy source would be the recharger plugged into its energy source.  In the battery model, initially electrically net neutral chemical species are dissociated and actively separated or pulled into different fluids. The oppositely charged particles move to opposite sides of a semi-permeable, somewhat electrically insulative bridge or barrier.  If it is too highly insulative, it can store the charges for long periods but then they are greatly retarded in their movement back through the barrier.  That limits the rate that such a field can be subsequently utilized to perform other work.  So to operate properly, this barrier must still be slightly conductive in order to allow oppositely charged particles to pass through at a useful rate and allow the charged particles to rejoin and neutralize their free charges.

In cells, that barrier is the phospholipid portion of the bilayer membrane.  Thus, across various cell membranes, if such electrochemical reactions take place asymmetrically across those membranes, charges can be separated and stored for various amounts of time across a two-dimensional surface.  This becomes a direct simulacrum to an electrical capacitor.  If an electrically-charged capacitor is subsequently connected to an electric motor, that stored electrical field energy can drive the motor as the potential electric field energy collapses and the charges flow back together to reform electrically neutralized species again.  In a more fundamental fashion, electrochemically charged  membranes can provide a moderate amount of electrical charge storage and a means of tapping that stored energy into various types of useful work when the cell needs such input to extend its existence and persist.  The first free-living protocell was the original Walkman.

Useful energy storage can also be obtained through simply moving non-dissociable molecular or atomic species from one side of the membrane to the other, to form a simple concentration gradient of a neutral non-ionizable chemical compound.  Such a type of energy storage is more analogous to a dam that collects water and then can be linked to some energy-converting mechanism, such as a hydro-electric power station or a simple water wheel.   It is also analogous to the physical work generated within a growing plant as it rises out of the soil and gains new height against gravity through forces generated by osmotic pressure.  The usable energy, or work, in this latter example is performed from building up the potential energy by movement of water molecules from one side of semi-permeable membranes to the other.  That forces the water collected on one side of a fixed volume cell to be at higher pressure relative to the other side.  Thus osmotic pressure is also equivalent to stored potential gravitational energy.  The system’s natural tendency creates this type of energy storage whenever non-ionizable substances are actively transported across membranes.  The gradients they form have the natural tendency to spontaneously return to their equilibrium distribution of the various compounds and water of solvation while in earth’s gravitational field.

In our actual cell, the concentration gradient is driven by attracting more water to one side of a water-permeable membrane through addition of more non-dissociable particles  to that side.  This can be accomplished in several ways.  One simple way is to have a trans-membrane transporter move a single molecule from the outside of the cell to its interior where it is subsequently split into two molecules.  Osmotic pressure is based upon simple numbers of particles that are solvated.  The thermodynamic behavior of this extra energy is then stored in the form of the heats of hydration for each of the transported species plus net movement of extra water towards the side needing added water for solvation.  If the volume of that side is forced to be constant, that drawing in of the extra water would be converted to a physical force or pressure.  That is the fundamental cause of osmotic pressure.  And for completeness, it should be stated that osmotic pressure can be associated with either ionizable or non-ionizable soluble matter.

All living cells are involved with both kinds of energy storage.  Certain states can exist, such as within ungerminated spores and suspended animation, that do not necessarily preserve those qualities during such transient states.  Thus we can allow for a living system to become temporarily “non-living” during suboptimal conditions and re-start again after restoration of minimally supportive conditions.

For all of the functioning that we have previously been looking at, we have to still describe how chemical or physical energy taken in by the early protocells was first converted to a storage form, then stored for extended periods, and subsequently linked to and drawn upon to energize various life processes during energy-poor moments.  Those non-equilibrium concentration gradients spanning across membranes, and electrical fields located on the membrane surfaces, need to be able to be physico-chemically linked to molecular engines or processes that can only continue to operate by absorbing useful energy from the gradient and converting it into new cell “stuff”?  Let’s look briefly at the energy contained within chemical gradients.

The Nernst Equation

A well-known relationship was described by early electro-chemists between the relative difference for the concentration gradient existing for two aqueous solutions of a charged ion separated across a semi-permeable bridge and the strength of that species’ tendency to return to equilibrium through transference of negatively-charged electrons.  The physical form that this type of energy storage manifests is as an attractive electrical field created between oppositely charged species separated by the thickness of a relatively low-conductive dielectric membrane.   Different ions have different intrinsic dissociation energies based on the electronegativity and electron configurations of the atoms that compose them.

As the science of electrochemistry developed, electron transfers were found to occur not only between two identical-type ions simply based upon their relative concentrations in two different solutions, but also between different ionizable species.  But such phenomena have been occurring throughout the entirety of earth’s history without the intervention of Man.  Such electrochemical reactions existed before the first protocell was capable of forming on primordial earth.  As general features of such reaction chemistry, the following should be noted.

  1. All electrochemical reactions can occur in aqueous media (there are non-aqueous electrochemical reactions but those will be reserved for our future considerations regarding origins and evolution of extra-terrestrial life).
  2. As electrons are transferred from one ion to another, water molecules are included in the reaction steps. This leads to a simultaneous uptake or release of protons causing the reactions to also be affected by, and be capable of changing, the pH of the medium.
  3. Useful work can be effected if the two reacting species, also known as half-cell reactions, are first separated by a semi-permeable bridge (salt bridge) or membrane possessing selective channels connecting the two reacting solutions. Such a path is needed to allow the net flow of one type of ion in the appropriate direction, depending on whether the gradient is being recreated by an energy-utilizing regeneration mechanism or allowed to flow back spontaneously towards its equilibrium point by some kind of work-accomplishing process.  Such a system is mimicked by standard man-made rechargeable batteries.  Asymmetric chemical catalysts and electron carriers and their attendant reactions are commonly found embedded in and spanning biological membranes.
  4. If one has two such selective membranes separating three different solutions in tandem, this would increase the electrochemical potential in the same way that two standard batteries in series would double their combined output voltage. This pattern is also analogous to that of the eucaryotic cell in which the extracellular medium is separated by the plasma membrane from the cytosol and the cytosol is further separated from the inner mitochondrial matrix by the inner mitochondrial membrane.  Such a stacked strategy is also found in the electrogenic glands of the electric eel, Electrophorus, as well as in stacked flattened membrane vesicles in chloroplasts (thylakoids) and photosensitive retinal cells.  Synapses within the nervous system have a similar but less recognizable similarity to such stacked membrane vesicle packages.  But in each of these examples, gated flows of ions and non-dissociable chemical species flow into and out of those individual compartments while being apposed to other membrane-bounded compartments or cells.  This gives rise to the well-known action potential behavior of excitable membranes.  And that is also a commonly found trait of such electrochemically energized membrane phenomena.  For this reason, it might be reasonably expected that not only were simple membranes required at the dawn of the living state, but a low grade version of a gated, excitable membrane’s type of behavior would have presented exceptionally enhanced survival advantages to whatever protocells had developed it.

This multi-compartmented design of the eucaryotic cells however is much more complex than this.

(Certain terms must be used due to their universal meanings.  “Design”, “plan”, and “create” are such terms.  These have also come to be used in less rigorous non-scientific contexts.  For the sake of this subject matter, I will at times use them but caution the reader that nothing is implied in their use regarding extra-scientific meanings.)

Multiple chemical species are separated among these three aqueous compartments.  Each relatively impermeable to the phospholipid membrane framework but is transferred through the membranes by selective carrier or transport molecules embedded in the membranes.  The membranes also isolate and help maintain differing pH, chemical and ionic compositions between compartments that are optimized for the specific kinds of compartment-specific enzymology contained within those same compartments.  Osmotic pressures are relatively well-maintained across each type of membrane.  The membranes also isolate the biochemical reactions in one compartment from potentially incompatible reactions going on in another compartment.

Some of the reactions taking place across a given membrane are of an electrochemical nature.  For those reactions, the protons or electrons must pass through the membrane channels or transport molecules and during that passage, useful chemical work is capable of being coupled to them.  In energy-capturing membrane structures such as thylakoids in chloroplasts, this process works in the reverse direction under illumination and generates the pH gradients through electrons being first raised to excited states by photon absorption followed by linked chemical reactions being driven by them as they return to their ground states.  Those reactions are of the utmost importance in the evolution of life on earth.  For those reactions finally link all life on earth, directly or indirectly, to the main form of energy output of our nearest stellar neighbor, the Sun.  These will be considered in greater detail later.

The simple mathematical relationship describing energy storage for electrochemically active chemical species separated by a salt bridge or semi-permeable membrane was first offered by Nernst as follows:

E = (RT / zF) Ln [ion] outside  / [ion] inside

where E is the electromotive force in volts

R is the universal gas constant

T is the absolute temperature

z is the number of charges on the ion

F is Faraday’s constant

and the square brackets indicate concentration units

This relationship exists for each type of ion that can move across the membrane.  There are many kinds of ions that transport across membranes and this relationship exists for each type.  Other equations have been developed to better describe biological membrane potential.  The overall resultant electrical potential is the mathematical sum of each ion’s contribution.  In resting animal cells, the plasma membrane typically exhibits a resting electrical potential of about 70 millivolts, outside negative.  Whenever a cell becomes active, ions flow in and out of the outer membrane and the membrane potential changes in either a depolarizing direction or a hyperpolarizing direction.  All of these properties and variations are intrinsically part of and define the living state as  contrasted to a state of death.  The alternative, a state of pre-biosis in which life’s processes have never previously occurred within an early version of protocell, would also be found in a similar equilibrium or near equilibrium condition.

Osmotic Mechanisms of Energy Storage and Management: High Resolution

As any chemical species, ionizable or non-ionizable, moves through its preferred molecular channel from one side of a semi-permeable membrane to the other, water molecules to keep each ion or molecule in solution must follow.  As previously described, the phospholipid portion of the biological membrane has an important property in that at normal growth temperatures it allows the smaller water molecules to easily slip between the hydrocarbon chains making up the interior of the bi-layer.

Membranes have elasticity, plasticity and resilience.  This provides a back pressure to resist the continuing and unlimited movement of solutes across that membrane.  Biological membranes have some interesting behavior as they are stretched by osmotic swelling.  They become more non-specifically permeable to smaller ions and molecules.  As the membrane is further stretched, larger molecular weight ions and molecules and even macro-molecules begin to passively leak back down their concentration gradients.  When cells are osmotically over-stressed even further, macro-molecular solutes in the cytosol slowly exit the cell through the unbroken, but now more “porous” plasma membrane.  Even molecules and ions that normally move through specific transporter membrane proteins can also flow through these non-protein leakage pathways.  These capabilities are both important and useful to understanding how the self-assembled pre-biotic membranes must have behaved as the first “life-capable” membranes could have formed for the first time.

Combining All to Form the Living State

Our origin of life model must also include an adequate and plausible description of how all of the starter molecules necessary for a rudimentary metabolism were captured by the first pre-biotic liposomes.  Further, our narrative must also have a basic set of means (trans-membrane transporters) by which necessary substances move into and out of those early cells.  But even with all of that inventory list of life’s minimum necessary molecular content, we must also include a rudimentary energy-capturing and/or energy-converting mechanism that could collect, store, supply and recharge the necessary forms of energy needed to keep all of the internal chemical and physical activities continuing indefinitely thereafter.  It is here that our greatest challenge to life’s genesis will probably be found.

So how does one combine a set of uni-directionally selective, asymmetrically and uniformally disposed membrane transport molecules?  How can one understand the first genesis of sets of sequentially-operating enzyme complexes and then finally find a plausible and testable means of linking the internal reaction chemistry to the active transport of ions and molecules across the membrane against their concentration gradients?

Each of these collective functions embodies a number of types of molecules having to be maintained in close proximity and accurately juxtaposed to one another for extended periods of time.  The combined functions taken together creates an almost insurmountably complicated perception of a collection of components that behave with integrated complexity.  The whole seems much greater than the sum of its parts.  It is almost like trying to build a tall house of delicately balanced cards beginning at the top!  But remember, we have already described a relatively direct means of bringing all these functional units together.  It is through the prior formation and then subsequent hybridization of each functional type of protocell.  The first generation of protocells captured and utilized formative sequences of combinatorial chemical reactions.  Multiple liposomal formation captured differing collections of molecules, that is to say, different rudimentary but operating fragments of biochemistry.  The second generation of protocells then could form following fusions of different functionally distinct types of liposomal ensembles.  The more successful combinations self-selected and became the dominant types in their niches.  Life would not be held in abeyance for long.  It was trying to develop all over earth and at probably more than one location, it would finally become fully capable of unlimited growth of mass and able to begin actively sensing its chemical and physical environment.  And that sensing allowed it to become more tropic…  more able to seek out and meet its needs for energy, raw materials and optimal growth conditions.  At this point our protocells are beginning to “come alive”.  But there is much more to the story before they can be officially called alive!

Next:  Pre-Biotic Evolution.  Part V.  The Evolutionary Importance of Chemi-Osmosis and Electron Transport

Scientific and Forensic Services, Inc., Delray Beach, FL. and Norfolk, VA


  1. Guth, J. H. “Pre-Biotic Evolution:  From Stellar to Molecular Evolution”.  Society for the Advancement of Metadarwinism, Volume 1, November 19, 2014.   Accessible at
  2. Guth, J. H. “Pre-Biotic Evolution:  Pre-Biotic Chemical Oscillations and Linked Reaction Sequences”.  Society for the Advancement of Metadarwinism, Volume 2, June 12, 2015.   Accessible at
  3. Guth, J. H. “Pre-Biotic Evolution:    Transitioning to Animacy”.  Society for the Advancement of Metadarwinism, Volume 3, January 5, 2016.   Accessible at

© Copyrighted by Joseph H. Guth, 2016.  All rights reserved.

Phenotype as Agent for Epigenetic Inheritance

By  John S. Torday, MSc. PhD.  & William B. Miller, Jr, M.D.

Abstract: The conventional understanding of phenotype is as a derivative of descent with modification through Darwinian random mutation and natural selection. Recent research has revealed Lamarckian inheritance as a major transgenerational mechanism for environmental action on genomes whose extent is determined, in significant part, by germ line cells during meiosis and subsequent stages of embryological development. In consequence, the role of phenotype can productively be reconsidered. The possibility that phenotype is directed towards the effective acquisition of epigenetic marks in consistent reciprocation with the environment during the life cycle of an organism is explored. It is proposed that phenotype is an active agent in niche construction for the active acquisition of epigenetic marks as a dominant evolutionary mechanism rather than a consequence of Darwinian selection towards reproductive success. The reproductive phase of the life cycle can then be appraised as a robust framework in which epigenetic inheritance is entrained to affect growth and development in continued reciprocal responsiveness to environmental stresses. Furthermore, as first principles of physiology determine the limits of epigenetic inheritance, a coherent justification can thereby be provided for the obligate return of all multicellular eukaryotes to the unicellular state.

Keywords: phenotype; Darwin; Lamarck; germline; epigenetic; life cycle; niche construction

  1. Introduction

The recognition that the cell is the basis for eukaryotic evolution [1] as a manifestation of perpetual cellular principles renders phenotypes as epiphenomena, i.e., subordinate to the actual event. Such a perspective alters many otherwise dogmatic aspects of evolutionary biology. In particular, the systematic error of the perception of evolution as a stochastic phenomenon yields instead to phenotypes as mechanistic products, always directed towards identifiable cellular needs [2]. Such a change of focus is similar in type to David Bohm’s insight into dual explicate and implicate orders in the physical realm [3]. He stipulated that our evolved senses mislead us into regarding our conscious experience as an explicate ordering of an entire reality. Instead, a truer reality is a continuous stream sustained by both explicates and an additional set of subjective implicates of which we are not typically aware. Similarly then, in biologic terms, it can be presented that an explicate phenotype is fully dependent upon a steady flow of epigenetic implicates in a cellular continuum that mechanistically interconnects evolutionary development with the larger environment across space and time.

  1. Background

In conventional terms, any phenotype is assumed to be the end result of descent with modification through Darwinian random mutation and natural selection [4]. However, with the emergence of a contemporary re-appraisal of the importance of Lamarckian inheritance, the role of phenotype must be reconsidered as the effective primary means by which all organisms acquire information from the environment in the continuous maintenance of essential cellular requirements. These necessities are

expressed through all the cellular mechanisms that are directed towards sustaining cellular activity within homeostatic limits, defending cellular integrity and self-recognition. Thus, in eukaryotic multicellular organisms, phenotype becomes an agent promoting and incorporating epigenetic inheritance, rather than a simple manifestation of the concordance of intergenerational vertical genetic transmission exclusively based on selection.

This perspective on the significance of the phenotype is consistent with Niche Construction Theory [5,6] critically enacted at the cellular level. When cellular imperatives or principles such as maintenance of preferential homeostatic status and self-protection in both individual and collective terms are subjected to environmental stresses through epigenetic inheritance, evolution is understood as much more dynamic and environmentally interactive than via any filtering mechanism of Darwinian evolution. Importantly, this perspective faithfully reflects evolution’s origin as a self-organizing, self-referential mechanism that originates within the unicellular domain, and always remains contingent on it [6]. In this manner, phenotype becomes a directed product of cellular activities in response to epiphenomena rather than a mere result of random forces [7].

Perhaps even more importantly, the impact of epigenetic inheritance on the cell, and its physiologic limits, is amenable to hypothesis testing and falsification in a manner beyond any generally accepted Darwinian evolutionary narrative [8]. Selection still applies, but its precise role and its center of action are deeply reconsidered.

  1. The Water-Land Transition as the Epitome of Epigenetic Inheritance

There is evidence that life was initiated and then propelled on its evolutionary course in response to the physical constraints imposed by the environment [9], and therefore evolved in response to such major effectors as gravity [10], carbon dioxide [11], oxygen [12] and calcium [13] as epiphenomena to the cell [1].

The vertebrate transition from water to land was caused by the evaporation of water globally about 300 million years ago due to the accumulation of carbon dioxide in the atmosphere, causing a “greenhouse effect” [14]. An essential set of evolved traits was necessitated for the transition from water to land, critically dependent upon the lung as homologous with the swim bladder of bony fish [15]. This is particularly the case for physostomous fish [15], which have the homolog of a trachea (called the pneumatic duct) connecting the esophagus to the swim bladder. The swim bladder is derived from the foregut in both fish and land dwelling vertebrates [16]. Functionally, the effective inflation and deflation of both the swim bladder and lung are dependent on the production of surfactant by the gas gland epithelium lining the bladder lumen [17,18]. In the case of the swim bladder, the surfactant has been speculated to be necessary for preventing self-adherence of the walls of the bladder [19]. In the case of the lung, surfactant is necessary for preventing atelectasis, or alveolar collapse [20]. Alveoli are very small in diameter, thereby generating high surface tension based upon the Law of Laplace [21]. The physiologic stress of hypoxia forced selection pressure for the remodeling of the alveoli. The cell-cell interactions between the epithelial and mesenchymal components that mediate surfactant production [22] were modified through phylogeny and ontogeny in order to allow for the reduction in alveolar diameter, increasing the surface area-to-blood volume ratio for efficient gas exchange [23].

This mechanism for facilitated gas exchange, ever-dependent upon lipids, refers to the inception of cholesterol synthesis and its critical insertion into the cell membrane [24]. The facilitation of gas exchange through this cellular example of niche construction exemplifies how first principles of cellular requisites are put in service for oxygenation in unicellular organisms, exapted over billions of years through the implementation of homologous genetic motifs [25].

Starting from its origins, the spontaneous generation of micelles as lipids immersed in water [26], the reduction in entropy [27], in combination with chemiosmosis [28] and homeostasis [29] are assumed to have fostered life [30]. Unicellular life dominated the Earth for the first 4 billion years [31]. Then, fewer than 500 million years ago, multicellular organisms evolved from unicellular forms, likely

due to competition among prokaryotes able to mimic multicellularity through traits such as Biofilm [32] and Quorum Sensing [33]. Rising levels of oxygen in the atmosphere put selection pressure on prokaryotes, producing hopanoids that caused increased order within the cell wall [34], making it more permeable. The generation of oxygen by bacteria was hypothesized to have given rise to cholesterol, given that eleven atoms of oxygen are required to form one molecule of cholesterol [35]. The presence of cholesterol in the cell membrane of primitive eukaryotes promoted metabolism, oxygenation and locomotion, the basic principles of vertebrate evolution [36]. There was also an increase in atmospheric carbon dioxide [37,38], which dissolved in the oceans to form carbonic acid [39]. That acidity leached calcium out of the bedrock, threatening life due to the denaturing effects of calcium on proteins, lipids and nucleic acids. In response, unicellular organisms formed peroxisomes, organelles that use lipids to buffer intracellular calcium [40]. In such a scenario, the formation of calcium channels from lipids for the excretion of calcium was exapted to protect burgeoning eukaryotes. During the Phanerozoic Era, the greenhouse effect of rising levels of carbon dioxide [13] forced some evolving vertebrates to transition from water to land [4143], marking the beginnings of terrestrial life [44]. The adaptation to land gave rise to novel physiologic traits that had their origins in fish. The increased force of gravity on land [45] put great selection pressure on the skeletal system, altering it at least five times based on the fossil record [44]. Rising, fluctuating oxygen levels in the atmosphere necessitated remodeling of the internal organs, though there is no fossil evidence for these events. For example, we now know that the same genes that determine the swim bladder of bony fish determine the development of the lung [14]. Functionally, both the swim bladder and lung depend upon surfactant for their function [19,22], and the mechanisms that facilitated the evolution of the lung alveoli from the swim bladder delineate how and why these structures became smaller and more numerous [23] due to cellular interactions fostering evolutionary change. Moreover, the genes responsible for both skeletal and pulmonary evolution are involved in the evolution of the skin, kidney and brain. These adaptive changes were the net result of physiologic stress mediated by cellular-molecular damage to specific tissues and organs due to shear stress on microvessels generating radical oxygen species causing gene mutations and duplications [46]. This epigenetic remodeling pathway based upon the water-land transition exemplifies how phenotype becomes a product of environmental stress based upon cellular requisites in the cellular context of niche construction.

As apart from these deeply rooted cellular/molecular developmental mechanisms, there are now many direct examples of epigenetic factors influencing phenotype. The phenotypic differences between human monozygotic twins are now ascribed to epigenetic factors [47]. As an outgrowth of twin studies, there is a greater understanding of that link between epigenetics and phenotype. Such studies on genetically identical organisms suggest that studying the effects of epigenetics on phenotypic outcomes can yield discrete molecular pathways and mechanisms [48]. Therefore, an exploration of the effect of contemporary epigenetic impacts on phenotype can be expected to integrate with deep evolutionary experiences along the same types of molecular pathways that have been outlined above.

Certainly contemporary impacts can have substantial phenotypic results. In humans, maternal body mass index and blood pressure directly correlate with fetal birth weights [49]. Neonates born to obese women are larger and at a higher risk of birth complications. A similar association exists for elevated maternal fasting blood glucose, whereas elevated maternal systolic blood pressure has been directly linked to low birth weight infants. Starvation is now known to have profound intergenerational effects on phenotype, fitness and health in many animals. In C. elegans, generations of progeny of starved animals demonstrate smaller size, diminished fecundity, smaller brood size, a greater number of male progeny, and an increased tolerance to heat [50]. These transgenerational starvation effects in C. elegans have been demonstrated to be due to small RNAs that persist for at least three generations [51]. Such effects are now acknowledged in humans with starvation-induced neonatal adiposity and an increased incidence of diabetes in progeny [52]. Gluckman and Hanson [53] include the periconceptional, fetal, and infant environments among those aspects of particular significance in the future incidence of adult human disease. They further stress the dependence of the mature

phenotype upon both any individual genome and its epigenome, which then, together and iteratively, influence subsequent responses to environmental stresses and disease incidence [54]. This emphasis on the reciprocating balance between the environment and phenotype via epigenetic intermediaries is a form of interrelating niche construction through which any particular organism receives feedback from the environment, is shaped by it in some manner, and then correspondingly affects the outward environment that it occupies.

Furthermore, the influence of epigenetic impacts on phenotype extends beyond those forms that are typically considered. The mammalian placenta represents an example of epigenetic interactions and their critical impact upon mammalian development that achieve complex phenotypic form. Mammalian placental development is partially dependent upon crucial reproductive protein expression that does not emanate from within any central genome. Instead, it is the product of early epigenetic impacts as a co-option of retroviral proteins. Such retro-elements are largely responsible for the formation of the placental syncytiotrophoblast [55]. The development of maternal immunosuppression enabling viviparity is itself critically dependent on proteins produced in response to accumulated retro elements as infectious epigenetic impacts on central genomes [56]. Furthermore, retrotransposon activity or suppression are now acknowledged as epigenetic mediators of phenotypic variation in mammals producing variations between genetically identical individuals [57]. Nor are such impacts of little consequence to genomic integrity since retrotransposons are the principal component of most eukaryotic genomes and alter the expression of a wide variety of genes in animals and plants [58]. Beyond these distant considerations, epigenetic impacts are actually now contemporaneously evident. The endogenization of HIV [59], Koala retrovirus [60], and the direct demonstration of heritable transmission of bacterial DNA [61] are crucial examples. Importantly, these transfers are emblematic of the self-same processes in which epiphenomena are either employed or withstood, within both the eukaryotic and prokaryotic realms [62]. Self-same processes is the descriptive term used to indicate that our evolutionary system is based upon cellular activities in which there are consistent adherencies to basic cellular principles. These basic mechanisms guide cellular interactions and reactions and are consistently reiterated at every scope and scale. In every circumstance, these physical and bioactive epiphenomena can now be appreciated as directing responses to environmental stress through reiterative means.

  1. Predictive Value of Phenotype as Epigenetic Agent

Based upon these considerations, niche construction, either as beaver dams or cities, and then even further as phenotypes, can be productively assessed as consequences of elemental cellular first principles and epigenetic underpinnings. When considered in this manner, even our protracted human infancy and childhood can now be understood as a necessary phase through which crucial environmental epigenetic marks are assimilated to foster human brain development.

With these clarifications, all phases of the life cycle can be understood as derivative of cellular needs and imperatives that determine the timing and expression of each developmental and life cycle stage of which, arguably, the endocrine system has primacy. Crucially though, the endocrine system itself is a cellular phenomenon that is its own summation of epigenetic marks and their differential activation, ever-dependent upon environmental stresses [63].

  1. Conclusions

In any typical Darwinian narrative, phenotype is an output of selection experienced through differential survival and reproductive success. However, heritable epiphenomena are now better understood. Therefore, it can be argued that epigenetic mechanisms are a primary means by which organisms evolve in matched reciprocation to environmental stresses best exemplified by niche construction. Phenotype can then be reappraised through a non-intuitive Bohmian shift [3] within biologic terms of co-existent implicates and explicates. Phenotype is a transient explicate upon which a series of epigenetic impacts gather, as a set of implicates, to be enacted according to

cellular imperatives. The obligatory return of eukaryotic multicellular organisms to the unicellular form becomes the critical phase for the settling of those implicates towards biological expression as phenotype. Thus, a renewed evolutionary narrative can be considered that centers upon the primacy of epigenetic inheritance within deeply rooted cellular mechanisms. In such circumstances, perpetual cellular imperatives determine our evolutionary course. Phenotype is no longer merely a result but is instead a means through which organisms explore the outward environment and its stresses. Those impacts are brought back to the eukaryotic unicell and then, upon reproductive elaboration, enable the reiterative extension of phenotype into the environment to experience a subsequent series of environmental impacts towards its next set of adjustments. It is this consistent reciprocation that shapes phenotype. When fully considered, this new concept becomes a novel and testable route towards further progress in evolutionary theory and biomedicine.


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Published with permission, courtesy Biology (Basel)

Citation:  Torday, J. S., & Miller, W. B. (2016). Phenotype as agent for epigenetic inheritance. Biology, 5(3), 30.

Man is Integral to Nature

By John S. Torday, MSc. PhD.  & William B. Miller, Jr, M.D.

“It is a country . . . with innumerable lakes and rapid streams, peopled with trout.”

                                                                                    —H.D. Thoreau, The Maine Woods

In the Beginning

The traditional perspective for physiology, as portrayed by Galen and Harvey, is like Lego Blocks, with one biochemical process linked to another until an entire biochemical structure is revealed. In contrast to that post facto narrative, a predictive approach can be asserted—there actually are founding first principles for physiology that originated in and emanate from the unicellular stage of life. Einstein’s insight to relativity theory emerged from a dream in which he traveled in tandem with a light beam, seeing it as an integral particle and wave. Similarly, viewing physiology as a continuum from unicellular to multicellular organisms provides fundamental insight to ontogeny and phylogeny as an integral whole, directly linking the external physical environment to the internal environment of physiology, and even extending beyond, to the metaphysical realm, bearing in mind that the calcium waves that mediate consciousness in paramecia and in our brains are one and the same mechanism.

Life probably began much like the sea foam that can be found on any shoreline, since similar lipids naturally form primitive “cells” when vigorously agitated in water. Algae, for example, are 90 percent lipid. Such primitive cells provided a protected space for catalytic reactions that decreased and stabilized the internal energy state within the cell, and from which life could emerge. Crucially, that cellular space permits the circumvention of the second law of thermodynamics. (The entropy of an isolated system such as a cell never decreases since such systems always decay toward thermodynamic equilibrium as a state of maximum entropy.) That violation of physical law is the essential property of life as self-organizing and self-perpetuating, always in flux, staying apace with, and yet continually separable from a stressful, ever-changing external environment.

Even from the inception of life, rising calcium levels in the ocean have driven a perpetual balancing selection for calcium homeostasis, mediated by lipid metabolism. Metaphorically, the Greeks called it Ouroboros, an ancient symbol depicting a serpent eating its own tail.

Ouoroboros, an ancient symbol depicting a serpent eating its own tail. (Image by Abake, Wikimedia Commons).
The Ouroboros embodies self-reflexivity or cyclicity, especially in the sense of something constantly re-creating itself. Just like the mythological Phoenix, it operates in cycles that begin anew as soon as they end. Critically, the basic cell permits the internalization of factors in the environment that would otherwise have destroyed it—oxygen, minerals, heavy metals, micro-gravitational effects, and even bacteria—all facilitated by an internal membrane system that compartmentalized those factors within the cell to make them useful. These membrane interfaces are the biologic imperative that separates life from non-life—“Good fences make good neighbors.”

The Advent of Multicellularity

Unicellular organisms dominated the earth for the first 4.5 billion years of its existence. Far from static, these organisms were constantly adapting. From them, the simplest plants evolved first, producing oxygen and carbon dioxide that modified the nitrogen-filled atmosphere. The rising levels of atmospheric carbon dioxide, largely generated by blue-green algae, acidified the oceans by forming carbonic acid, progressively leaching more and more calcium from rock into the ocean waters, eventually forcing a proliferation of life from sea to land.

The existence of a protected space within primitive “cells” allowed for the formation of the endomembrane system, giving rise to chemiosmosis, or the generation of bioenergy through the partitioning of ions within the cell, much like a battery. Early in this progression, the otherwise toxic ambient calcium concentrations within primitive cells had to be lowered by forming calcium channels, composed of lipids embedded within the cell membrane, and the complementary formation of the endoplasmic reticulum, an internal membrane system for the compartmentalization of intracellular calcium. Ultimately, the advent of cholesterol synthesis facilitated the incorporation of cholesterol into the cell membrane of eukaryotes, differentiating them (our ancestors) from prokaryotes (bacteria), which are devoid of cholesterol. This process was contingent on an enriched oxygen atmosphere, since it takes six oxygen molecules to synthesize one cholesterol molecule. The cholesterol-containing cell membrane thinned out, critically increasing oxygen transport, enhancing motility through increased cytoplasmic streaming, and was also conducive to endocytosis, or cell eating.

All of these processes are the primary characteristics of vertebrate evolution. At some point in this progression of cellular complexity, impelled by oxygen-promoting metabolic drive, the evolving physiologic load on the system resulted in endoplasmic reticulum stress, periodically causing the release of toxic calcium into the cytoplasm of the cell. The counterbalancing, or epistatic mechanism, was the “invention” of the peroxisome, an organelle that utilizes lipids to buffer excess calcium. That mechanism became homeostatically fixed, further promoting the movement of ions into and out of the cell. Importantly, the internalization of the external environment by this mechanism reciprocally conveyed functional biologic information about the external surroundings, and promoted intracellular communication—what Claude Bernard referred to as the “internal milieu.”

Walter B. Cannon later formulated the concept that biological systems are designed to “trigger physiological responses to maintain the constancy of the internal environment in face of disturbances of external surroundings,” which he termed homeostasis. He emphasized the need for reassembling the data being amassed for the components of biological systems into the context of whole organism function. Hence, in 1991, Weibel, Taylor, and Bolis tested their theory of “symmorphosis,” the idea that physiology has evolved to optimize the economy of biologic function; interestingly, the one exception to this theory was the lung, which they discovered was “over-engineered,” but more about that later.

Harold Morowitz is a proponent of the concept that the energy that flows through a system also helps organize that system. West, Brown, and Enquist have derived a general model for allometry (the study of the relationship of body size to shape, anatomy, physiology, and behavior). They proposed a mathematical model demonstrating that metabolism complies with the 3/4 power law for metabolic rates (i.e., the rate of energy use in mammals increases with mass with a 3/4 exponent). Back in 1945, Horowitz hypothesized that all of biochemistry could be reduced to hierarchical networks, or “shells.” Based on these decades of study, investigators acknowledge that there are fundamental rules of physiology, but they do not address how and why these rules have evolved.

As eukaryotes thrived, they experienced increasing pressure for metabolic efficiency in competition with their prokaryotic cousins. They ingested bacteria via endocytosis, which were assimilated as mitochondria, providing more bioenergy to the cell for homeostasis. Eventually, eukaryotic metabolic cooperativity between cells gave rise to multicellular organisms, which were effectively able to compete with prokaryotes. As Simon Conway Morris has archly noted, “First there were bacteria, now there is New York.” Bacteria can act like multicellular organisms through such behavioral traits as quorum sensing and through biofilm formation, thus behaving, even at this primitive stage, as a pseudo-multicellular organism. The subsequent counter-balancing selection evolution of cellular growth factors and their signal-mediating receptors in our vertebrate ancestors facilitated cell–cell signaling, forming the basis for metazoan evolution. It is this same process that is recapitulated each time the organism undergoes embryogenesis.

This cellular focus on the process of evolution serves a number of purposes. First, it regards the mechanism of evolution from its unicellular origins as the epitome of the integrated genotype and phenotype. This provides a means of thinking about how and why multicellular organisms evolved, starting with the unicellular cell membrane as the common origin for all evolved complex traits. Further, it offers a discrete direction for experimentally determining the constituents of evolution based on the ontogeny and phylogeny of cellular processes. For example, it is commonplace for evolution scientists to emphasize the fact that any given evolved trait had its antecedents in an earlier phylogenetic species as a pre-adapted, or exapted, trait. These ancestral traits can then subsequently be cobbled together to form a novel structure and/or function. Inescapably, if followed to its logical conclusion, all metazoan traits must have evolved from their unicellular origins.

Evolution, Cellular-Style

Moving forward in biologic space and time, how might such complex traits have come about? Physiologic stress must have been the primary force behind such a generative process, transduced by changes in the homeostatic control mechanisms of cellular communication. When physiologic stress occurs in any complex organism, it increases blood pressure, causing vascular wall shear stress, particularly in the microvascular beds of visceral organs. Such shear stress generates reactive oxygen species (ROS), specifically at points of greatest vascular wall friction. ROS are known to damage DNA, RNA, and proteins, and to particularly do so at those sites most affected by the prevailing stress. This can result in context-specific gene mutations, and even gene duplications, all of which can profoundly affect the process of evolution. So we should bear in mind that such genetic changes are occurring within the integrated structural-functional context of that tissue and organ. However, understanding the biochemical processes undergirding the genetic ones equips a profound and testable mechanism for understanding the entire aggregate of genetic changes as both modifications of prior genetic lineages, and yet “fit enough to survive” in their new form.

Over evolutionary time, such varying modifications of structure and function would iteratively have altered various internal organs. These divergences would either successfully conform to the conditions at hand, or failing to do so, cause yet another round of damage-repair. Either an existential solution was found or the organism became extinct; either way, such physiologic changes would have translated into both phylogenetic and ontogenetic evolution. Such an evolutionary process need not be unidirectional. In the forward direction, developmental mechanisms recapitulate phylogenetic structures and functions, culminating in homeostatically controlled processes. And in the reverse direction, the best illustration lies with the genetic changes that occur under conditions of chronic disease, usually characterized by simplification of structure and function. For example, all scarring mechanisms are typified by fibroblastic reversion to their primordial signaling pathway. This sustains the integrity of the tissue or organ through the formation of scar tissue, albeit sub-optimally, yet allowing the organism to reproduce before being overwhelmed by the ongoing injury repair.

The Water-Land Transition and Vertebrate Evolution

Nowhere are such mechanisms of molecular evolution more evident than during the water-land transition. Rises in oxygen and carbon dioxide in the Phanerozoic atmosphere over the course of the last 500 million years partially dried up the oceans, lakes, and rivers, forcing organisms to adapt to land through remodeling of tissues and organs, or else become extinct. There were two known gene duplications that occurred during this period of terrestrial adaptation—the parathyroid hormone-related protein (PTHrP) receptor and the β adrenergic receptor (βAR). The cause of these gene duplications can be surmised from their effects on vertebrate physiology. PTHrP is necessary for a variety of traits relevant to land adaptation—ossification of bone, skin barrier development, and the formation of alveoli in the lung. Bone had to ossify to maintain the integrity of skeletal elements under the stress of higher gravitational forces on land compared to relative buoyancy in water. PTHrP signaling is necessary for calcium incorporation into bone. We know from the fossil record that there were at least five attempts to breach land by fish ancestors based on fossilized skeletal remains. Those events would have been accompanied by the evolution of visceral organs, based on both a priori reasoning, and the fact that the genes involved in skeletal development are also integral to the morphogenesis of critical internal organs, particularly PTHrP. In the aggregate, the net effect of shear stress on PTHrP-expressing organs like bone, lung, skin, and kidney may have precipitated the duplication of the PTHrP receptor, leaving those progeny best able to adapt to land. These, then, were the founders of the subsequent terrestrial species.

As a result of such positive selection pressure for PTHrP signaling, its genetic expression also evolved in both the pituitary and adrenal cortex, further stimulating adrenocorticotrophic hormone and corticoids, respectively, in response to the stress of land adaptation. This pituitary-adrenal cascade would have amplified the production of adrenaline in the adrenal medulla, since corticoids produced in the adrenal cortex pass through the microvascular arcades of the medulla on their way to the systemic bloodstream. This passage of corticoids through the medullary labyrinth enzymatically stimulates the rate-limiting step in adrenaline synthesis, catechol-O-methyltransferase, or COMT. Positive selection pressure for this functional trait may have resulted from cyclic periods of hypoxic stress. Episodes of intermittent large increases and decreases in atmospheric oxygen over geologic time, known as the Berner Hypothesis, may have triggered lapses in the capacity of the lung to oxygenate efficiently, demanding alternating antagonistic adaptations to hyperoxia and hypoxia as a result. The periodic increases in oxygen gave rise to increased body size, whereas hypoxia is the most potent vertebrate physiologic stressor known. Such intermittent periods of pulmonary insufficiency would have been alleviated by the increased adrenaline production, stimulating lung alveolar surfactant secretion, transiently increasing gas exchange by facilitating the distension of the existing alveoli. The increased distention of the alveoli, in turn, would have fostered the generation of more alveoli by stimulating stretch-regulated PTHrP secretion, which is both mitogenic for alveolarization, and angiogenic for the alveolar capillary bed. This would allow for iterative evolution of the alveolar bed in the interim through positive selection pressure for those members of the species most capable of increasing their PTHrP secretion.

And it is worthwhile highlighting the fact that the increased amounts of PTHrP flowing through the adrenal may also have been responsible for the evolution of the capillary system of the medulla. Such pleiotropic effects typify the positive selection that has occurred during the evolutionary process.

This scenario would also have explained the duplication of the βARs. The increase in their density within the alveolar capillary bed was necessary for relieving a major constraint during the evolution of the lung in adaptation to land. The βARs are required for a ubiquitous mechanism for blood pressure control in both the lung alveoli and the systemic blood pressure. The pulmonary system has a limited ability to withstand the swings in blood pressure to which other visceral organs are subjected. PTHrP is a potent vasodilator, so it had the capacity to compensate for the blood pressure constraint in the interim. But eventually the βARs evolved to coordinately accommodate both the systemic and local blood pressure control within the alveolar space.

The glucocorticoid (GC) receptor evolved from the mineralocorticoid (MC) receptor during this same period through a third gene duplication. Since blood pressure would have tended to increase during the vertebrate adaptation to land in response to gravitational demands, there would have been positive selection pressure to reduce the vascular stress caused by the blood pressure stimulation by the MC aldosterone during this phase of land vertebrate evolution. The evolution of the GC receptor would have placed positive selection on GC regulation by reducing the hypertensive effect of the MCs by diverting steroidogenesis toward cortisol production. In turn, the positive selection for the GC cortisol would have stimulated βAR expression, potentially explaining how and why the βARs superseded the blood pressure–reducing function of PTHrP. It is these ad hocexistential interactions that promoted land adaptation through independent local blood pressure regulation within the alveolus. This integration of blood pressure control in the lung and periphery by catecholamines represents allostatic evolution.

The net result of PTHrP-mediated pituitary-adrenal corticoid production would have fostered a more potent “fight or flight” mechanism in our amniote ancestors. These were small, shrew-like organisms that would have been advantaged by such a mechanism, making them “friskier,” able to more likely survive the onslaught of predators during that turbulent era.

Moreover, increased episodes of adrenaline production in response to stress may have fostered the evolution of the central nervous system. Peripheral adrenaline mediates and limits blood flow through the blood-brain barrier, which would be expected to cause increased adrenaline and noradrenaline production within the evolving brain. Both adrenaline and noradrenaline promote neuron development. One might even speculate that this cascade led to human creativity and problem solving as an evolved expression of that same axis as an alternative to fight or flight, since it is well known that learning requires stress.

The duplication of the βAR gene may also have been instigated by the same intermittent cyclical hypoxia resulting from the process of lung adaptation, subsequently facilitating independent blood pressure regulation within the alveolar microvasculature; both of these mechanisms would have been synergized by the evolution of the GCs during this transition.

The bottom line is that all of the molecular pathways that evolved in service to the water-land transition—the PTHrP Receptor, the βAR, and the GC Receptor—aided and abetted the evolution of the vertebrate lung, the rate-limiting step in land adaptation. Perhaps that is why Weibel, Taylor, and Bolis observed that the lung had more physiologic capacity than was necessary for its normal range of function (see above), since only those organisms fit to amplify their PTHrP expression survived the stress of the water-land transition. The synergistic interactions of the lung and pituitary-adrenal axis producing adrenaline relieved the constraint on the lung through increased PTHrP production, fostering more alveoli. Perhaps this is the reason why the lung has excess capacity—either that, or become extinct.

The Cellular Approach to Evolution Is Predictive

This reduction of the process of evolution to cell biology has an important scientific feature—it is predictive. For example, it may answer the perennially unsolved question as to why organisms return to their unicellular origins during their life cycles. Perhaps, as Samuel Butler surmised, “a hen is just an egg’s way of making another egg” after all. It is worth considering the proposal that since all complex organisms originated from the unicellular state, a return to the unicellular state is necessary in order to ensure the fidelity of any given mutation with all of the subsequently evolved homeostatic mechanisms, from its origins during phylogeny through all the elaborating permutations and mutational combinations of that trait during the process of evolution. One way of thinking about this concept is to consider that perhaps Haeckel’s biogenetic law is correct after all—that ontogeny actually does recapitulate phylogeny. His theory has been dismissed for lack of evidence for intermediary steps in phylogeny occurring during embryonic development, like gill slits and tails. However, that was during an era when the cellular-molecular mechanisms of development were unknown. A testament to the existence of such molecular lapses is the term “ghost lineage,” which fills such gaps in the fossil record with euphemisms. We now know that there are such cellular-molecular physiologic changes over evolutionary time that are not expressed in bone, but are equally as important, if not more so in other organ systems. In all likelihood, ontogeny must recapitulate phylogeny in order to vouchsafe the integrity of all of the homeostatic mechanisms that each and every gene supports in facilitating evolutionary development. Without such a fail-safe mechanism for the foundational principles of life, there would be inevitable drift away from our first principles, putting the core process of evolution in response to environmental changes itself at risk of extinction. S.J. Gould famously wondered whether an evolutionary “tape” replayed would recapitulate. In this construct, the answer would resoundingly be yes, at least qualitatively, since all of the same components—bacteria, oxygen, minerals, heavy metals—are still present, and it would be expected that first principles would still remain as they are.

One implication of this perspective on evolution—starting from the unicellular state phylogenetically, being recapitulated ontogenetically—is the likelihood that it is the unicellular state that is actually the object of selection. The multicellular state—which Gould and Lewontin called “Spandrels”—is merely a biologic probe for monitoring the environment between unicellular stages in order to register and facilitate adaptive changes. This consideration can be based on both a priori and empiric data. Regarding the former, emerging evidence for epigenetic inheritance demonstrates that the environment can cause heritable changes in the genome, but they only take effect phenotypically in successive generations. This would suggest that it is actually the germ cells of the offspring that are being selected for. The starvation model of metabolic syndrome may illustrate this experimentally. Maternal diet can cause obesity, hypertension, and diabetes in the offspring. But they also mature sexually at an earlier stage due to the excess amount of body fat. Though seemingly incongruous, this may represent the primary strategy to accelerate the genetic transfer of information to the next generation (positive selection), effectively overarching the expected paucity of food. The concomitant obesity, hypertension, and diabetes are unfortunate side effects of this otherwise adaptive process in the adults. Under these circumstances, one can surmise that it is the germ cells that are being selected for; in other words, the adults are disposable, as Dawkins has opined.

Hologenomic evolution theory provides yet another mechanism for selection emerging from the unicellular state. According to that theory, all complex organisms actually represent a vast collaborative of linked, co-dependent, cooperative, and competitive localized environments and ecologies functioning as a unitary organism toward the external environment. These co-linked ecologies are comprised of both the innate cells of that organism and all of the microbial life that is cohabitant with it. The singular function of these ecologies is to maintain the homeostatic preferences of their constituent cells. In this theory, evolutionary development is the further expression of cooperation, competition, and connections between the cellular constituents in each of those linked ecologies in successive iterations as they successfully sustain themselves against a hostile external genetic environment. Ontogeny would then recapitulate phylogeny since the integrity of the linked environments that constitute a fully developed organism can only be maintained by reiterating those environmental ecologies in succession towards their full expression in the organism as a whole.

Another way to think about the notion of the unicellular state as the one being selected for is to focus on calcium signaling as the initiating event for all of biology. There is experimental evidence that increases in carbon dioxide during the Phanerozoic era caused acidification of the oceans, causing leaching of calcium from the ocean floor. The rise in calcium levels can be causally linked to the evolution of the biota and is intimately involved with nearly all biologic processes. For example, fertilization of the ovum by sperm induces a wave of calcium, which triggers embryogenesis. The same sorts of processes continue throughout the life cycle, until the organism dies. There seems to be a disproportionate investment in the zygote from a purely biologic perspective. However, given the prevalence of calcium signaling at every stage, on the one hand, and the participation of the gonadocytes in epigenetic inheritance on the other, the reality of the vectorial trajectory of the life cycle becomes apparent—it cannot be static, it must move either toward or away from change.

By using the cellular-molecular ontogenetic and phylogenetic approach described above for the water-land transition as a major impetus for evolution, a similar approach can be used moving both forward and backward from that critically important phase of vertebrate evolution. In so doing, the gaps between unicellular and multicellular genotypes and phenotypes can realistically be filled in systematically. But we should bear in mind that until experimentation is done, these linkages remain hypothetical. Importantly, though, there are now model organisms and molecular tools to test these hypotheses, finally looking at evolution in the direction in which it occurred, from the earliest iteration forward. This approach will yield a priori knowledge about the first principles of physiology and how they have evolved to generate form and function from their unicellular origins.

We Are Not Just in This Environment, We Are of It

The realization that there are first principles in physiology as predicted by the cellular-molecular approach to evolution is important because of its impact on how we think of ourselves as individual humans and as a species, and on our relationship to other species. Once we recognize and understood that we, as our own unique species, have evolved from unicellular organisms, and that this is the case for all of the other organisms on earth, including plant life, the intense and intimate interrelationships among all of us must be embraced. This kind of thinking has previously been considered in the form of genes that are common to plants and animals alike, but not as part of a larger and even more elemental process of evolution from the physical firmament. This perspective is on par with the reorientation of man to his surroundings once he acknowledged that the sun, not the earth, was the center of the solar system. That shift in thought gave rise to the Age of Enlightenment! Perhaps in our present age, such a frame-shift will provide insight into black matter, string theory, and multiverses.

In retrospect, it should have come as no surprise that we have misapprehended our own physiology. Many discoveries in biomedicine are serendipitous, medicine is post-dictive, and the Human Genome Project has not yet yielded any of its predicted breakthroughs. However, moving forward, knowing what we now do, we should countenance our own existence as part of the wider environment—that we are not merely in this world, but literally of this world—with an intimacy that we had never previously imagined.

This unicellular-centric vantage point is heretical, but like the shift from geocentrism to heliocentrism, our species would be vastly improved by recognizing this persistent, systematic error in self-perception. We are not the pinnacle of biologic existence, and we would be better stewards of the land and our planet if we realized it. We have learned that we must share resources with all of our biological relatives. Perhaps through a fundamental, scientifically testable and demonstrable understanding of what we are and how we came to be so, more of us will behave more consistently with nature’s needs instead of subordinating them to our own narcissistic whims. As we become deeply aware of our true place in the biologic realm, such as we are already witnessing through our increasing recognition of an immense microbial array as fellow travelers with us as our microbiome, we may find a more ecumenical approach to life than we have been practicing for the last five thousand years.

Bioethics Based on Evolutionary Ontology and Epistemology, Not Descriptive Phenotypes and Genes

By definition, a fundamental change in the way we perceive ourselves as a species would demand a commensurate change in our ethical behavior. Such thoughts are reminiscent of a comment in a recent profile of the British philosopher Derek Parfit in The New Yorker magazine, entitled “How to be Good,” in which he puzzles over the inherent paradox between empathy and Darwinian survival of the fittest. These two concepts would seem to be irreconcilable, yet that is only because the latter is based on a false premise. Darwin’s great success was in making the subject of evolution user-friendly by providing a narrative that was simple and direct. Pleasing as it may be, it is at best entirely incomplete. Think of it like the transition from Newtonian mechanics to relativity theory. As much is learned about the unicellular world with its surprising mechanisms and capacities, new pathways must be imagined. It is clear that we as humans are hologenomes, and all the other complex creatures are, too. In fact, there are no exceptions. The reasons for this can only be understood properly through a journey from the “Big Bang” of the cell forward, with all its faculties and strictures. By concentrating on cellular dynamics, an entirely coherent path is empowered. Tennyson’s line about “Nature, red in tooth and claw” is only the tip of what the iceberg of evolution really constitutes. As pointed out above, we evolved from unicellular organisms through cooperation, co-dependence, collaboration, and competition. These are all archetypical cellular capacities. Would we not then ourselves, as an example of cellular reiteration, have just those self-same and self-similar behaviors?


In summary, by looking at the process of evolution from its unicellular origins, the causal relationships between genotype and phenotype are revealed, as are many other aspects of biology and medicine that have remained anecdotal and counter-intuitive. That is because the prevailing descriptive, top-down portrayal of physiology under Darwinism is tautological. In opposition to that, the cellular-molecular, bottom-up approach is conducive to prediction, which is the most powerful test of any scientific concept. Though there is not a great deal of experimental evidence for the intermediate steps between unicellular and multicellular organisms compared to what is known of ontogeny and phylogeny of metazoans, we hope that the perspectives expressed in this essay will encourage more such fundamental physiologic experimentation in the future. 

Pre-Biotic Evolution: III. Transitioning to Animacy

Joseph H. Guth

Published by the

Society for the Advancement of Metadarwinism, Volume 3 2015

One Scientist’s Overview and Perspectives


In the first two parts of this series1,2, we have attempted to describe and summarize the steps and processes that in the author’s view, provide an almost invariably likely pathway from the production and evolution of various atomic species, formation of huge varying complex chemical mixtures under early earth conditions, and predictable interactions that then could lead to a high probability for the self generation and self-selection of complex dynamic, chemically-reactive systems and processes that ultimately we would come to define as organic-based terrestrial life.  This may not be the only such pathway but is argued to be a logical, consistent and realistic one that merits serious consideration.  Most would consider the ultimate occurrence of such a completely integrated process as highly unlikely, as one of very low probability.  That would be true if the universe only worked with one pair of molecules at a time and in a linear, sequential, trial-and-error fashion.  But if these molecular “experiments” occurred in parallel and simultaneously, and in uncountable high numbers of combinations and permutations for such events, as occurs in combinatorial chemistry, but which extended over very long timeframes while under a wide variety of operating conditions, the unlikely could then become the inevitable.  This is not unlike the old posit of having an infinite number of monkeys typing on an infinite number of typewriters eventually by random chance and accident being able to create all of Shakespeare’s novels.

Up through this point, we have collected a set of basic chemical and physical facts and allowed them to be blended into a plausible story of genesis of the first set of chemical species and energy sources that are thought by many to exist throughout the universe.  It provides a certain compelling argument for expecting similar living processes and organisms being likely to have developed throughout the cosmos.  But it also provides us with a more specific set of targets and signs to scan and search for other life in our extra-terrestrial explorations now and into the far future.

Animacy, one of life’s most useful characteristics when it occurs, is the ability to engage in some kind of movement, relocation or translocation.  Such movements are not always present or apparent at the individual organismal level but can usually be found if not individually, then collectively in colony-forming species or multicellular organisms.  Even rooted, as well as non-rooted single-celled or colony-forming plants, have the ability to move or reorient themselves in response to their environmental needs (e. g., phototaxis, water seeking tap root growth, gravitaxis, specialized food-trapping structures and flagellated motion).  It is hard to overstate how great an evolutionary advantage was gained when living systems of all types finally were able to react to environmental conditions and the presence of other living organisms by development and self-selecting for directed translocation.

From Molecules in Random Motion to Functional Movements in Evolving Protocells

Motion, reorientation or other movement can be found in different functional modalities in modern day living systems.  For instance, the movement  of the outer eukaryotic cell membrane as it pinches off at the end of the mitotic cycle performs a different function than the pinching off of membrane vesicles during phago-, endo- and exo-cytosis.  Such membrane-based movements are further expanded on with the ruffling leading edge of the plasma membrane in certain types of amoeboid movement.

Figure 1.  Both mammalian cells are demonstrating membrane edge ruffling behavior.  In this animated gif image file, the cell on the left is stationary because the entire 360 degree perimeter is involved.  The cell on the right is rapidly moving from left to right with the leading hyaline edge on its right side ruffling strongly.  In such directional motility, the cell’s underlying cytoskeleton plays a major role in its ability to keep it moving in a continuous fashion in one direction.  (Taken from  Accessed 12/2/2015)

Of course many of these modern day capabilities are also greatly assisted by underlying intracellular structures and molecular processes3 such as microfilament-microtubule interactions4 and the concurrent visco-elasticity changes through reversible cytoplasmic gel-sol modification, the internal relocation of mitochondria and increased rates of intra-cytoplasmic streaming and mixing of chemicals and enzyme complexes (Figure 2).

Figure 2.  Cytoplasmic flow generated by microfilaments and microtubule polymerization-depolymerization cycles is seen in this animated image as a mechanism for internal cytoplasmic mixing, relocation of subcellular organelles and for generating the underlying forces for formation of pseudopodia, ruffling edges and other mechanical movements by the cell.  The efficiency of such an evolutionary development represents a highly efficient use of structure and function for the early cells. (From Dr. R. Wagner,, accessed 12/6/2015)

But there has been a well-demonstrated ability by various purified synthetic lipid membrane model systems preparations to also spontaneously possess very similar behavior and capabilities.5  Figure 3 provides a still record of this behavior but microscopic videos have recorded it as a dynamic budding process that is sometimes surprisingly rapid.  So the first protocells composed of these chemical compounds simply incorporated the natural tendencies of the self-organized chemicals they were made up of.  It was simple, undirected, relatively low-functional movement, but movement and change, nevertheless.  And from any kind of change comes the opportunity to evolve into something more adapted to the existing conditions.

Figure 3.  Spontaneous microsphere formation, pinching off and fusion behavior occurs in synthetic preparations and is intrinsic to both lipids, like lecithin, and to proteins with higher contents of less polar amino acid compositions.  (Taken from, accessed 12/6/2015)

Thus we likely actually have the initial seed for cellular mobility and animation actually built into the molecular properties of the membrane matrix.  It can be enhanced with other underlying molecular machinery, such as microfilament and microtubule structural and dynamically-changing mechanical frameworks, but the primary basics for animacy are built into the physical properties of the lipids, and to a narrower extent, the membrane proteins and carbohydrate-containing molecules that compose such structures in later protocells as well as up through modern cells.  For more highly developed mechanisms and structure emerging in cell evolution, such as flagella and cilia, we will have to wait until we discuss the developments after the first prokaryotic and eukaryotic cells transitioned into existence in a later installment.

With the intrinsic capability for limited movement already inherent or built-in at the molecular level in protocellular membranes, directed movement could provide a dramatic improvement in survival potential for more successfully composed protocells.  For instance, the ability to avoid damaging conditions, such as excessively high or low temperatures (thermotaxis), or seek out better sources of nutrients or avoiding toxic substances (chemotaxis), or find optimal levels of illumination (phototaxis) would be indispensable for enhancing the growth rate, reproduction rate, and survival potential of protocells as it does in modern living cells.  In this sense, one might extend the idea of Darwinism and survival-of-the-fittest to the basic animation processes deriving from the molecular properties of various molecules making up the cell membranes.

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Figure 4.  An animated image providing an example of a directed chemotactic response by a white blood cell that is tracking and phagocytizing bacteria in blood.  (Taken from and attributed to David Rogers, Vanderbilt University.  Accessed 12/2/2015)

This capability is well-recognized to confer a significant modern advantage to present day life as exemplified in the above figure, but such a capability would have profound advantages to a simpler protocell that was only getting its initial nutrition through soluble compounds.  Such dissolved nutrients were rate-limited by slow passive diffusion rates through the surrounding medium.  Non-soluble sources of nutrients would have been unavailable to such passive cells.

Membrane Basics and Molecular Bases for Membrane Dynamic Behaviors

Many things can modify those animation-related properties of modern cell membranes.  Such membranes all have a basic common design.  The major framework is composed of various phospholipids that preferentially auto-assemble into two-dimensional sheets in aqueous media in the temperature range allowing fluid behavior.  One of the most abundant membrane phospholipids, lecithin, is composed of three bonded components.  Those are a phosphate moiety, glycerol, and one or two fatty acid molecules (see Figure 5).

Figure 5.  The most common molecule that provides the basic framework for modern day cell membranes is the phospholipid, lecithin.  It is made of a phosphate group (blue ball) covalently bonded to a glycerol molecule to which two fatty acid molecules (elongated downward legs) are also attached.  This diagram also demonstrates the saturated fatty acid molecule on the left is flexible but can become linear while the fatty acid molecule on the right has a permanent kink in it due to its being unsaturated (that is, containing a double bond between two adjacent carbon atoms in it).  Not all phospholipid molecules contain the same fatty acids nor the same proportions of saturated to unsaturated fatty acids.  The lengths of the fatty acid chains and the number of unsaturated bonds within the unsaturated fatty acids also modify the temperature at which that part of the membrane transitions between its more fluid to less fluid state, and thus its deformability and fusibility in that same region.  (Taken from Wikipedia, “Phospholipids”. Image authored by Ties van Brussel .  Accessed 11/30/2015).

The elongated phospholipid molecules always have a highly polar end, that is attracted to other polar molecules such as water, and a low- or non-polar end that is not bondable with polar molecules.  Those hydrophobic ends tend to become more stably associated with other non-polar molecular regions.  When these phospholipid molecules finally finish their segregating and reorientation in a bulk water phase, they initially can form a monomolecular layer at the air-water surface interface, with the polar phosphate ends bonding downwards with water molecules and their non-polar, hydrophobic ends pointing upwards towards the less polar molecules making up air.  In this “head down” orientation, the fatty acid hydrophobic tails are stabilized by lower strength van der Waal forces.  But that is still sufficient to hold them in a closely packed two dimensional sheet.  But in any average suspension of such phospholipids in aqueous media, these monolayers will tend to spontaneously form bilayers in which two monolayers pair up with their hydrophobic fatty acid tails directed inwards towards each other and their more polar, electrically-charged phosphate heads oriented outwards and in intimate contact with the water phase.  Water molecules are polar and can hydrogen bond with each other as well as other, more polar molecules.  Because of the stronger electrical charge fields existing around the phosphate moiety, water molecules tend to strongly interact with this end of the phospholipids and arrange themselves in a loose cage-like structure surrounding each phosphate group.  This overall electrical charge repulsive force and clathrate structure interaction is what generates the natural but slight rigidity in a molecular aggregate that is only two molecules thick!  It generates a physical back-resistance to membrane deformation that allows there to be different flexibility between different zones of the membrane.  This is energetically a more stable arrangement for a sheet-like structure.

In an aqueous phase there are edges where phosphate groups are absent that are exposed to the more polar aqueous environment and those seek to lower their energy state even more by minimizing the exposed internal hydrophobic interior.  To minimize this disruption to the water structure surrounding the flat sheet of bilayer membrane, the membrane edges continue the self association process ultimately resulting in an even lower energy arrangement, a closed vesicle shape with no exposed edges.  Thus the simplest bilayer membrane geometry that is most stable at normal temperatures is the single bilayer, membrane-enclosed vesicle.  Studies have demonstrated that multiple- layered bilayers are also capable of forming.  These are not as rapidly permeable as a single bilayer under normal circumstances so it is unlikely that such added barriers between the internal life chemistry and the external medium would confer greater advantages to the newly created protocells.  Though we will revisit this thought in a later installment.

Analytical and synthetic chemists have a rule of thumb that is based upon this difference in solubility based on the polarity and charges of the molecule’s bonds.  Termed the Solubility Rule, it is well known and stated that “like dissolves like”.  What this means is that when looking at a molecule’s structure, one can make a fairly accurate prediction of what kinds of solvents would best be useful to dissolve such a compound in. Many kinds of chromatographic and affinity separations of complex mixtures use this relationship as well.  It is based upon the thermodynamic properties and bond strengths of intermolecular bonding and rearranging the positioning and orientation of electrically charged and uncharged molecules (or portions thereof) with respect to one another.  Its applicability also extends one level further in that when dealing with two or more types of molecules within a single category, such as hydrophobic molecules, that molecules with identical structure tend to further segregate themselves into regions with others of the same structure.  Such pooled regions of identical composition in a larger background of molecules of similar low charge and polarities is essentially what is found in modern day biological membrane structure.  This has also been termed membrane microenvironments.  A good example is the “raft-like” behavior of cholesterol and other steroidal molecules that cluster around embedded and transmembrane proteins within a broader area of bilayer phospholipid membrane (see figure 6).

Figure 6.  A cross-section of a modern animal cell plasma membrane illustrating the different microenvironment regions of such a membrane.   Phospholipid molecules showing the two fatty acid hydrophobic tails (light blue) attached to the hydrophilic heads (darker pink circles) align in two monolayers to form a bilayer framework within which many other molecules associate and function.  Region 1 is the microenvironments where no other molecules are present.  Region 2 is a raft region which is enriched in cholesterol content (number 7 in light pink) and membrane proteins (numbered 3 and 4 in dark green) as well as other kinds of functional molecules (numbered 5, 6 and 8).  The raft regions provide a more physically and chemically unique microenvironment within which membrane-bound, multi-protein complexes are attracted and operate.  Phase transition temperatures within these regions are expected to be somewhat different than those in region 1 zones.  Physical deformability and fusion/membrane coalescence capabilities would also be expected to be modified in those regions as well.  (Taken from  Wikipedia.  “Lipid Raft”.  Image authored by Artur Jan Fijałkowski.  Accessed 11/30/2015).

One purpose of such segregated regions within a phospholipid bilayer is to provide regions of differing solubility for other lower polarity transmembrane molecules, such as transmembrane proteins, to reside in.  Such transmembrane molecules can provide specialized transmembrane functions, such as semi-permeability, electrochemical reaction platforms, cell signalling, intracellular communication, and catalysis-assisted processing and transport of specific substances.  Obviously, the appearance and conservation of cholesterol-rich, self-associated membranes during protocell evolution would also be a major advantage to increasing the likelihood of more potential functions being able to be added to the developing protocell membrane in our evolutionary story of the perfecting of cells from protocells.

Under typical biologically favorable conditions, these phospholipids tend to aggregate together in different packing geometries and with different bonding energies defined, in part, by the kinds of fatty acids that are part of their composition.  Regularly-shaped saturated fatty acids tend to “crystallize” into more tightly packed arrangements at some pseudo-solidification temperatures while the presence of unsaturated bonds tends to keep them from crystallizing at those temperatures.  What this means is that bilayer membrane lipids will have regions in their 2 dimensional surfaces at any given temperature where the internal, more saturated fatty acids will be more tightly packed and less fluid.  Lower fluidity necessarily hinders molecular motions and transmembrane processes while at other, more unsaturated locations those transmembrane processes occur more quickly and with larger scale molecular motions.  In such higher fluidity regions of the membrane surface, the packed membrane components can quickly reconfigure to allow surface-localized transport to occur.

Other concurrent membrane properties that are affected by the rafting and saturated/unsaturated behavior of the lipid components are membrane flexibility, folding and fusibility/pinch-off potential.  Such are the physicochemical foundations for modifying the fluid/semi-fluid/nonfluid behavior potential of all membranes.  In studies using modern cell types, the fatty acid compositions of biological membranes actually changes in a single cell type as a function of the growth temperature.  This is an active homeostatic control mechanism of cells that is linked to their lipid metabolism.  The membrane fatty acid composition is continually readjusted so that the fluid transition temperature is maintained just below the actual growth temperature, assuring transmembrane processing will remain fully functioning.  Because these membrane lipids can have several different transition temperatures that can modify both the packing geometries of a single kind of lipid as well as the fluidity and flexibility of that part of the membrane it is found in, membranes are considered liquid crystalline in nature.  This provides for thermal transitions between several different phases and thus allows a wide variety of fluidity-based functions to co-exist within a single membrane at the same time at a given temperature.

Each transition temperature built into a particular membrane’s lipid framework confers both mechanical and transport limitation ranges as well as the optimum temperatures for each kind of membrane-based process or movement.  When one looks at cells that are undergoing some directed dynamic process or behavior, it is tempting to envision that such a large scale movement is caused by or connected to highly localized transmembrane molecular events underlying and inseparably-linked to the overall function being carried out.  As an example, if one views the formation and extrusion of pseudopodia in a modern day amoeba moving along a surface towards a nutrient-laden particle, it would be greatly aided in its food-seeking chemotaxis if the concentration of soluble nutrient substances diffusing from that particle be the first event that triggers the amoeboid cell movement in that direction.  This first event entails recognizing that a source of food is closely present and can be reached by moving in the direction closest to the point on its membrane where the greatest flux of soluble nutrients are passing through.  Areas of this closest approach would become enriched in more fluid-like lipids that concurrently would allow a greater degree of deformation at a given temperature while the other parts of the membrane are reduced in their fluidity.  They would take on a somewhat more deformation-resistant quality and give the cell a better way of maintaining its directionality of overall translocation or movement.  And embedded within the differing functional and structural micro-areas of this food-seeking cell’s membrane are other translocation molecules, like proteins, that are further enhancements to connect the overall membrane alterations with internal biochemical changes needed to capture and process the food source once it is physically encountered.  Not only higher fluxes of soluble nutrient molecules would be expected in this initiation of a chemotactic movement, but active triggering of internal cellular motility machinery through localized increases of calcium ion influxes would be coordinated at the same time.  Such localized increased calcium concentration just below the deforming membrane surface would also activate microfilament movement creating an internal current of cytoplasmic flow in that same region.  That flow would further deform the membrane in that location leading to a general cellular movement in that direction.

In protocells, we do not yet have that level of functionality and complexity built in but the co-presence of other lipids with differing transition temperatures and self-association energies can begin to provide such infant cells with a beginning blush of chemotactic potential.  Darwinian evolution must then follow to increase rates and improve performance of such a complex set of molecular events found in modern day cells.  One very basic aspect of cell animacy is likely to have sprung from the molecular structure and properties of the membrane lipids that are to be found in modern cells and likely in early earth protocells as well.

The Evolutionary Impact of Cell Membrane Fusion and Scission

Experiments with both phospholipids have demonstrated that when the temperature was above the phase transition temperature for membrane fluidity, the closed membrane vesicles were capable of two overall actions.5,6  The first was of a scission or budding process in which a vesicle could pinch off and separate into two or more smaller vesicles without loosing, leaking or diluting its internal fluid content or otherwise modifying that internal compartment chemistry noticeably (See Figure 7).  That is an extremely useful built-in behavior for such a packaging that needs to indefinitely maintain its complete integrity and uncompromised contents.Figure 7.  This greatly magnified phospholipid vesicle can exhibit bud formation and ultimate scission into smaller vesicles without any additional internal mechanical or chemical assistance. This example is indistinguishable from an identical behavior in modern organisms’ cell membranes.  (Taken from E. Sackmann, “Physical Basis of Self-Organization and Function of Membranes: Physics of Vesicles”, Chapter 5 in Handbook of Biological Physics, Vol. 1, R. Lipowsky and E. Sackmann, eds., Elsevier Science B. V., 1995)

Figure 7.  This greatly magnified phospholipid vesicle can exhibit bud formation and ultimate scission into smaller vesicles without any additional internal mechanical or chemical assistance. This example is indistinguishable from an identical behavior in modern organisms’ cell membranes.  (Taken from E. Sackmann, “Physical Basis of Self-Organization and Function of Membranes: Physics of Vesicles”, Chapter 5 in Handbook of Biological Physics, Vol. 1, R. Lipowsky and E. Sackmann, eds., Elsevier Science B. V., 1995)

The second action is that of the coalescence and fusion of two or more different vesicles into one larger vesicle, again without loosing or modifying their contents through external medium leakage into them.  The fact that such actions are naturally found to occur in most living cells provides us with a directly observable behavior of cells that is based upon the inherent molecular properties of phospholipid membrane composition, internal membrane component rules of association, and the overall tendency to form vesicular structures in aqueous environments.

What these two properties allowed the earliest protocells to accomplish was to provide a rather robust and putatively permanent housing for the complex chemical processes that were developing and occurring in the locations on early earth where the building blocks of life were being endlessly and spontaneously generated and modified.  These properties of membrane fusion and scission allowed the large pre-protocellular bulk liquid phases containing the various combinations of linked reaction sequences to not only be initially captured, but even more important, to give each micro-collection of complex reactive chemistry the ability to resist dissipation and to remain longer-lasting.  It provided the desirable internal environments to maintain the reactants in a more concentrated fashion within small protected environments.  With this increased longevity, these reactive vesicles could combine their contents together in a multitude of different trials-and-errors while conserving the higher internal concentrations of ingredients.  Such a design would improve the chances for development of even faster and more controlled internal chemical kinetics.

But another and even more valuable evolutionary capability was inherent in this protocellular membrane design.  The ability to become permanently self-sustaining through the spontaneous scission and generation of vesicular copies of more successful protocell compositions would be drastically improved if the more rapidly growing versions could bud or pinch off into independently growing units.  Multiple backup copies of successful versions would be a strategy to conserve a new and more suitable species of protocell.  And retaining a smaller vesicular size (i. e., surface-area-to-volume ratio) increases the transmembrane flow of metabolites into and out of the protocells without becoming limited by internal diffusion rates or increased thickness of membrane surface diffusion-limiting solvent boundaries.  The forces occurring in natural fluid settings that abound in nature are quite adequate to provide the energy and forces necessary to induce such scissions prior to the evolutionary development of enhanced molecular machinery specifically designed to carry out such gross modern cell membrane modifications as occurs in cell division (Figure 8a and 8b).

Figure 8a (top) and 8b (bottom).  Simple fluid cascades and waves breaking can provide the shearing forces to stretch and pinch off the membranes of larger vesicles and produce multiple copies of smaller versions of the parent version of vesicles without losing the original content compositions or internal chemistries.  It is argued here that these were the first forces available and involved in the earliest versions of protocell formation, division and even subsequent fusions of protocells initially possessing different internal constituents. (Unknown sources).

So with this rudimentary mechanism on early earth operating and which caused an increase in protocell numbers, this could be construed as a form of protocell division.  It is important to highlight at least one process that would have naturally occurred to show we have a likely physical process for protocell division.  Not only would it have been likely to occur anywhere in the world of early earth, it would have sped up the evolution of more successful versions of protocells.  Thus it would have increased their survival as well as rates of generation.

As long as nutrients are present and waste elimination is not limiting, each separated vesicle of a more successful composition would become another center of growth through mass accumulation.  This is a seminal feature of the application of both Chaos Theory and Complexity Theory.  This combination of advantages is all proposed to have occurred as a precursor to the development of a genetic code, genomic information recording and directed synthetic management  system (DNA, RNA and protein biosynthesis).  It provides a plausible mechanism for enhanced multiplying of the more successful combinations of protocellular compositions over those that were less successfully composed in a non-genetic manner through the Butterfly Effect.

Self-sealing, membrane-bounded vesicles can also fuse together, safely combining their protected internal contents into new admixtures.  This mechanism also provides a higher order potential of developing larger steps in the protocell design changes during the continuous march of trial-and-error experimentation that evolution rests upon.  Membrane fusion is also useful for the development of later capabilities of phagocytosis found in many kinds of modern day cells.  For instance, a cell engulfs a food particle and takes it into its cytoplasm where that membrane-bounded particle now fuses with lysosomes or other membrane-bounded organelles for further processing.  For protocellular survival, the ability to take in non-soluble structures, such as other protocell membrane fragments and less successful protocell versions, and reduce them to soluble nutrients would provide new sources of nutrition.  Without that property, insoluble protocell debris would continue to accumulate with no means to rapidly remove it from the niche.  This competition could quickly develop into a very early version of survival-of-the-fittest in a predator-prey relationship of protocell types.

Modern day cell membranes have many molecular “helpers” to accelerate various membrane-based mechanical movements and modifications.  But the intrinsic molecular properties of the phospholipid framework upon which it, as well as protocells, were built was already designed to allow such actions to occur given the right environmental conditions and an adequate, naturally-occurring energy source.  The later developments of pino-cytosis, endo-cytosis, exo-cytosis, phago-cytosis and syncytium formation all have highly developed machinery to aid and control these processes.

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Figure 9.  Dictyostelium slime mold amoeboid movements, chemotaxis-based association behavior and differing functional movements at both the individual cellular and multicellular organizational levels. (Produced by John Bonner, Princeton University.  Taken from public posting on YouTube at  Accessed 12/5/2015)

But none would have developed had not the phospholipid component of cell membranes already possessed, in a chemically built-in fashion, its very unique set of associative, disassociative, fusibility and localized variation of permeability properties.  And all attendant functions connected to those developments would also have suffered or been unsupportable and lost.  Such functions are paramount to the organization and operation in multi-cellular organisms and as such, may be necessary to attain before such larger collective and specialized multicellular organisms could evolve.

Animacy is a broad collection of evolutionarily useful potential mechanisms based upon the physicochemistry of protocell membrane lipids.  It is an almost incomprehensible testament to the persistence, protection, permanency and survival power of the membrane design that finally occurred in the first successful protocell.  That membrane, only two molecules in thickness, had to capture, retain and protect the living chemical processes operating within it in a continuous, never-ending sequence of growth, multiplication and interaction with the outside world for billions of years.  It had to have the plasticity to move through even the smallest of passages and instantly seal when any sharp physical insult threatened its integrity.  The complex chemistry leading up to the production of membrane-forming molecules is seemingly pre-destined to always form anywhere in our cosmos.  What a marvel!

Next:  Pre-Biotic Evolution.  Part IV.  The Development of Electrochemically-Generated Energy Linkage, Extraction and Storage in Protocells

* Scientific and Forensic Services, Inc., Delray Beach, FL. and Norfolk, VA


  1. Guth, J. H.  “Pre-Biotic Evolution:  I. From Stellar to Molecular Evolution”.  Society for the Advancement of Metadarwinism, Volume 1, November 19, 2014.   Accessible at
  2. Guth, J. H.  “Pre-Biotic Evolution:  II. Pre-Biotic Chemical Oscillations and Linked Reaction Sequences”.  Society for the Advancement of Metadarwinism, Volume 2, June 12, 2015.   Accessible at
  3. Taylor, D. L., J. S. Condeelis, P. L. Moore and R. D. Allen.  “The Contractile Basis of Amoeboid Movement, I. The Chemical Control of Motility in Isolated Cytoplasm”.  J. Cell Biol. 59:378-94 (1973)
  4. Norberg, B., U. Bandmann and L. Rydgren.  “Amoeboid movement in human leucocytes: basic mechanisms, cytobiological and clinical significance”.  J. Mechanochem. Cell Motil.  4:37-53 (1977)
  5. Dobereiner, H.-G., J. Kas, D. Noppl, I. Sprenger and E. Sackmann.  “Budding and Fission of Vesicles”.  Biophys. J. 65:1396-1403 (1993)
  6. Takakura, K. and T. Sugawara.  ” Membrane Dynamics of a Myelin-like Giant Multilamellar Vesicle Applicable to a Self-Reproducing System”.  Langmuir, 20:3832–3834 (2004)

© Copyrighted by Joseph H. Guth, 2015.  All rights reserved.

Mega-evolution: a Metadarwinian Extended New Synthesis (MENS)

Arnold De Loof

Functional Genomics and Proteomics Group, Department of Biology, KU Leuven, University of Leuven, Belgium. Address: Zoological Institute, Naamsestraat 59, 3000 Leuven, Belgium.



It is very logical to first formulate an unambiguous definition of Life before engaging in analyzing the parameters instrumental to its evolutionary change. However, nearly everybody assumes that catching the essence of Life in a single sentence that coherently lists all previously described properties of living matter is impossible. Yet a plausible but as yet undervalued definition that meets all essential criteria according to some philosophers of science already exists since two decades. It starts from the observation that all living matter is invariably organized in sender-receiver compartments that incessantly handle information (= communicate), thereby solving problems, most of it in an automated way. It reads: The verb ‘Life’ (as an activity) denotes nothing else than the total sum of all communication acts executed, at moment t, at all levels of its compartmental organization: L = ∑C. The key question in evolutionary theory becomes: “How can signaling activity change using both neo-Darwinian genetic- and Lamarckian non-genetic mechanisms?” At the cellular level, any act of communication is a problem-solving act because any message is coded. Hence it can be logically deduced that not Natural Selection itself but communication/problem-solving activity preceding selection is the universal driving force of evolution.

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Introductory remark

This invited paper is a concise compilation of 3 recent papers that describe in more detail the essence of Mega-evolution. For the complete set of figures, see the Open Access papers:

De Loof, A.  2014. Organic and cultural evolution can be seamlessly integrated using the principles of communication and problem-solving: The foundations for an Extended Evolutionary Synthesis (EES) as outlined in the Mega-Evolution concept. Life: The Excitement of Biology 2(4):247-269. http://dx.doi: 10.978444/LEB2(4)DeLoof.01

De Loof, A. 2015b. How to deduce and teach the logical and unambiguous answer, namely L = ∑C, to “What is Life?” using the principles of communication? Communicative & Integrative Biology. (accepted and scheduled to be published very soon)

De Loof, A. 2015c. From Darwin’s On the Origin of Species by Means of Natural Selection… to The evolution of Life with Communication Activity as its very essence and driving force (= Mega-Evolution) Life: the Excitement of Biology 3(3) 153-187.

  1. Introduction

The subject of micro-evolution is the change in allele frequencies that occur over time within a population. It is relevant to the emergence of new species. Thanks to Charles Darwin [1] and Alfred Russel Wallace [2] evolutionary theory with focus on species formation and Natural Selection as its driving force was founded. Macro-evolution acts on a scale of separated gene pools. It occurs at or above the level of species. Mega-evolution is a recent approach that takes into account recent insights in fundamental biological/biochemical/physiological processes [3]. It does not specifically focus on genetic changes like micro- and macroevolution do. It attempts to describe the evolution of ‘Life’ in its totality, irrespective of the way ‘Life’ manifests itself in the wealth of prokaryotic and eukaryotic species and their communities. Such approach requires that one first unambiguously defines what ‘Life’ exactly is before engaging in analyzing the various mechanisms/parameters instrumental to its change in the long run (evolution). From the standpoint of philosophy of science this is the logical way to proceed. Yet, such approach did not really get ground in the mainstream of current evolutionary theory, as exemplified in the formulation of the neo-Darwinian New Synthesis (NS) [4, 5, 6, 7, 8 and many others). Kutschera and Niklas [9] summarized the historical development of the term New Synthesis, from George Romanes who introduced the term to refer to the version of evolution advocated by Alfred Russel Wallace [2] and August Weismann (1834-1914) with its heavy dependence on Natural Selection, to Stephen J. Gould [8].

The main reason for the duality in which theory and practice do not match resides in the widely accepted idea that it is impossible to catch the nature of Life in a single sentence, a statement that features in textbooks of general biology, edition after edition for at least half a century up to the present (e.g. in Raven et al. [10]. Such (unwarranted) statements restrain younger newcomers who have an interest in such a fundamental question to engage in searching for a plausible answer.

This paper recapitulates these recent papers [11, 12, 13] describing that if one first defines ‘Life’, a novel paradigm emerges which brings unity in Biology, and which also enables us to answer the question whether Natural Selection should be replaced by Problem-solving activity as the universal driving force of evolution.

  1. An unambiguous definition of Life has already been deduced 20 years ago

In the late 1980’s, I was challenged by my undergraduate students to come up with a plausible definition to “What is Life?”, this to make me more credible as a professor of biology, the science of Life. Their reasoning was: “Why should we engage in the study of Life (= Biology) if one cannot define what ‘Life’ is”?. I accepted the challenge but I soon experienced that many had tried before me, without much success. No wonder as one gets confronted with all the criteria a good definition of Life should meet according to the philosophers of science Schejter and Agassi [14]. Their wording was: “Apart from its not being trite and uninformative (circular, to use a traditional term), it should be neither too wide nor too narrow; it should not exclude living things and it should not include dead ones. Furthermore, it should not make biology part-and-parcel of chemistry and physics (meaning that there should be room for an immaterial dimension).” I add: “and it should organize all known dimensions and properties of living matter in a logical order and context, and it should pave the way for defining what exactly happens at the moment of Death”.

But why did (and still does) nearly everybody assume that ‘Life’ cannot be defined? To my own surprise, the very reason turned out to be the result of the combination of asking the wrong question, with an understandable but nevertheless fatal thinking error.   Indeed, the common procedure at that time (the 1980’s-90’s) was to try to deduce the properties of Life by comparing the properties of ‘living matter’ with those of ‘non-living or inanimate matter’, assuming that these two conditions are true opposites like warm-cold, high-low etc. True opposites can only have one counterpart. But a given living entity, e.g. a dog can be opposed to a myriad of non-living entities: a bottle, a ring, a brick, a ship etc. Thus they are false opposites. The true opposites with respect to ‘Life’ are: ‘still alive versus ‘just dead’. This urges for answering: “What exactly changes at the very moment of Death?” Answering that question in a non-circular way e.g.”death ensues when life ends” imparted the following insights that: 1. Death ends an activity of a given system. That activity turned out to be communication activity of systems organized as sender-receiver communicating compartments; 2. There are numerous degrees of communicational complexity of most living systems, at least 16 in my classification system (see later).

The following definitions emerged:

  1. Death ensues when a given communicating compartment irreversibly (to exclude regeneration) loses its ability to communicate at its highest level of compartmental organization, the total number of such levels . It follows that the essence of being alive, or of ‘Life’ as an activity, is communication activity.
  2. Communication. Numerous definitions of communication have already been formulated one more complex than the other, but seldom all encompassing. My preferred definition reads: “Communication is transfer/handling of information in a system organized as a sender-receiver communicating compartment” (Fig. 1A). Any act of communication is generated as follows. A sender or the environment produces and releases a message(s) which is always written in coded form into what is called ‘a communication channel’ (blood, water, air etc.). The message (usually transported with the help of some carrier) will eventually arrive at a competent receiver (= with matching receptors). Here it will be captured, decoded, amplified and responded to by causing the mobilization of part of the stockpiled energy to do some ‘work’ sooner or later, e.g. by engaging in feedback. Depending upon the complexity of the system, numerous acts of communication can be simultaneously executed. All parts of any communicating compartment are subject to change; the sender, the message/messenger system, the transmission channel, the receiver, the feedback loops etc.

Feedback is not a circular but a spiral-like unidirectional activity (Fig. 1B). When the complexity of a signaling system increases, the possibility for generating more than one answer may arise; this happens at bifurcation points (Fig. 2B. In my opinion this necessity to make a choice is the very basis of ‘free will’. The more bifurcation points, the more possibilities for making use of free will.

Figure 1. The classical sender-receiver compartment (A) is a better alternative than the cell for functioning as the universal unit of structure and function of all living matter. Feedback is a spiral-like, unidirectional process (B). At bifurcation points, a choice has to be made as to how to proceed with communication. In digital-era wording, the Temple of Life has only 4 pillars (C), in contrast to the classical PICERAS Temple of Life that has seven. From De Loof (2015c).


  1. Information. My definition of information reads: A message contains ‘information’ when, upon being decoded by a competent receiver (= a receiver with the proper receptor(s)), part of the stored energy in that receiver is mobilized for doing some sort of ‘work’. This is the meaning of AT WORK in Fig. 1A. Information is itself immaterial, but it usually needs a carrier for being transported [12].
  2. A logically deduced unambiguous definition of Life (as an activity) reads: ‘Life’ sounds like a noun, but it is a verb. What we call ‘Life’ is nothing other than the total sum of all acts of communication exerted by a given sender-receiver compartment at moment t, at all levels of its compartmental organization (cell organelle, cell, tissue,…, whole organism,…, population,  community, Gaia level). The simplest symbolic notation reads: L = ∑C [12, 15].

Because “Life” is an activity of a given sender-receiver compartment of which there exist many different forms, one can specify it further as:





L= Life; S= type of compartment; t = moment at which the communication acts are executed; 1 = lowest level of compartmental organization (1 = prokaryotic cell or cell organelle in a eukaryotic cell); j = highest level of compartmental organization (cell, tissue, organ, organism, …, aggregate, …, population, community, the Gaia-level). For a symbolic notation that highlights how to compare ‘biological’ and ‘mechanical’ life (e.g. computer-life), see [12,15].

Thus ‘Life’ has both a qualitative (nature of the communication acts) and a quantitative (number of communication acts) aspect.

As to the origin of ’Life’: it came into being at the very moment that the first act of communication was executed [3, 16]. Fig. 2. illustrates this event in cartoon form (from [11]). How in pre-biotic conditions a living entity could have come into existence chemically has been discussed by Guth [17, 18]. The reasons why I think that the synthesis of actin-like molecules may have preceded the synthesis of RNA or DNA as information carriers have been outlined in [16].

Figure 1. Cartoon illustrating my view that Life came into being at the very moment that the first act of communication was executed.  Which act that was and under which environmental conditions this happened is unknown. Adapted from De Loof [3, 11].

  1. Time. If one thinks that the definition of ‘Life’ also requires that ‘Time’ has to be defined as well, I tried to do so: In my opinion ‘Time’ is invariably a property of a given energy-converting system. It is a measure for the inertia of the conversion of a given form of energy (heat, light, chemical etc) into another form(s) plus increase in entropy of the system (second law of thermodynamics). There are as many different times as there are energy converting systems [3]. This definition does not at all unveil why there is inertia in energy conversion, thus why such conversions do not proceed at an infinitely high speed. This continues to be a big mystery in physics.
  2. Evolution of Life

If L=∑C is an acceptable symbolic notation for ‘Life’, the simplest symbolic notation for its evolution becomes:

ΔL(T2-T1) =  Δ∑C(T2-T1)

  1. Some of the novelties in Mega-evolution as compared to classical evolutionary theory (neo-Darwinian New Synthesis)

4.1. The common descent principle was never better documented

This principle represents the very heart of Darwinian/Lamarckian evolutionary theory in both the New Synthesis and in the Mega-Evolution approach. Today it is very well experimentally documented [19[. In the past, a truly major novel insight has been formulated by the late Lynn Margulis. According to her symbiogenesis theory (1981), the eukaryotic cell came into being when at least 3 different ancient prokaryotic species established a functional symbiotic novel level of compartmental organization. Later in evolution, ever more complex multicellular eukaryotic entities came into being, requiring ever more complex coordinating signaling systems. The consequence of Margulis’ theory is that, in fact, all life forms on the planet earth, thus both the contemporary ‘genuine’ prokaryotes as well as all eukaryotes are manifestations of the only existing planetary form of life, which is bacterial in origin and nature. No other forms of life are known. According to [21] acquiring genomes was an important issue in the origins of species.

4.2. Not ‘the cell’ but ‘the sender-receiver’ as the universal unit of structure and function of all living matter

It has been outlined before [3, 11] that the ‘sender-receiver’ (Fig. 1A) better serves the role of universal unit of both structure and function of all living matter than ‘the cell’. In origin the term ‘cellulae’ was used by Robert Hooke (1635-1703) to denote the small chambers in cork. Later Schleiden and Schwann described that all living matter is made up of ‘cellulae’. The prokaryotic cell is the smallest sender-receiver. The Gaia-level is the highest one.

4.3. Levels of complexity in communicating compartments: more numerous than in classical biology

In introductory textbooks of biology, the usual levels of complexity are; cell organelle, cell, organism, population and community. In the Mega-evolution approach which uses communication as criterion for grouping ‘entities’, there are at least 16 levels of compartmental organization in living matter. This has been described at length elsewhere [3, 11]. The ≥ 16 levels can be grouped into three categories. Witzany [22] handles a similar communication-based classification system.

  1. Compartments restricted to a single individual (levels 1-8): prokaryote, eukaryote, cell aggregate, syncytium, mono-epithelium, polyepithelium, segmented organism, tool utilizing compartment.
  2. Compartments with individuals of the same species (levels 9-14): colony, heterosexual and social compartments, baby inside mother (internal budding) compartment, population/species, electrosphere compartment (e.g. humans linked by telephone, radio etc.).
  3. Compartments with individuals belonging to different species (levels 15-16): the community (with nutritional and/or protective aspects), and the planetary or Gaia compartment.

In classical evolutionary theory the main focus is on the population and species (genetics), which is level 13 (out of 16) in my classification system that takes into account the signaling pathways at all levels.

4.4. Instead of ‘Body and Mind/Soul’ rather ‘Hardware and Software’

One of the many reasons why it took so long before a plausible definition of Life was formulated [15] resided in the absence of an adequate vocabulary. The dichotomy ‘Soma or Body’ and – for humans- ‘Soul’ or ‘Mind’ reigned in Western culture for millennia. In Asian culture, that distinction was less clear-cut. Yet, defining ‘Soul’ was not evident. The term disappeared from the core of Psychology as a discipline (psyche = “soul” in Greek),but it continues to be an essential element in (some) religions. The question whether only humans have a soul or whether other organisms, in particular animals, also have a soul and are conscious, is no longer a scientifically valid question, but it continues to be asked again and again [23].  It is better replaced by the questions how the cognitive memory works, how widespread such memory system is and what its relation is with consciousness and problem-solving. Since the start of the digital era, the terms hardware and software (Fig. 1C) became widely accepted for computers. In biology and in particular in evolutionary theory, they are useful [3, 12, 13], be it that this is not yet common practice. ‘Hardware replaces ‘Soma’. Chemically the hardware of organisms is made up of fossil stardust ([3]. “Software” helps to describe some aspects of the cognitive memory. It is not a substitute for ‘Soul’.

4.5. Organisms have two memory systems. Two possible types of progeny

Like any sender-receiver all prokaryotic or eukaryotic cells on earth have probably two memory systems, a genetic- and a cognitive one, each with its own set of rules. The first central dogma DNA →RNA → Proteins [24] represents the very heart of the functioning of the genetic memory. Today its functioning is well understood. In contrast, despite all progress in the neurosciences, the biochemical functioning of the cognitive memory largely remains a black box [25]. One of the results is that “All inclusive inheritance” [26] uses heredity (= through genes) for all transfer of information to the next generation instead of transferability of information to the next generation (and laterally as well) which allows also taking into account teaching-learning involving the non-genetic aspects of the cognitive memory.

Physical children are the progeny generated through the principles of the genetic memory that underlies the formation of the hardware of organisms. Pupils are the progeny generated through the cognitive memory system.

4.6. Any act of communication is a problem-solving act by definition and can hence be instrumental to adaptation. Semiosis.

Why is an act of communication, at the cellular level, invariably a problem-solving act (Fig. 1A)? This follows from the fact that any message, whatever its nature is coded. Hence, when the message (often, if not always transported with the help of some carrier) arrives at the receiver and is captured there, it next needs to be decoded before it can trigger the receiver to ‘do something with it’, either instantly, or later after storage for some time, or it can be deleted. We understand our mother tongue but no other (foreign) languages because in our childhood our parents, family members, our broad environment etc. installed – by teaching – in our brain the decoding programs for our mother tongue. That gives us the impression that understanding our mother tongue is not a problem-solving activity. This interpretation is wrong; it is an automated decoding activity. The causal link between signaling and problem-solving is not commonly emphasized in the exact biological sciences, contrary to its status in the humanities, in particular in linguistics. Here the term ‘semiosis’ or ‘sign process’ is routinely used [27, 28, 29, 30 and others]. It was introduced by Charles Sanders Peirce (1830-1914) to denote any form of activity, conduct, or process that involves signs, including the production of meaning. I agree with Kull and Emmeche [29] that because it incessantly interprets signs and signals, “Life is semiosis”. My wording L = ∑C [15] said the same but in the wording of the exact biological sciences.

4.7. Adaptation to an environment poisoned by high Ca2+-concentrations

Organisms have to adapt to changing external conditions. The environment can become dryer, wetter, colder, warmer, less rich in food supply, populated by more parasites etc. When chemical pollution as an adverse condition is at stake, one usually thinks at man-caused pollution by pesticides, heavy metals, CO2 etc. Yet the most toxic pollutant on earth (O2 not taken into account) is the omnipresent Ca2+-ion. This may look strange because we encounter the beneficial aspects of Ca2+ in our daily life: our calcareous bony skeleton, Ca2+-rich milk, the egg shell of birds, and Ca2+ as a secondary messenger [29]. Yet, because above a very low threshold a rise in cytoplasmic Ca2+ concentration is very toxic as it causes changes in the conformation of some essential macromolecules, in particular proteins. In fact it is because of this toxic effect on proteins that Ca2+ can act as secondary messenger. The intracellular Ca2+ concentration in the cytoplasm of unstimulated cells amounts to about the vey low value of about 100 nanomolar.  The extracellular concentration is many orders of magnitude higher, namely about 1-60 millimolar (2 mM in blood). Thus there is a gradient of about 100,000 times in Ca2+-concentration cytoplasm-outside cells. If the intracellular Ca2+-concentration rises too much for too long, cells can get damaged and may even enter the apoptosis cell death cycle (Calcium-induced apoptosis: Orrenius et al. 31]). The duration of the heart contraction cycle which is based upon periodic Ca2+-release from the SER followed by fast re-uptake is an indication of what “too long” means, namely in the order of seconds rather than of minutes in most cell types. The toxicity of Ca2+ means that cells have to continuously fight against the influx of excess Ca2+ from the outside world (environment). Their major weapon is the different types of ATP-driven Ca2+-ATPases in both the plasma membrane and in the internal membrane systems. The cellular system for maintaining Ca2+-homeostasis and other types of homeostasis as well, a most important issue in cellular physiology and evolution is complex [32, 33, 34]. It is in this context that the self-generated inorganic ion-based cellular electricity and the lipid nature of cell membranes has to be understood. This is well worded by John Torday [33] as” The history of physiologic cellular-molecular interrelationships can be traced all the way back to the unicellular state by following the pathway formed by lipids ubiquitously accommodating calcium homeostasis, and its consequent adaptive effects on oxygen uptake by cells, tissues and organs”.

Lipid membranes are not permeable to inorganic ions unless they harbor proteinaceous ion channels and pumps. They are permeable to electrons which means that self-generated cellular electricity could not function if it were electron-based.  Self-generated inorganic ion-based electricity is vital to life. A cell is dead when its electrical dimension collapses [35, 36]. An overlooked key feature of cells is that all cells are able to drive an electrogenic electrical current through themselves, at least during part of their developmental cycle, and that they are polarized (for figures, see [13].  This is contained in “The cell as a miniature electrophoresis chamber concept” [35].

4.8. Not Natural selection but problem-solving activity preceding selection is the universal driving force of evolution

Neo-Darwinists hearing somebody contesting the generally accepted view that Darwin’s Natural Selection is the universal driving force of evolution probably experience this as cursing in a cathedral. Metadarwinists may consent (see website: The third way of evolution [37]). The problem for both is that at present it remains difficult to clearly define the mode of action of Natural Selection and to present examples where it has been at work [38]. If selection would nevertheless not be the driving force, what is the alternative? I argue that if one changes paradigm away from the NS by starting to define at first ‘Life’ and next analyses what mechanisms may be instrumental to its variability, problem-solving activity preceding selection emerges as the long-sought for alternative. In a former paper [13], I used the example of students doing an exam to illustrate this principle. The general perception is that the examiner, not the students taking the exam does the selection. Yet, if one analyses the system, the opposite conclusion emerges. The teacher-examiner formulates the questions. In evolutionary wording, he/she constructs some gradient, like nature would build temperature-, light- etc.  gradients. It is up to the students to show their ability to overcome the exam-gradient. Thus, they engage in self-selection, a principle advanced by [3] as ‘Gradient-Provoked Swelling/Shrinking Self-Selection or GP-Triple S Principle’. The examiner only lists their success or failure. The principle of self-selection is further strengthened when the student succeeds in solving the problem by feedback, i.e. by answering in such a way that the sender/teacher will (deliberately or not) lower the gradient (e.g. by changing the subject of examination).

But problem-solving activity is inherent to communication activity which itself is a synonym for Life (as an activity). This leads to the unexpected and counterintuitive conclusion that Life itself is the driving force of its own evolution. In other words, the principle of Life being an activity of compartments that are invariably organized in sender-receiver entities contains the endogenous mechanism for driving its own evolution. In my opinion, this is a magnificent principle.

4.9. Cultural evolution is evolution “the software way”

Neo-Darwinism did not yet succeed in plausibly incorporating cultural evolution into the mainstream of evolutionary theory [11]. The main reason is that the New Synthesis reduces all causes of variability under the common denominator of genetic changes (Fig. 3).

Figure 3. Major genetic and non-genetic causes of (Communicational) variability. Not only Charles Darwin (1809-1882) but his contemporary Alfred Russel Wallace (1823-1913) as well independently conceived the theory of evolution through natural selection. Jean-Baptiste Pierre Antoine de Monet, Chevalier de Lamarck but commonly referred to as simply Lamarck is best known for his theory of inheritance of acquired characteristics that was proven wrong in the context of classical genetics. Epigenetics is a form of temporary transfer of genetic information (through DNA- and/or histone modification) to the next (few) generation(s). According to some researchers such transfer is Lamarckian in nature. Cultural evolution is also mainly Lamarckian in nature.

As long as one assumes that the principles of the cognitive memory are inherent to those of DNA → RNA → Proteins, this assumption is the only possible one. But the assumption is wrong. The cognitive memory system has its own rules and mechanisms which include self-generated electrical activity. This activity is based on the transport of inorganic ions, and thus only partially dependent upon the central dogma [24]. Cultural evolution is mainly achieved through the possibilities of the cognitive memory. In digital era wording, it is evolution ‘the software way’ while organic evolution is evolution ‘the hardware way’, using the principles of the genetic memory [11]. In fact, organic- and cultural evolution are the two sides of the very same coin, which is evolution of Life with its two memory systems. As stated before, cultural evolution is achieved through pupils who function as the software progeny counterpart of physical children.

4.10. Evo-Devo. Haeckel’s “Ontogeny recapitulates Phylogeny”.

Development may be regarded as the accumulation of changes during lifetime, thus as evolution in the very short run [39, 40]. The key issue in development is the differential use of the same genome in all cells of a differentiating organism, a few exceptions not taken into account. The generation of cells which all differ in their membrane-cytoskeletal properties by what has been called ‘The double asymmetry principle’ (for figure see [13]) is causal to this differential use of the same genome [41, 42]. There is no role for mutations in development, this in contrast to long-term evolution in which mutations do play a key role. In my opinion, Ernst Haeckel’s law “Ontogeny recapitulates phylogeny” remains a valid key concept in evolutionary theory.

5. Discussion

The 2014 Nature paper of Laland et al. [43] shows that among evolutionary biologists the conviction is gaining ground that the neo-Darwinian New Synthesis needs an upgrade, but unanimity on this opinion has not yet been reached. Whether one is pro or contra an upgrade may be influenced by one’s major study object. If one focuses on sessile organisms like e.g. plants, one may be inclined to assume that the NS explains well enough the mechanisms of evolution, some details not taken into account. If one focuses more on free living organisms like e.g. animals, one may favor the view that some systems partially direct their own evolution [44], and therefore an upgrade is urgently needed. Free living organisms benefit more from adaptations in mobility and from the possibilities offered by the cognitive memory system for elaborating strategies for improving their survival and reproductive success. Another cause of pro-contra thinking may concern the type of evolution one is interested in. The humanities are primarily interested in ‘cultural evolution of the Homo sapiens. The exact biological sciences consider the Homo sapiens not as a special case for which another type of evolution needs to be invoked, but as one of the numerous terrestrial species. They are more interested in the organic-chemical evolution of this and other species, no matter whether they live in an aquatic or terrestrial habitat.

If one agrees that the numerous novel insights generated by the novel disciplines in biology [13, 45, 46, 47, 48, 49, 50, 51 etc.] need to be incorporated in an extended evolutionary new synthesis, one faces the question how such integration and unification can be achieved. The importance of communication for understanding Life and its evolution has been approached in various ways by e.g. [15, 29, 52, 30, 33 and others]. In my opinion, the most straightforward approach is to start from a plausible definition of Life that is acceptable to both the humanities and the exact biological sciences. Communication activity executed by sender-receiver compartments is the key issue in such definition [12, 15]. It leads to the question how the architecture and functioning evolved from the probably simple Progenote as the primordial sender-receiver into the multitude of organismal and supra-organismal entities that function as sender-receivers.

Neo-Darwinists and Metadarwinists both agree on the common descent principle, the very heart of Darwinism. They differ in opinion(s) on a number of topics, e.g. on the relative importance of epigenetics, on the weight one should give to the overall importance of genetic changes as instrumental to bringing about (all) evolutionary change as well as on the significance of Natural Selection as the universal driving force of evolution. NS primarily focuses on the effects of all kinds of mutations (Fig. 3) and on species formation through the possibilities of only one memory system, namely the genetic memory and the central dogma DNA → RNA → Proteins. This is apparent from the formulation of ‘the all inclusive inheritance principle’ [26] that acknowledges that in addition to all sorts of mutations, there are indeed other causes of variability instrumental to evolution. But in the end their effects can all be explained by one memory system, the genetic one. But cells/organisms have in addition to their DNA memory, a cognitive memory system. Although it continues to be a (biochemical) black box, there is no reason to neglect its existence and importance. Darwin did not know the principles of the genetic memory but he took them into account. As a result, NS fails to adequately incorporate cultural evolution into the mainstream of evolutionary theory. This type of evolution relies more on the cognitive memory system. As long as NS does not accept a software upgrade, it will remain a theory of the evolution of the hardware of living matter as governed by the principles of genetics. Such type of evolution is very slow. It usually (but not always) operates at the geological time scale. Mega-evolution takes two memory systems into account. Through teaching and learning which are mainly enabled by the cognitive memory, evolution by non-genetic mechanisms (which is ‘evolution the software way’ in my approach) can be very fast as illustrated by the recent evolution of the species Homo sapiens. Another example is the coming into existence of a new Darwin finch species on the Galápagos island Daphne Major that took only 4-5 generations, starting in 1981 [53]. These data illustrate the power of the introduction of a dialect in a language as instrument for reproductive isolation, an important issue in species formation.

Some people may not like the idea that our body is in fact a clump of some 100,000 billions (= 1011) eukaryotic cells that by themselves are the symbiotic result of a few (3?) ancient bacteria. Mitochondria are modified bacteria. Each eukaryotic cell contains several mitochondria. In addition, numerous bacteria live on the surface and in the alimentary canal of animals. All these subcellular and cellular entities have to cooperate which means that the communication networks (signaling pathways) inside any multicellular organism are numerous. The complexity can be orders of magnitude higher in populations, communities etc. In a recent (2015) internet discussion forum) Kalevi Kull posted the quote that “Life is semiosis. Life is a network of sign processes and that this is obviously the most exact and brief definition of life”.

I advocate replacing the widely accepted concept that “Natural Selection is the driving force of evolution” by “Problem-solving activity preceding selection (like when doing an exam) is that universal force”. One could argue that in the end it does not make much difference: the best adapted (which are not necessarily the ‘strongest’ ones) will do better. Yet, the formulation does make a substantial difference because it necessitates answering the question which biological principle enables problem-solving. The answer is that problem-solving does neither follow in full from the central dogma nor from the fact that all living matter is cellularly organized. It is inherent to the organization of all living matter in senders-receivers that continuously handle information, thereby solving problems, most of them in an automated way. This approach necessitates that one rethinks several aspects of evolutionary theory. For example, should one continue to attribute so much weight to “species formation”? Or, what is the unit of selection: the cell, the organism, the species etc. or the signaling pathway as instrumental to problem-solving or the sender-receiver compartment? How to better incorporate the principles of physiology in evolutionary theory [13, 48, 54].

Because of the multitude of signaling pathways and their endless interactions the scope of Metadarwinism (in particular the MENS approach as explained in [13] and in this paper) is much broader than that of NS. MENS is better rooted in physiology, a weak point of NS. Instead of ‘heredity’, MENS prefers “transferability of information to the next generation(s) (and where relevant, laterally as well) by all means, thus also by the possibilities of a second memory system, the cognitive memory. This way it manages to seamlessly integrate both organic- and cultural evolution [11].

The Mega-evolution approach urges for changes in teaching biology. For the moment textbooks of biology seldom explain the principles of communication, probably because the authors assume that these principles are self-evident (because we communicate all the time without any problem), and that therefore they do not need extensive explanation. In reality, the opposite is true. The fact that most communication happens in an automated way indicates that it is far from simple. How could it become automated? Upon analysis, it becomes clear that the mechanisms of communication are at least as sophisticated as those of genetics. In particular, the role of the cognitive memory in signaling is still a black box, despite all progress in neurobiology.

In the recent past I repeatedly stated that, paraphrasing Theodosius Dobzhansky (1973), “Nothing in biology and evolutionary theory makes sense except in the light of the ability of living matter to communicate, and by doing so, to solve problems”. Given its continuing observational and descriptive nature the discipline of Biology keeps missing a unifying principle comparable to E = mC2 for physics or the atomic model for chemistry. Torday [34] summarized this with the characterization by Earnest Rutherford as ‘stamp collecting’. In my opinion, if properly incorporated in teaching L = ∑C harbors the potential for shedding the (not fully mistaken) perception that many biologists insufficiently grasp in full the very nature and importance of the principle that can integrate all subdisciplines of Biology, namely communication.


I thank all students and colleagues who helped me to streamline my communication-based view of evolution. My thanks too to Julie Puttemans, Marijke Christiaens and Katrien Becuwe for help with the figures, and to Michael Gaffney for text correction.

Conflict of interest



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Legends to Figures

Figure 1. The classical sender-receiver compartment (A) is a better alternative than the cell for functioning as the universal unit of structure and function of all living matter. Feedback is a spiral-like, unidirectional process (B). At bifurcation points, a choice has to be made as to how to proceed with communication. In digital-era wording, the Temple of Life has only 4 pillars (C), in contrast to the classical PICERAS Temple of Life that has seven. From De Loof (2015c).

Figure 2. Cartoon illustrating my view that Life came into being at the very moment that the first act of communication was executed.  Which act that was and under which environmental conditions this happened is unknown. Adapted from De Loof (2002, 2014).

Figure 3. Major genetic and non-genetic causes of (Communicational) variability. Not only Charles Darwin (1809-1882) but his contemporary Alfred Russel Wallace (1823-1913) as well independently conceived the theory of evolution through natural selection. Jean-Baptiste Pierre Antoine de Monet, Chevalier de Lamarck but commonly referred to as simply Lamarck is best known for his theory of inheritance of acquired characteristics that was proven wrong in the context of classical genetics. Epigenetics is a form of temporary transfer of genetic information (through DNA- and/or histone modification) to the next (few) generation(s). According to some researchers such transfer is Lamarckian in nature. Cultural evolution is also mainly Lamarckian in nature



How Can a Molecule Behave as If It is a Brain?

Jon Lieff, M.D.
Past President, American Association for Geriatric Psychiatry
Assistant Clinical Professor, Tufts University Medical School

There are several competing theories of what mind could be in nature. Is it an emergent property of molecules and neurons in the brain, an electromagnetic energy field interacting with molecules, cells, organisms and brains or something else? There is no definite answer. The unusual intelligent behavior of cells including bacteria, intestinal epithelial cells, skin cells, platelets, T cells, astrocytes and microglia is now acknowledged. This complex behavior of cells, without a brain, raises the question of whether mind exists throughout nature and somehow interacts with molecules and cells. The question asked in each of these articles is how can a cell behave as if it has a brain?

Surprisingly this same decision-making, communication and complex behavior is also observed in viruses.  A question then arises whether jumping genes and prions demonstrate similar types of complex behavior as viruses.  This would imply that even a molecule could exhibit a form of intelligence.

This article describes the remarkable behavior of the small molecule mTOR that forms many large complexes along with other molecules. By sensing many different cellular processes at once and simultaneously responding to all of them, this one molecule appears to behave as if it had a brain.

The information in this article is distilled from a variety of more complex research reviews. It is a simplified description of behavior outlined in extreme detail in molecular biological research articles. Two excellent recent reviews of this material are The Neurology of mTOR, Jonathan O. Lipton and Mustafa Sahin,

Neuron 84, October 22, 2014 a2014 Elsevier Inc. and Nutrient-sensing Mechanisms and Pathways, Alejo Efeyan, William C. Comb, doi:10.1038/nature14190. Other references appear at the end of the article.

Many Functions of mTOR

How can one complex protein function as if it is a nervous system? mTOR is able to monitor a large amount of different external and internal information and use this data to make critical decisions and take many actions. The decisions involve multiple pathways controlling cellular growth and the amount of protein manufacturing. Corresponding actions include triggering specific genetic networks for many different tasks including balancing of basic metabolism and energy production. It is difficult to imagine how mTOR can perform so many different critical functions that integrate so much information related to the cell or an organisms relation to its environment.

Because its effects are so vital for survival of the cell and the organism, many different diseases are related to mTOR dysfunction, such as diabetes, cancer, tumors, epilepsy, degenerative brain disorders, depression and autism. In addition, even while behaving like a brain itself, mTOR has critical functions throughout the human brain.

Major accomplishments of mTOR are directly regulating the amount of proteins created , the amount of RNA made from DNA (transcription), critical elements of energy metabolism, the creation and maintenance of many different organelles and programming cellular death. mTOR is also directly involved in brain functions of all types, such as making neural stem cells to populate the developing brain, creation of neuronal circuits, neuroplasticity and very specific functions of sleep, eating, and circadian clocks.

mTOR Stands for Mammalian Target Of Rapamycin

The strange name of this vastly complex vital protein is derived from the discovery of the antibiotic rapamycin in a microbe on Easter Island, known locally as Rapa Nui. Rapamycin was found to block a protein, which was called “mammalian target of rapamycin” or mTOR. Rapamycin stops the fungus cell division cycle. It also stops this same division cycle in human B-lymphocytes and is now used to suppress the immune system after transplants. Later, many more functions were discovered.

mTOR is the critical mediator of many different pathways and signals including the very important functions of making proteins and regulating nutrients and energy. It integrates many critical inputs, such as insulin, growth factors, amino acids, oxygen and general energy levels. As a result, it is also critical in many diseases such as diabetes, obesity, depression, cancers and brain diseases.

Rapamycin forms a large complex, which binds to a part of mTOR and decreases its activity. There are, in fact, two different mTOR Complexes—mTORC1 and mTORC2—which operate both independently and together. They are often found in different cellular compartments, but act together for many different functions.

mTOR is the molecule activating, regulating and inhibiting the functions of the two large complexes. Both complexes 1 and 2 consist of many large proteins that form the structure of the complexes and aid in the many functions of stimulating and inhibiting the most important pathways and cascades in the cell. Rapamycin’s action is to block a particular protein only when this special protein is connected to the complexes. Rapamycin acts differently in the two complexes.

Two Interacting Large Complexes

mTOR Complex 1 (mTORC1) senses nutrients, energy and oxidation pathways and controls the manufacture of proteins with messenger RNA and ribosomes. For many years it was known that the amino acid leucine stimulated mTOR and it was thought that leucine was the critical signal for all amino acids, for example in starvation of calorie restriction experiments. But, recently a completely different second mechanism has been found for the amino acid glutamine, which opens the question of whether mTOR, in fact, responds to many other amino acids. The mechanisms for other amino acids are not known. Since it is very difficult to study metabolic pathways, the many ways that mTOR senses nutrients is only now being discovered. For, examples, mTOR is stimulated by insulin, growth factors, blood factors, phosphatidic acids and oxidative processes.

mTOR Complex 2 (mTORC2) is itself composed of many different, equally complex proteins. This is known to regulate the cytoskeleton of the cell. It stimulates the addition of high-energy phosphate particles to proteins and many other molecules for important metabolic functions. Regulation of the cell scaffolding, and therefore its shape in building axons and dendrites, is through the action of actin. mTOR’s regulation of actin is also related to programmed cell death and cell survival.

mTOR1 is critically related to the function of ribosomes and ribosomes are necessary to activate the second complex. The first complex stimulates building of a ribosome, which activates the second complex. The many interactions of the two complexes make study even more difficult.

Activation of mTOR

The activation of mTOR is very complex, stimulated by a wide variety of factors. Their functions are based on complex shape as with other large proteins.

A variety of powerful neurotrophic factors stimulate mTOR pathways including glutamate, special guidance molecules, BDNF (brain derived neurotrophic factor), IGF1 (insulin like growth factor), VEGF vascular endothelial growth factor) and CNTF (cilliary neurotrophic factor).

Another powerful signal is Rheb that is suppressed by many different other factors including TSH (tuberous sclerosis complexes -1 and 2). TSH, itself, is highly regulated by multiple cascades with many important well-known kinases (ERK, AKT, GSK, Wnt). These various pathways stimulate mTOR is various ways, both increasing and decreasing different activity.

There are many different mechanisms to regulate mTOR. Some of these pathways relate to the use of energy in the cell, when more or less is needed at different times and places. mTOR triggers or suppresses metabolic cycles to accomplish these goals.

Amino Acid Sensing

It was thought that a specific amino acid, leucine, was the most important sensor for mTOR among amino acids. Recent dramatic research findings demonstrate that the mechanisms for this protein to sense the amount of two different amino acids are entirely different. They not only have different mechanisms, but the mechanisms are in different cell compartments. Yet, they both interact with the “growth regulatory complex” of the cell.

Sensing the amount of different nutrients available is highly linked to the metabolic processes that make large important molecules for the cell to grow and multiply. There are special systems of receptors that signal to mTOR through the vital sphosphoinositide-3-kinase (PI3K) cascade. There are multiple enzymes that communicate with mTOR to signal that there are enough amino acids present.

The lysosome organelle (usually thought of as a large membrane sac that breaks down large molecules and microbes) is part of this mTOR activation for amino acid sensing. A super-complex on the lysosome’s membrane surface is where mTOR is activated. Since lysosomes take apart large molecules, it is possible that mTOR monitors a pool of amino acid materials in the lysosome. Four hundred genes in humans make protein carriers for the lysosome membrane to transport many different substances, including ions, into the lysosome. Amino acid transporters are just now being identified. It is not yet clear how many different mechanisms there are for different nutrients.

Many Factors Stimulate mTOR

 One of mTORs very important functions is regulating the translation of messenger RNA into proteins at the ribosome. A series of enzymes are involved in this function. The cap of the messenger RNA (methylated guanosine repeat at 5’ end of the DNA) is affected by enzymes that start the process of making proteins. Several factors compete in this process through mTOR.

A series of molecules controlled by mTOR are transcription factors. Transcription factors are proteins that bind to specific places in the DNA, triggering the start and stop of the process that will make proteins. Transcription factors can promote or activate (promoters and activators) or stop (repressors) and they, also, attract the important enzyme that transfers code form DNA to messenger RNA—RNA polymerase.

An important group of mTOR factors are involved in regulating the use of lipids for energy in the cell. These factors sense lipid nutrients, regulate axonal myelin and produce neuronal action potentials. They are also related to several neurodegenerative diseases.

mTOR responds to the lack of oxygen in the cell by controlling the DNA and ribosomes that make a particular protein transcription factor regulating the response to low oxygen. These factors shift metabolism from an oxidative state to glycolytic pathways. This same mechanism is also involved in stimulating the growth of more blood vessels when low oxygen is caused by a stroke or other damage causing low blood flow to tissues.

mTOR activates another factor through complex mechanisms that affect mitochondria function. In fact, blocking mTOR can create many different problems that occur in mitochondria.

mTOR Control of Autophagy

Autophagy is a complex process that cells use to recycle its material. It takes apart amino acids, large molecules and dysfunctional organelles. mTOR inhibition triggers autophagy, which is related to many degenerative diseases and cancer.

Autophagy in the brain is complex and important, but not well understood. For example, altering the pathway through mTOR causes movement disorders, destruction of neuronal axons, a particular ubuiquitin tagging of important proteins and possible death. These mTOR related problems are correlated with mis-folded proteins that are the hallmark of brain disease.

It is now known that autophagy is very involved in the creation of dendrite spines and their elimination when not needed. This may be the way mTOR is involved in the social behavior of the organism. Importantly, it is found that autophagy in neurons is unique with several distinct opposing mTOR pathways for inhibition and stimulation. Therefore, in the brain, mTOR is a vital point of regulation.

mTOR Signaling is Vital in The Brain

In animal experiments, alteration of mTOR in the fetus eliminates many crucial regions in the developing brain. Without mTOR, the neuronal stem cells do not produce enough neurons. In fact, mTOR appears to be critical in creating the windows of brain development seen in babies. But, this effect when exaggerated in research goes both ways—stopping neuron production and overproducing neurons. When the molecules that regulate mTOR are not present, excessive signals can dramatically change brain structure—multiple axons on neurons and alterations of dendrite structure, with increased size but fewer spines.

 In the brain, it has been difficult to distinguish the effects of mTOR 1 and 2. Altering either of these factors created smaller, abnormal brains. In disease, mTOR 1 might have a greater effect on myelin than 2, but they clearly operate together in regulating the lipids for myelin.

mTOR is also part of signaling between different types of cells such as astrocytes and neurons.

When mTOR was disrupted in research it altered visual circuits. This occurs through an unusual mechanism wherein mTOR affects the guidance molecules for axon travel. In order for axons to travel to regions far away from the neuron’s cell body, the axon responds to cues along the way. These cues interact with mTOR pathways producing local stimulation of ribosomes and manufacturing proteins that are needed in particular places for the growth and direction of the axon.

Research is now focusing on identifying critical RNAs for axon growth and creation of synapses. The synapse needs a tremendous amount of local production of highly specialized molecules. While the messenger RNA comes all the way from the nucleus, the ribosomes, transfer RNAs and protein factors are right at the synapse. It is mTOR that stimulates this entire process.

There are many other ways that mTOR is regulated in the brain

  • Immune systems interact with mTOR in creating neuronal circuits, such as insulin regulating synapses using MHC immune molecules
  • During brain injury or spinal cord injury, mTOR uses(((“uses fetal mechanisms that have been quiescent from the fetal period until the injury to stimulate axon growth and local production of proteins.
  • Ketamine triggers glutamate NMDA receptors stimulating mTOR, which increases critical proteins at the synapse and dendrite spines.
  • mTOR regulates potassium channels in dendrites.
  • Activities of mTOR stimulate creation of the specific brain neuronal circuits.

mTOR In Neuroplasticity Learning and Memory

It was first learned that rapamycin stopped the strengthening of synapses for neuroplasticity by stopping production of critical proteins. Later, it was found that as is the case for all actions of mTOR, there are many different interacting pathways. The receptor 1 is linked to the form of neuroplasticity called long-term depression—a decrease in the strength of a synapse (as opposed to long term potentiation).

When glutamate mGluRs receptors are triggered, they stimulate mTOR and increase proteins at the synapse to make it stronger in long-term potentiation. Blocking mTOR stops this learning. mTORC2 then regulates the actin cytoskeleton, critical to the growing axons, dendrites and synapses.

 Alterations in mTOR pathways have produced hippocampus deficits in animals, as well as decreased general learning and memory and abnormal fear conditioning and spatial learning. In fact, any change in the mTOR pathways can have dramatic effects on all types of learning and cognitive behavior. It appears that mTOR is the center of many interactive pathways that have dramatic positive and negative effects on synapses, neuronal circuits and behavior.

mTOR in Energy Regulation

The very complex regulation of cellular energy allocation is critically involved in the monitoring of all types of different nutrients and the specific needs of different cells involved in growth and all other types of energy usage. Energy needs are also  related to the activity and motivation of the organism. mTOR is at the center of all of this.

mTOR is very involved in the extremely complex regulation of eating. There are many different pathways involved in appetite but two primary ones are centered in the hypothalamus using different cells producing opposing neurotransmitters and peptides. mTOR is involved in both increased eating with obesity and decreased eating with starvation. Leptin is a critical signal when full. This signal is channeled through mTOR to produce special molecules to initiate or  to stop eating. mTOR also appears to be critical to the anti aging effects of calorie restriction.

mTOR is further critically involved in circadian rhythms that triggers gene networks and specific protein production that regulates it. However, the exact mechanisms are just being discovered. Although it is not known exactly how the extremely complex and varied actions of mTOR regulate circadian rhythms, it is clearly related to the creation of special proteins for synapses that have profound effects on sleep. It is known that the learning that increases in sleep is meditated by mTOR neuroplasticity.

Diseases Caused by Alterations in mTOR

It is natural that such complex pathways would be significant in many different diseases, such as tuberous sclerosis, some forms of severe autism, neurofibromatosis, Fragile X syndrome, epilepsy, Alzheimer’s, Parkinson’s, Huntington’s, depression, schizophrenia and many tumors.

  • Reactive oxygen decreases high-energy phosphates in mitochondria and inhibits mTOR pathway decreasing manufacture of proteins by stopping ribosomes. Altering mTORC1 stops mitochondria respiration and production of energy.
  • In tuberous sclerosis and other tumors, abnormal pathways produce abnormal neurons and large glia.
  • mTOR abnormalities produce symptoms autism among many other symptoms.
  • Genetic mTOR tumor diseases cause seizures, which respond to treatment with rapamycin, possibly related to effects on migration of neurons and axons, production of axons and dendrites and regulation of action potentials.
  • Factors that trigger mTOR pathways such as lack of oxygen, inflammation and the electrical events in neurons can interact with genetic defects in the complex pathways.

Because mTOR is the most important measure of nutrients and energy needs, it can, also, be related to aging. Although, the mechanism is not known in animals, rapamycin increases life span. It is also involved in the mechanism by which calorie restriction increases life span, probably through the control of protein manufacturing.

  • mTOR signaling is associated with both increased amyloid and abnormal tau in Alzheimer’s. Lowering mTOR signals lowers both. mTOR’s relation to autophagy is critical for the elimination of the mis-folded amyloid and tau.
  • In Parkinson’s, rapamycin increased autophagy and decreased abnormal alpha-synuclein proteins.
  • Both autophagy and mTOR are related to increases in Huntington’s abnormal protein clumps.
  • The rapid, but temporary, improvement in depression with ketamine (a glutamate NMDA antagonist) is based on mTOR pathways. Also, rapamycin blocks this effect.
  • One gene that has been associated with schizophrenia is related to mTOR pathways.

How Can a Molecule Behave As If It is a Brain?

How can one molecule be a sensor for many different nutrients, oxygen, energy, and then control protein synthesis and help remodel the brain? Previous posts have noted that cells, such as microbes, behave as if they have a brain by integrating many senses and making many different decisions simultaneously. Others have demonstrated unusual behavior of viruses with only a handful of genes and proteins, but are still able to perform hundreds of complex behaviors, while tricking vastly larger human immune cells. Observing the actions and capabilities of viruses, jumping genes and prions, the question has to be raised about mind interacting with these individual molecules.

But, how can one molecule behave as if it is a brain by itself, performing the same feats as the microbe—analyzing many different sensory inputs and making many simultaneous complex decisions and actions?

 How can the evolution of this molecule be explained, when so many interlocking different processes depend upon its exact structure?


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Jewell JL, Kim YC, Russell RC, Yu F-X, Park HW et al. 2015. Differential regulation of mTORC1 by leucine and glutamine. Science 9 January 2015: Vol. 347 no. 6218 pp. 194-198 DOI: 10.1126/science.1259472

Lipton JO, Sahin M. 2014. The neurology of mTOR. Neuron 84 (2): 275-391. 22 Oct


Wang S, Tzun ZY,  Wolfson RL, Shen K, Wyant GA et al. 2015. Metabolism: Lysosmoal amino acid transporter SLC38A9 signals arginine sufficiency to mTORC1. Science. 2015 Jan 9;347(6218):188-94. doi: 10.1126/science.1257132. Epub 2015 Jan 7.

Evo-DevoENV the Science Concerned With the Record at Ontogenic Stages of the Interaction of Living Organisms and Their Enivornment During the Evolutionary Process

Jorge Herkovits

Instituto de Ciencias Ambientales y Salud, Fundacion PROSAMA, Paysandu 752 (1405) Buenos Aires, Argentina.

Key words: ontogenesis, developmental biology, environment, biomarkers,        evolution,  record, evo-devo, evoecotoxicology, paleoecotoxicology, gaia theory

Abstract. Although it is generally accepted that environmental features can select resistant individuals within species and resistant species within ecosystems, most of our understanding of the evolutionary process´ interactions between the environment and life forms is extremely limited and controversial.  This article focuses on the hypothesis that living organisms at ontogenic stages could be considered biomarkers of the successive interactions with the environmental agents during the evolutionary process including their influence on the environment toward a living planet.  A proposal for a universal brand name to link all fields related to ontogenesis and evolution is suggested.

  1. The established links between ontogenesis and evolution

The link of ontogenesis to evolution originates in the comparative embryology of the nineteenth century in the work of von Baer and Haeckel whose “laws” -embryonic divergence and recapitulation- were presented as being generally applicable to the way in which phylogenesis evolves. Although ontogenesis had a minor role both in the work of Lamarck, Darwin/Wallace  and the establishment of the modern synthesis in evolution, the developmental patterns of otogenesis help to explain the branching evolutionary divergences in the metazoan and the relationships among phyla (Maier 1999).  As developmental genetics advanced it became increasingly evident that every species is like a history book. Present day evo-devo erupted out of the discovery of the homeobox and the conservation of spatiotemporal expression pattern of those developmental genes. It is increasingly evident from evo-devo studies on organisms ranging from plants to mammals that the evolution of distinct morphologies relies on the reutilization of a relatively small set of master regulatory genes such as the Hox genes, which establishes a segmental pattern of activation (Meyerowitz 2002). This process of developmental reprogramming implies that if a particular ontogeny is represented by a trajectory through multidimensional phenotypic space, then many types and degrees of morphological differences can be caused by programming a different trajectory (Swalla, 2002; Davidson 2002).  For instance, the great extent to which changes in plant forms have been engendered was exemplified by heterochrony (temporal shifts in developmental pathways) or heterotopy (spatial shifts in developmental pathways). Assessment of heterochrony or heterotopy is usually attributed to morphological features and seems to occur due to the expression of developmental genetic pathways in a new context. The ontogenetic trajectory is often influenced by environmental factors (the reaction norm). This sometimes occurs in a rapidly adapting manner, as in the case of aquatic plants that make different kinds of leaves above and below water (heterophylly), or the insects that produce winged and wingless forms at different population densities. Moreover, the whole ontogenetic trajectory could be deflected in a continuously variable way in response to environmental conditions. Eco-evo-devo, a study focused on the interactions between an organism’s environment, genes, and development, contributes to understand the evolutionary implications of these phenomena. Its major goals are to uncover the rules that underlie the interactions between an organism’s environment, genes, and development, and to incorporate these rules into evolutionary theory; for a recent review see Abouheif et al (2014).

2. The record of evolutionary environmental signatures during the ontogenesis of living organisms.

Environmental features essential for ecosystems, conservation of biodiversity and human health evolved from the time of the Earth´s accretion around 4.5 billion years ago. These features were crucial for the development of biota from its early forms, around 3.8 billion years ago. Present day biodiversity, estimated to comprise more than 100 million species, has developed on the basis of the ability of life forms to adapt to the evolving environmental scenarios at a planetary level and multiply at a rate that surpassed both background and mass extinction events. Aside from the abiotic inputs of chemicals in the environment, a living planet was established with environmental conditions increasingly of biological origin as life expanded worldwide and biodiversity increased. For example, the increased O2 in the water and atmosphere was due to the achievement of photosynthetic water-splitting capacity about 2.4 billion years ago (Anbar and Knoll 2002). However, our understanding of the evolutionary processes linking the various life forms and their environment is very limited, especially for the initial 3.3 billion years of evolution. This fact is due to the sampling intensity from different periods of time, as well as the preservation of fossils which bias our knowledge on the existent biodiversity and environmental conditions during different periods of the evolutionary process. The uncertainty for these ancient times is so profound that even in a concept article on the development of multicellular biodiversity, no time for this basic evolutionary step is suggested (Wolpert and Szathnary 2002).  Thus, it could be highly valuable to explore alternative avenues to contribute to the understanding of the evolutionary processes on the Earth.  In this article, I will revisit case studies to research how radical changes in metabolic features and the susceptibility to noxious agents during ontogenesis can be used as biomarkers of environmental features during the evolutionary process (Herkovits, 2006).


  1. i) The oxygen signature. The oxygen consumption of amphibian embryos exhibit marked changes as embryonic development advances: at the egg cell stage 5.7, tail bud 35, open mouth 118 and at complete operculum 180uL/hr for 100 embryos (Herkovits and Jatimliansky 1982). Conversely, survival times in anoxia shifted from more than 30 h at 2 days after fertilization to 20 h at 10 days of age, to only 2-4h at 14 days of age (Adolph 1983). A shift in the mitochondrial enzymes towards an aerobic metabolism was reported at the gastrula stage (Lovtrup-Rein and Nelson, 1982). Anaerobic metabolism in early embryonic stages was reported in a wide range of species and therefore could be generalized to embryonic development. For instance, the energy that mammals require in the preimplantation embryo are generated by anaerobic metabolism. (Adolph 1983, Robkin 1997; Burton 2003). Moreover, reducing oxygen concentration from atmospheric levels during in vitro culture generally improves early embryonic development across a range of species (Booth et al 2005). The same pattern of low oxygen uptake by early stage embryos occurs in invertebrates such as intertidal crabs, which increases its O2 consumption over 10-fold by the time of hatching (Taylor and Leelapiyanart 1997). Artemia is an extreme example, as 60% of early life stage embryos could survive 4 years of continuous anoxia at physiological temperatures (Clegg 1997). The anaerobic metabolism at early developmental stages in different species can be interpreted as an evolutionary trait that protects the embryo from oxidative stress damage and/or as an array of adaptations that enable them to survive a wide variety of environmental extremes. As Though the embryo should successfully pass through all of its developmental stages, this does not seem to be advantageous for present day environmental conditions (which have existed through the last 2 billion years), to achieve anaerobic metabolism as an adaptive feature just for the initial developmental stages (Herkovits, 2006).

Is this critical metabolic change during ontogenesis related to an environmental signature during the evolutionary process?  From an O2 perspective, it is generally accepted that the biological and geochemical history of Earth can be separated into two supereons (Anbar and Knoll 2002). Based on independent geochemical evidence on oxygen availability, it can be deduced that surface water was already oxygenated between 2.4 and 2 billion years ago (Rue and Bruland 1995; Johnson et al 1997). O2 is usually considered the first biogenerated environmental pollutant to appear in large quantities on the planet, after which anaerobes died or restricted themselves to environments that O2 did not penetrate, while other organisms began the evolutionary process of evolving the use of O2 for metabolic transformation e.g. for efficient energy production (the mitochondrial oxidative phosphorylation system).  In this context, the fact that living organisms exhibit anaerobic metabolism at early developmental stages is generally accepted to be the case for living forms in the ancient anoxic Earth. This provides support for the hypothesis that living organisms recapitulate their metabolic features at embryonic stages during their phylogenetic evolution and conversely, they could be considered as biomarkers of environmental features such as the transition from an anoxic to an oxic Earth. Moreover, taking into account that intercellular adhesiveness exists in protists and is achieved in amphibian embryos at very early blastula stage (Herkovits 1978), the earliest multicellular life forms seem to be a very ancient achievement in the anoxic Earth. In any case, it is noteworthy that the recapitulation at early embryonic stages of ancient life has features like anaerobic metabolism, which seem to be essential for the accomplishment of normal ontogenesis even for vertebrates. Anaerobic multicellular organisms exist in present day, but their evolution in complexity is very limited compared to those achieved by aerobic metazoans.

The transition towards an aerobic metabolism from the gastrula stage in living organisms corresponds within the EVO-DEVOenv perspective to an evolutionary period with major achievements in cell differentiation and morphogenesis in the rising free oxygen environment. In the case of amphibian phylogeny it  implies the rearrangement of the cells toward a tridermic organism, and within this process the evolution led to early chordate life forms. Moreover, an increasing aerobic metabolism seems to be essential for the development of the cephalic nervous system (Herkovits, 2014).   Based on a battery of studies including phylogenetics, structural biology, protein engineering, metabolism, competition and genomics, it was concluded that adaptation to environmental conditions was already in place 3.5 billion years ago (Zhu et al 2005). Therefore, the creatures from the Ediacaran, Tommotian, Oman, Chengjiang and Burgess Shale can be viewed as part of a broader picture with roots in multicellular life forms in the deep anoxic Earth history. The EVO-DEVOENV theory implies a new dating method, e.g.  the shift from anaerobic to aerobic metabolisms during early embryonic stages versus the transition from an anoxic to an oxic Earth, allowing us to anticipate that multicellular organisms flourished over 2 Gy ago (Herkovits, 2006);  they were discovered in 2010 (El Albani, et. al. 2010).

The colonization of terrestrial habitats by amphibians around 380 million years ago (Carol 1987) implied not only the shift from uptake of oxygen from water to the higher oxygen concentrations in the air, but also the possibility  of mobilization requiring less energy demand from the media from which oxygen is obtained. Among biomarkers of the access to the higher oxygen environment and the concomitant need for enhanced protection against the increasing oxidative stress, a novel and more efficient glutathione S-transferases isoenzyme is highly expressed in post-metamorphic amphibian liver (Amicarelli et al 2004). It is well accepted that this last evolutionary step related to oxygen consumption in amphibian phylogeny corresponds to a process working its way towards occupying a new niche, the terrestrial habitat. The access to higher free oxygen levels at least 200 million years ago seems to have rendered living organisms with homoeothermic metabolism (Carol 1987).  It is noteworthy that both the mammalian as well as the bird embryo have poikilothermic metabolism till birth (the homoeothermic condition is provided by the parent organism). The usual explanation of this fact is based on the energy needs of the embryo, which are considerably less than if it were a comparably sized homoeothermic organism. Although in present environmental conditions this explanation could be acceptable, from the EVO-DEVOENV perspective poikilotherm metabolism of the avian and mammalian embryo  reflects the evolutionary process starting from anaerobic  living conditions during the anoxic period of the Earth onwards to the last metabolic achievement.


3. The susceptibility to noxious agents at ontogenic stages versus environmental signature during evolution

Could the well documented stage dependent susceptibility to physicochemical agents during developmental stages (Herkovits et al 1997; Rutledge, 1997; Degitz 2000;  Kast-Hutcheson et al. 2001; Fort et al., 2004) be related to environmental features during the evolutionary process?  The high resistance at the blastula stage to physicochemical stress (Perez-Coll et al 1990; Herkovits et al 1997), enhanced in free living embryos by protective barriers like the vitelline membrane and jelly coats could reflect very aggressive environmental conditions during the evolution of early multicellular organisms. For instance, the high toxicity associated with UV-B irradiation in the anoxic Earth could be associated with the very high resistance of amphibian embryos at the blastula stage to UV-B (Castañaga et al, 2007), as well as other agents exerting oxidative stress (Pérez-Coll and Herkovits 1990; Herkovits et al. 1997; Vismara et al 2001). Again, the reciprocal elucidation of ontogenic features and environmental signatures during the evolutionary process provides support that metazoan organisms existed in the ancient anoxic Earth.  Moreover, by focusing on these early developmental stages, there is a remarkable stage-dependent susceptibility within the blastula (Bustuoabad et al. 1977) reflecting environmental changes during the early phases of multicellular life forms in the anoxic Earth. This is not surprising when evaluated from an EVO-DEVOenv perspective, as blastula-like organisms existed during hundreds, if not over a billion years of evolution in the anoxic Earth.


The high resistance to environmental agents during the initial developmental stages contrasts with the high susceptibility of the organism as cell differentiation and morphogenetic processes achieve increasing complexity (Pérez-Coll and Herkovits 1990; Herkovits et al. 1997; Vismara et al 2001; Bogi, 2003; Christensen et al 2005). This juxtaposition is directly related to a gradual increase in aerobic metabolism and the associated oxidative stress. The fact that the organogenic stages are very susceptible to noxious agents in spite of the high capacity of the embryo at those developmental stages to recover from adverse effects (Herkovits and Faber 1978) contributes to the  idea that the increasing complexity of cell differentiation and  morphogenesis could be achieved during the evolutionary and ontogenetic processes  under benign, low environmental stress conditions. Metamorphosis, also a complex cell differentiation and morphogenetic process in both invertebrate and vertebrate organisms, is also a period of very high susceptibility to a variety of environmental agents  (Howe et al. 2004; Wilson 2004). Thus, the stage dependent resistance profile to noxious agents during the ontogenesis of any species could reflect the environmental stresses supported by their ancestors during the evolutionary process.


4. Ontogenesis and the construction of a living planet.

Global environmental conditions can be altered by both abiotic and biotic inputs (Herkovits, 2006). The biota has a significant effect on the Earth’s environment, such as the rise of free oxygen during the evolutionary process (Anbar and Knoll  2002) and oxygen production  and consumption from that time onwards. According to the Gaia hypothesis the whole ecosystem can be seen as a giant organism in which life tends to optimizes both the physical and chemical environments to best fit their needs (Lovelock, 1986;  Lenton 1998).  The rise of O2 in the water and atmosphere initiated by photosynthetic cyanobacteria about 2.4 billion years ago represents an example of the magnitude of the impact of living organisms on the Earth´s environment and the effects of environmental changes on living forms. We discovered that amphibian embryos neutralize the acidic condition produced by different noxious agents like glyphosate, aluminum or even citrate buffer (Piazuelo et al., 2011; Herkovits et al., 2015). As acidic conditions were documented in ancient environmental scenarios (Knoll et al., 1996), the capacity of amphibian embryos to neutralize acidic pH could be considered  a biomarker of ancestral organisms actively adjusting environmental conditions to their needs. Thus EVO-DEVOENV also provides the possibility  of studying the effects of living organisms on ancestral environmental conditions. This could  contribute  to a better understanding of the coeveolution of living organisms and their environment, providing the possibility to obtain  experimental data on the participation of individual species or a set of species in the buildup of environmental condition that benefit life.  As a whole, based on the EVO-DEVO ENV synthesis, our study provides some evidence that a pH of 4 probably was the lower pH condition in the habitats of  South American amphibian ancestors,  it  is the limit for embryo survival, their capacity to modify environmental pH and thus their lower limit within  the resilience phenomenon. During the last few years several model systems have emerged for addressing the interconnectedness between an organism’s environment, its development, and its ecological interactions in natural populations (Van Valen’s 1973; Ledón-Rettig and Pfennig 2011).  Our results point out that besides internal mechanisms of defense against toxicity, living organisms could contribute to modifying environmental pH toward their benefit.


  1. Developing a unique brand name to link ontogenesis and evolution.

During the last 20 years, there has been a notable increase in contributions that are oriented to link ontogenesis and evolution from different perspectives. There are journals and interest groups devoted to this field like the European Society for Evolutionary Developmental Biology, the Society for the Advancement of Metadarwinism, the Pan American Society for Evolutionary Developmental Biology, etc. Generally, most of the traditional scientific societies in the biological field could be interested in contributions linking ontogenesis and evolution.  As contributions have emerged from different disciplines, several brand names, acronyms and definitions have been presented  in the literature e.g. eco-evo-devo in the case of the ecological impact on development, Evoecotoxicology in the case of ecotoxicological criteria to report the record of environmental signatures during the evolutionary process during the ontogenesis of living organisms, etc. EVO-DEVO is by far the most popular term and acronym. Thus, my proposal is to use EVO-DEVO as the brand acronym for all the studies oriented to reporting links between ontogenesis and evolution and by means of a superscript identifying the main discipline of each contribution. For instance GEN if the study focuses on genetic aspects (EVO-DEVOGEN), ENV in the case of environmental issues (EVO-DEVOENV), ECO in the case of ecological subjects (EVO-DEVOECO), PHYSIO in the case of physiology (EVO-DEVOPHYSIO), PATH in the case of pathology (EVO-DEVOPATH), etc. By using this method, we will construct a robust multidisciplinary and interdisciplinary brand name for all the contributions focused on establishing the links between ontogenesis and evolution orienting from  the brand name  the nature of each contribution.


  1. Conclusion

The possibility of considering living organisms at ontogenetic stages as biomarkers of the evolutionary process of both environmental features and living forms allows the reconstruction of the evolutionary process  on Earth. Some biomarkers of environmental conditions, like those related to the rise of oxygen starting about 2.4 billion years ago and the subsequent changes towards aerobic metabolism, have global significance and therefore it could be anticipated that they appear in all aerobic organisms at specific developmental stage(s) according to their phylogenetic trajectory. Conversely, biomarkers related to very local features like the case of serpentinite-hosted hydrothermal field beneath the mid-ocean ridge (Kelley et al. 2005) will occur only in organisms living in those environmental conditions.  From a methodological point of view, simple ecotoxicological studies can provide the unique opportunity to study in just one experiment the natural selection process affecting individuals (the survival of the most resistant individuals) and the capacity of a population to adjust to environmental conditions (e.g. pH in the case of glyphosate, aluminum, etc) to their benefit (e.g. Herkovits et. al., 2015). The reciprocal elucidation approach of ontogenic features and environmental signatures during the evolutionary process could  integrate information from toxico-ecotoxicology, geochemistry, paleontology, cell differentiation, morphogenesis, physiology, metabolism, genetics, epigenetics, pathology, evolution, phylogenetics, etc. as a multidisciplinary and interdisciplinary approach to understanding the evolutionary process by means of a rational and holistic explantation. It includes  the  susceptibility/resistance features to noxious agents (Herkovits, 2006). As an overall picture, multicellular, blastula-like organisms existed at least 2.4 Gy ago, but may even date back to 3 Gy ago in the deep anoxic world, while aerobic tridermic organisms emerged around 2.4 billion years ago. Initially, living forms had to survive in very adverse environmental conditions, which is the reason  for their high resistance to noxious agents, including  UV-B irradiation, a fact reflected in the present day by the high resistance to noxious agents at early developmental stages. Conversely, organogenesis and metamorphosis, both  exhibiting complex cell differentiation and morphogenetic processes, were achieved on an evolutionary scale during low level environmental stress conditions  expressed in present day as high susceptibility to a wide range of different noxious agents. This reflects  complex processes requiring very low stress levels in order to achieve success.  In summary, EVO-DEVOENV provides the possibility for considering each species  as a history book of both the environment and life forms contributing to a better understanding of the evolutionary process on Earth.

Jorge Herkovits is a scientist of the National Council of Science and Technology (CONICET), Argentina. This study received support from Fundacion PROSAMA and I thank the skilful revision of English by Lilly Backer


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Pleiotropy and Evolving Novelty

by  Pleiotropy and Evolving Novelty

John S. Torday, MSc, PhD.

Key Words: pleiotropy, growth factor, growth factor receptor, cell-cell interaction, Parathyroid Hormone-related Protein, Prostaglandin E2, βAdrenergic Receptor, type IV collagen, Adipocyte Differentiation Related Protein, Neutral Lipid Trafficking

Pleiotropy, the Deus ex Machina (Ghost in the Machine)

Based on the conventional ‘snapshot’ of an organism’s physiology, pleiotropy is generally construed as the same gene randomly utilized for various differing and flexible purposes. As a classic example of pleiotropy’s pervasive effects, the preeminent evolutionist George Williams utilized this phenomenon to explain, for example, that senescence occurs as the price for Darwinian reproductive advantage. He described this phenomenon as Antagonistic Pleiotropy (1) – when one gene controls more than one trait, one of these traits being beneficial to the organism’s fitness, and another detrimental to it.

However, might pleiotropy actually occur deterministically rather than by chance, based on specific physiologic principles, thereby revealing the true nature of evolution? It can be productively advanced that pleiotropy has fostered evolution through iterative interactions between the First Principles of Physiology and the ever-changing environment (2). Pleiotropic novelties emerge through recombinations and permutations of cell-cell interactions for phenotypic adaptation based on both past and present conditions, in service to the future needs of the organism for its continued survival. Thus, in contrast to Antagonistic Pleiotropy based on descriptive biology, based on a cellular-molecular mechanistic approach senescence can be seen as the loss of cellular communication due to the natural decline in bioenergetics resulting from selection pressure for optimal reproductive success earlier in the life cycle of the organism (2).

Rubik’s Cube as a Metaphor for Pleiotropic Evolution

Erno Rubik invented his eponymous ‘cube’ [Fig. 1] to teach his students about spatial relationships and Group Theory (3). By twisting and turning the cube, you can generate 4 x 1019 permutations and combinations of green, yellow, white, orange, red and blue squares in space and time. Similarly, as an embryo ‘twists and turns’ in biologic space and time during development it generates hundreds of different cell-types via cell-cell signaling to form the human body; Lewis Wolpert, the renowned Developmental Biologist has said that “It is not birth, marriage, or death, but gastrulation, which is truly the most important time in your life”. That may be because it is at that stage in embryogenesis that the single layered cell membrane of the embryo becomes three-layered, generating the endoderm, mesoderm and ectoderm that ultimately give rise to the various cell-types of the organism (4, 5). Moreover, those various cell-types formulate tissue-specific homeostatic interactions to accommodate structure and function. The fact that the genes of each cell are all the same, and yet are all phenotypically different, both within and between tissues, has remained enigmatic within evolutionary development.

Pleiotropy is the expression of a single gene that generates two or more distinct phenotypic traits (6) – much like twisting a Rubik’s cube and forming various permutations and combinations of colors. In the case of the biologic process, it generates the various cellular phenotypes that compose the body, with equally varied homeostatic interactions (7). If this process is followed phylogenetically and ontogenetically, it provides insights to the mechanisms of evolution (8), just as unicellular organisms gave rise to multicellular organisms under the iterative, interactive influences of both internal and external environmental selection pressures (9). The Rubik’s cube metaphor for pleiotropy aids in understanding how one gene can affect multiple phenotypes. For example, it can be noted that there are images of cells on the faces of the Rubik’s cube in Fig. 1 that are associated with different colors. As the cube is twisted to reconfigure the color combinations, those cellular images are re-permuted and recombined. The inference is that the cellular phenotypic traits are modified in much the same way as they would have been during the process of evolutionary adaptation. Therefore, the reallocation of genes and phenotypic traits is not due to random selection, but rather is determined by homeostatic constraints within each newly-established cellular niche (10). Those constraints evolved from the unicellular bauplan and its homeostatic needs during the transition from one life cycle to the next, remaining consistent with those homeostatic constraints at every scale, phylogenetically, developmentally and physiologically alike (11). If they do not, they can either be compensated for by other genetic motifs (12), they can be ‘silenced’ (13), or they can be embryonically lethal (14). It is this process that explains how and why physiologic traits are pleiotropically distributed throughout biologic systems. More importantly, it provides the mechanism for evolutionary novelty, since pleiotropy offers the opportunity to ‘repurpose’ pre-existing genetic traits for different phenotypic functions, as needed.

In the book Evolutionary Biology, Cell-Cell Communication, and Complex Disease (9), the pleiotropic property of biology was utilized to explain the evolutionary mechanisms for both physiology and pathophysiology. In the former case, our work demonstrated how the alveolus of the lung and the glomerulus of the kidney are virtually the same functionally at the cell signaling level (9, 15, 16), even though they nominally seem to be structurally and functionally unrelated when seen from a descriptive perspective- one mediates gas exchange between the environment and the circulation, the other mediates fluid and electrolyte balance in the systemic circulation. However, both of these organs sense and transduce pressure signals, and thereby regulate homeostasis through stretch-regulation of Parathyroid Hormone-related Protein (PTHrP) production by the epithelium and its receptor-mediated signaling to specialized neighboring fibroblasts. Ironically, their homologous physiologic origins are recognizable through pathophysiologic conditions like Congestive Heart Failure and Goodpasture’s Syndrome. In the case of the former, heart failure commonly disrupts homeostatic control in both the lung and kidney (17) due to PTHrP dyshomeostasis in both (18, 19). In the case of the latter, the evolutionary adaptation to land mediated by the Goodpasture’s Syndrome Type IV collagen alpha3(IV)NC1 isomer, which is hydrophobic, protecting against water loss (19), can cause death due to the generation of autoantibodies that can cause heart and kidney failure (20) .

Regarding the physiologic commonalities between the lung and kidney, in the case of the lung, the stretch-regulated PTHrP produced by the epithelial type II cell feeds-back to its receptor on the lipofibroblast to regulate lung surfactant production, reducing surface tension to maintain alveolar homeostasis (21); in the case of the kidney, PTHrP produced by the epithelial podocytes that surround the fluid-filled space within the glomerulus regulate the mesangium, the thin mesodermal membrane supporting the glomerular capillary loops, homeostatically monitoring and regulating fluid and electrolyte balance in the systemic circulation (22).

The Lung as the Prototypical Pleiotropic Mechanism

The evolution of the lung was existential for the survival of land-dwelling vertebrates, since the rise in atmospheric temperature due to the Green House Effect of rising levels of carbon dioxide caused the drying up of bodies of water, forcing our forebears to adapt to land (23). The physicochemically-integrated developmental and phylogenetic cell-cell interactions regulating lung surfactant offer the means of understanding the ontogenetic and phylogenetic structural-functional interrelationships at the cellular-molecular level between the decrease in alveolar diameter and the increased lung surface area for gas exchange. The counterbalancing of the otherwise pathological increase in alveolar surface tension due to the decrease in alveolar diameter would have resulted in its collapse, or atelectasis (24); conversely, the evolution of epithelial-mesenchymal interactions for the concomitant thinning of the alveolar wall and the progressive efficiency of the surfactant system facilitated alveolar accommodation of gas exchange.  This is the only biologic means for increasing oxygenation (25).

Beginning with the fish swim bladder as a biologic mechanism for adapting to water buoyancy- inflating to float, deflating to sink- fish have successfully exploited gas to optimize their adaptation to water buoyancy. All of the key molecular features of the mammalian lung as a reciprocating gas exchanger were already present in the fish swim bladder- surfactant phospholipid and protein to prevent the walls of the bladder form sticking together (26), PTHrP functioning during swim bladder development (27), and the βAdrenergic Receptor regulating the filling and emptying of the swim bladder with gas absorbed from or secreted into the circulation (28). These components of the evolutionary process were capable of re-permutation and recombination within the physiologic constraints of the existing structure and function to form the lung (29). The only additional critical physiologic adaptation to be acquired was Neutral Lipid Trafficking (NLT) (30), mediated by Adipocyte Differentiation Related Protein (ADRP) (31), a member of the PAT (Perilipin, ADRP, TIP47) family of lipid transport and storage proteins wherever lipids are stored (32). NLT likely evolved from the adaptive advantage of lipofibroblast neutral lipid storage, initially for protecting the lung gas exchange surface against oxidant injury (33), followed by its regulatory role as a means of more efficiently producing surfactant in response to the ever-increasing excursions of the alveolar wall in response to metabolic demand (30). This is the epitome of the mechanism of pleiotropy, repurposing adipocyte metabolism for both the respiratory system and for the emergence of homeothermy (11), synergistically facilitating vertebrate adaptation to land through a common functional homolog.

The Lung as an Interactive Barrier- Homolog of the Plasma Membrane, Skin and Brain

Developmentally, the lung emerges from the foregut as an expansion of the surface of the alimentary tract (34). As a homolog of the gut, the lung also acts as an interface between the internal and external environments of the body. However, the homology goes much deeper molecularly since the stratum corneum of the skin forms a lipid barrier on its surface much like the alveolar surfactant, forming tubular myelin as a membrane barrier (35)- in both cases (35,36), the epithelium secretes lamellar bodies composed of lipid-protein complexed with antimicrobial peptides. And the skin and brain are structurally-functionally homologous, both phylogenetically and pathophysiologically- the nervous system of the skin in worms gave rise to the central nervous system of vertebrates, referred to as the ‘skin-brain’ (37). Pathophysiologically, the skin and brain share common lipodystrophies in such neurodegenerative diseases as Niemann-Pick (38), Tay Sachs (39) and Gaucher’s Disease (40). It has been speculated by some that this is a reflection of “too much of a ‘good thing’ going bad” (41). In this case, the excessive myelination of axons in the brain causes tandem skin lipid lesions in association with brain neuronal pathology.

For example, the functional homology between the lung alveolus and kidney glomerulus are enacted by shared mechanotransducers for the physiologic stretching of their respective walls- in the case of the lung, alveolar PTHrP signals to increase surfactant production, preventing its collapse due to increased surface tension (42). In the case of the kidney, the podocytes lining the glomerulus also secrete PTHrP, which then signals the mesangium to regulate water and electrolyte economy as a function of fluid distension (43). In either case, the calcium-regulatory activity of PTHrP, which is ubiquitously expressed in all epithelial cells (44), has been embellished due to its myriad functionally evolved properties (45-48). For example, due to its angiogenic property (48), PTHrP promotes microcirculatory capillary formation for gas exchange in the alveolar bed, and fluid and electrolytes in the glomeruli. Phylogenetically, within the fish kidney, the growth of the primitive filtering capillaries of the glomus would presumably have been stimulated locally by PTHrP, ultimately culminating in the expansion of the capillary network to form glomeruli, increasing the efficiency of water and electrolyte homeostasis in service to land adaptation (49).

NKX2.1, Thyroid, Pituitary and Lung Pleiotropy

The foregut is a plastic structure from which the thyroid, lung, and pituitary arise through the Nkx2.1/TTF-1 gene regulatory pathway (50). Evolutionarily, this is consistent with the concept of terminal addition (51), since the deuterostome gut develops from the anus to the mouth (52). Developmentally, when Nkx2.1/TTF-1 is deleted in embryonic mice, the thyroid, lung, and pituitary do not form during embryogenesis (53). This provides direct experimental evidence for a genetic common denominator for all three organs. Their phylogenetic relationship has been traced back to amphioxus, and to cyclostomes, since the larval endostyle (a longitudinal ciliated groove on the ventral wall of the pharynx for gathering food particles) is the structural homolog of the adult thyroid gland (54).

The Phylogeny of the Thyroid

The endostyle is retained in post-metamorphic urochordates (55), and in adult amphioxus (56), but the post-metamorphic lamprey has a follicular thyroid gland, which is an evolved endostyle (57). The presence of an endostyle in larval lampreys does not suggest direct descent of lampreys from protochordates, but rather that the evolutionary history of the lamprey is deep and ancient in origin, and that it shares the common feature of having a filter-feeding mechanism during its larval stage of development (58). However, it is noteworthy that the other extant agnathan, the hagfish, possesses thyroid follicles before hatching (59). Since hagfish evolution is considered to be conservative, going back 550 million years, this suggests that thyroid follicles could also be considered to have an ancient history (59).

An Evolutionary Vertical Integration of the Phylogeny and Ontogeny of the Thyroid

Mechanistically, the increased bacterial load consequent with the facilitation of feeding by the endostyle may have stimulated the cyclic AMP-dependent protein kinase A (PKA) pathway, since bacteria produce endotoxin, a potent PKA agonist (60, 61). This cascade may have evolved into regulation of the thyroid by Thyroid Stimulating Hormone (TSH), since TSH acts on the thyroid via the cAMP-dependent PKA signaling pathway (62). This mechanism potentially generated novel structures such as the thyroid, lung, and pituitary, all of which are developmentally induced by the PKA-sensitive Nkx2.1/TTF-1 pathway (63). The brain–lung–thyroid syndrome, in which infants with Nkx2.1/TTF-1 mutations develop hypotonia, hypothyroidism, and respiratory distress syndrome, or surfactant deficiency disease, provides further evidence for the coevolution of the lung, thyroid, and pituitary (64).

Developmentally, the thyroid evaginates from the foregut in the embryonic mouse beginning on day 8.5 (65), about one day before the lung and pituitary emerge, suggesting that the thyroid may have been a molecular prototype for the lung during evolution, providing a testable and refutable hypothesis. Adaptationally, the thyroid rendered molecular iodine in the environment bioavailable by binding it to threonine to synthesize thyroid hormone (66), whereas the lung made molecular oxygen tolerable, first by inducing fat cell–like lipofibroblasts as cytoprotectants (67), which then stimulated surfactant production by producing leptin (68), relieving the physiologic oxygenation constraint on the blood–gas barrier by making the alveoli more distensible (69). This, in turn, would have further facilitated the use of rising oxygen in the atmosphere metabolically, placing further selection pressure on the alveoli, giving rise to the stretch-regulated surfactant system mediated by PTHrP and leptin (9). Subsequent selection pressure on the cardiopulmonary system may have facilitated liver evolution, since the phylogenetically increasing size of the heart (70, 71), accommodating the water-land transition, would have induced precocious liver development- induction of liver development is determined by the physical interaction between the heart and liver (72)- fostering increased glucose regulation, e.g. gluconeogenesis and glycogen storage/release. In turn, this may have fostered brain evolution since the brain is a glucose ‘sink’ metabolically (73, 74). Further evolution of the brain, specifically the pituitary, would have served to foster the evolution of complex physiologic systems, culminating in endothermy/homeothermy in mammals and birds (11).

Both the thyroid and lung have played similar adaptive roles by accommodating otherwise toxic substances in the environment during vertebrate evolution. The thyroid has facilitated the utility of iodine ingested from the environment (75), whereas the lung has accommodated the rising oxygen levels during the Phanerozoic era (76). Importantly, both the thyroid and lung have interacted synergistically in facilitating vertebrate evolution- for example, Thyroid Hormone stimulates embryonic lung morphogenesis during development (77), while also accommodating the increased lipid metabolism needed for surfactant production by driving fatty acids into muscle to increase motility (78), as opposed to maladaptively oxidizing circulating lipids to form toxic lipoperoxides (79). The selection pressure for metabolism was clearly facilitated by the synergy between these foregut derivatives.

A Retrospective Understanding of Evolution

Looking at the definitive structure and function of the mammalian alveolus [Fig. 2], one can see the signature for phylogenetic traits that facilitated the evolution of land vertebrates from fish in a step-wise fashion. Referring to the Schematic [Fig. 2], at the far left is the transition from prokaryotes to eukaryotes, which may have been the result of the effect of rising oxygen tension in the atmosphere on sterol production (80).  This scenario would resolve the age-old debate as to whether evolution was gradual or salutatory – it was both. This is a key insight to understanding mechanistic evolution. Historically, Darwin thought that evolution was a slow and gradual process (81).  He did not think that this process was smooth, but rather, that it should be presumed to be stepwise, with species evolving and accumulating through small variations over long periods of time.  Darwin further speculated that if evolution were gradual that there would be fossil evidence for small incremental change within species.  Yet Darwin and his supporters have been unable to find most of these hypothesized ‘missing links’.  Darwin surmised that the lack of fossil evidence was due to the low likelihood that such critical transitions would have been preserved. Then, in 1972 (82) evolutionary biologists Stephen Jay Gould and Niles Eldredge suggested that the “gaps” in the fossil record were real, representing periods of stasis in morphology, calling this mode of evolution “punctuated equilibrium.” This infers that species are generally morpholgically stable, changing little for millions of years.  This slow pace is “punctuated” by rapid bursts of change resulting in new species.  According to this theory, changes leading to new species do not result from slow, incremental changes in the mainstream population. Instead, changes occur in populations living on the periphery, or in isolated populations where their gene pools vary more widely due to slightly different environmental conditions.  When the environment changes, such peripheral or isolated species possess variations in morphology that might allow them an adaptive advantage.

A bridging concept can account for both Gradualism and Punctuated Equilibrium. The kinds of mechanisms that have been invoked for pleiotropy would account for both scenarios. As Darwin had surmised, evolution could have occurred on a continuous molecular basis microscopically in response to physiologic stress (83), occasionally leaving fossilized evidence once form and function reached a macro-scale, only making it seem as though evolution had occurred in bursts (yet the molecular evidence can be seen in the continuum from ontogeny and phylogeny to pathophysiology!).

A scenario for two differing rates of evolutionary change is all the more cogent when one superimposes the episodic increases and decreases in atmospheric oxygen that have been documented over the last 500 million years, referred to as the Berner Hypothesis (76). Within this theory, the increases in atmospheric oxygen caused the well-documented increases in the size of land animals (84). However, the decreases have never been considered before, yet would predictably have had profound effects on vertebrae evolution, given that hypoxia is the most potent effector of complex physiologic systems (85). Elsewhere (11), a novel mechanism for the evolution of endothermy/homeothermy based on the interactions between the pulmonary and neuroendocrine/endocrine systems has been evoked that allows for the arc of the Cambrian Burst, culminating in the crown species of mammals and birds. This perspective is validated by the pleiotropic effects of the specific gene duplications for the PTHrP Receptor (86), the βAdrenergic Receptor (87), as well as the differentiation of the Glucocorticoid Receptor (88) and the evolution of the Goodpasture’s Syndrome Type IV collagen isomer (19), all of which occurred during the water-land transition. These events corroborate the repurposing of pre-existing genes for novel phenotypic adaptations.

Even earlier in vertebrate evolution, sterols may have liquified the bacterial cell wall (80), possibly due to rising levels of oxygen in the atmosphere (89) stimulating sterol production. That event would have marked the phenotypic transition from prokaryotes to eukaryotes, the former having hard exterior walls, the latter having compliant cell membranes. That transition may have been further catalyzed by the nascent synthesis of cholesterol (90), under positive control by Hypoxia Inducible Factor-1 (89), catalyzing the evolution of eukaryotes (see Fig. 2). The two horizontal, bolded arrows for ‘endothermy’ and ‘oxygen’ were the major drivers of vertebrate evolution. All three of highlighted processes- prokaryote/eukaryote evolution, oxygen and endothermy- have acted synergistically to promote vertebrate evolution, indicated by the dotted arrows that interconnect them.


The seemingly serendipitous occurrences of pleiotropy based on the conventionally descriptive understanding of biology are over-arched by the synchronically mechanistic basis for pleiotropy, emanating from the cell-cell signaling principles elucidated above. Thus, the deep, otherwise-unobvious pleiotropic homologies transcend the superficialities of comparative anatomy, only being revealed by knowledge of molecular developmental and phylogenetic physiologic motifs (91, Shapiro, J.; Evolution: a view from the 21st Century; FT Press Science, Upper Saddle River, New Jersey, United States, 2011.

92). The deepest of these are related to the physiologic effects of stretching, or mechanotransduction, on surfactant metabolism, which refers all the way back to biologic adaptation to gravitational force, the most ancient, omnipresent and constant of all environmental effectors of evolution (93,94).

For example, the alveolar type II (ATII) cells produce Prostaglandin E2 (PGE2) (95), particularly when they are distended (96), causing secretion of lipid substrate from lipofibroblasts for lung surfactant phospholipid production by the ATII cells (96); without PGE2, the lipids would remain bound within the lipofibroblasts. This effect of PGE2 on the secretion of Free Fatty Acids (FFAs) from lipofibroblasts is homologous with the release of FFAs from peripheral fat cells, a trait that hypothetically evolved as a consequence of the evolution of endothermy (11). To alleviate the periodic hypoxic constraints on the evolving alveolar bed, stress induced adrenalin stimulated surfactant secretion to increase gas exchange transiently until the indigenous PTHrP mechanism could generate more alveoli (97). Thus, the pleiotropic co-evolution of the PGE2 mechanism facilitating FFA utilization in both the lung and fat pad was not a chance event; it was synergistic when viewed within the context of the evolving lung’s effect on endothermy (11). In further support of this hypothesis, the role of the lung in the evolution of endothermy is further evidence for the causal evolutionary interrelationship between the pulmonary and neuroendocrine systems, both mediated by PTHrP signaling (98, 99). Yet again, this is not a chance event; periods of hypoxia due to the continuous evolution of the lung would have caused physiologic stress, stimulating adrenalin production by the adrenal medulla. Adrenalin production would have had the dual adaptive benefit of increasing alveolar oxygenation (100), and releasing FFAs from the peripheral fat pads (101). The release of excess FFAs from the fat pad would otherwise have been toxic (102), but instead adaptively increased body temperature (11), complementing the evolution of dipalmitoylphosphatidylcholine, the surface-active phospholipid in mammalian alveoli, which is 300 percent more surface-active at 37oC than at 25oC (103).

A similar physiologic evolutionary interrelationship emerges from the etiology of Goodpasture’s Syndrome. The disease is caused by an autoimmune reaction to an evolved isoform of Type IV collagen (104). Alpha 3(IV)NC1 Type IV collagen is absent from worms and flies, but appears in fish (19). However, it does not generate the pathogenic Goodpasture’s Syndrome antibody (19). It is ubiquitous in amphibians, reptiles, birds and mammals. It has the evolutionarily-relevant physicochemical characteristic of being more hydrophobic than other Type IV collagens, offering a functional role in preventing water loss across the lung and kidney epithelia in adaptation to land. The fact that this specific Type IV collagen isoform evolved during the process of land adaptation is unlikely to have occurred merely by chance, given its ability to prevent water loss on land (19).

Thus, not unlike Chemistry and Physics, Biology, is also founded on First Principles that can be understood ontologically and epistemologically rather than through dogmatic teleologic mechanisms and tautologic concepts (105). George Williams’ Antagonistic Pleiotropy hypothesis for senescence was alluded to above- in large part, this perspective is reflective of the systematic error authored by Ernst Mayr (106) that there are proximate and ultimate mechanisms of evolution that must be dissociated from one another based on Darwinian principles of mutation and selection. However, that dictum was formulated more than sixty years ago. Theorists that offered differing perspectives, such as Haeckel, Spemann and Lamarck have generally been dismissed. However, in the interim a great deal more about biology has been learned that re-energizes some previously disregarded principles towards understanding evolutionary development. This is particularly true within cell biology, where pathways can be identified that inform us that there is a continuum between the proximate and ultimate mechanisms of evolution- Mayr exemplified this principle using bird migration, which was then too complex to be understood as one continuous process, yet we now know how ambient light affects the neuroendocrine system to foster migratory behavior.

As an extension of the insights gained by seeing pleiotropy through the lens of mechanistic pleiotropy, repurposing of the same genetic signaling cascade to form novel phenotypes, heterochrony can be seen in the same way- the mechanism of heterochrony has never been provided before, it has only been described (107). Haeckel described the concept of Heterochrony as a way of expressing how development could facilitate evolutionary change (108). To this day, no one has expressed heterochrony as a mechanism for reallocating cell-cell signaling to accommodate adaptive change, yet it is the premise we have used throughout this book.

It was Thomas Kuhn, the author of The Structure of Scientific Revolutions (109), who said that an indicator of a paradigm shift was a change in the language- going from a descriptive to a mechanistic way of thinking about pleiotropy and heterochrony would reflect such a paradigm shift.

Indeed, Haeckel, Spemann and Lamarck had many correct surmises about the mechanistic biologic principles that they each addressed- recapitulation theory, the embryologic ‘organizer’, and acquired characteristics. In their time, they lacked the technical ability to support their hypotheses. However, the novel perspective on pleiotropy expressed herein honors both old concepts and new. Our own evolving understanding of evolutionary mechanisms generates a compelling narrative for evolution as a continnum of physiologic adaptations towards rewarding homeostatic mechanisms that permit cells to thrive in diverse environments.

Cells solve problems- they use the tools that they have or can generate (110). Many generations of scientists have attempted to discern the puzzle of evolutionary development, yet they have lacked the tools that can be productively employed today. What we have now learned is in many ways unexpected. Contrary to our expectation, what was old can again become new. In that sense, this paper is dedicated to those who have labored before us. Their efforts can now be married to compelling research. Through this combination, a new paradigm for evolutionary development unfurls that is congruent with the dominant truth that can be asserted about our physiologic path from First Principles. It is clearly evident that all complex organisms unavoidably must return to their unicellular roots (10, 11). The physiological pathways and the cellular communication mechanisms that underscore it explain the imperative for this immutable recapitulation.

The resolution of the evolutionary significance of pleiotropy is tantamount to Niels Bohr’s eloquent explanation for how light could be both wave and particle based on principles of Quantum Mechanics. In his Complementarity lecture at Lake Como, Switzerland in 1927 he resolved this paradoxical duality by explaining that it was an artifact of the way in which the light was measured (Bohr Como Lecture) (111). Similarly, the cell is both genetic and phenotypic, depending upon the metric, yet in reality it is integral whole whose fate is determined by the ever-transcendent mechanisms that perpetuate it (11). In his groundbreaking book Wholeness and the Implicate Order the physicist David Bohm (112) explains how our subjective senses cloud our perception of reality. As in Physics, recognizing this dichotomy is key to future progress in biology and medicine.

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