Dollo's Law: made to be broken?

Maybe it's time to retire Dollo's Law, the idea that once a trait has disappeared from a lineage, it can't reappear.  Louis Dollo was a Belgian paleoanthropologist who proposed in the late 19th century that once gone, a trait was lost forever.  Evolution could not repeat itself.  We have blogged about this before, e.g. when a paper appeared in the journal Evolution in 20011 suggesting that mandibular teeth had reappeared in a frog lineage after more than 200 million years.

Holly brought a new paper in Evolution to our attention, also detailing instances in which traits long lost have reappeared.  Most previous examples have been of hard tissue reversions, but in this paper, Rui Diogo and Bernard Wood document numerous instances of reversions to previous muscle structures in primates, and suggest what this might mean about development and evolution.

Diogo and colleagues have been involved in a long term comparative study of the anatomy of non-primate vertebrates, and of primates, looking at "homologies and evolution of the head, neck, pector and forelimb muscles of all major groups...based on dissection of hundreds of specimens and on a review of the literature".  They used this extensive data set to do parsimony and Bayesian cladistic analyses (a statistical method for classifying organisms into biologically similar groups based on whatever trait of interest) of the muscle data for primates.  They built a phylogenetic tree based on the cladistic analysis of 166  characters of head, neck, pectoral and upper limb muscles. 

...of the 220 character state changes unambiguously optimized in the most parsimonious primate tree, 28 (13%) are evolutionary reversions, and of these 28 reversions six (21%) occurred in the nodes that lead to the origin of modern humans; nine (32%) violate Dollo's law.

Without going into the anatomical details covered in the paper, suffice it to say that they found more anatomical reversions to an earlier state in head and neck muscles than in the chest or upper limb, and conclude that evolutionary reversions were significant in primate and human evolution.  Their explanation for why so many exceptions to Dollo's Law have been documented is that the developmental pathways that formed these structures were maintained over evolutionary time, perhaps because the pathways were used in the development of other structures, so that they could be readily recruited for the re-development of a once-lost trait.  Chickens, for example, still have some of the developmental pathways for teeth, although they haven't actually had teeth for 60 million years.  Some constraint on the pathways would explain its continued existence. 

It has also been found that during ontogeny, say of the hand muscles, various muscles develop that are subsequently lost as the embryo grows.  One example is the contrahentes muscle that extends to various fingers in an early human embryo, but is then lost later in development.  This is a muscle that adult chimpanzees do have, though adult humans do not.  Diogo and Wood report this same developmental story for multiple muscles.  This means that the developmental pathway has been retained, even if the specific trait has not -- 'hidden variation'. 

According to some authors, cases where complex structures are formed early in ontogeny just to become lost/indistinct in later developmental stages (the so called 'hidden variation') may allow organisms to have a great ontogenetic potential early in development, that is if there are for instance external perturbations (i.e., change in the environment, e.g., climate change, environment occupied by new species, etc.) evolution can use that potential (adaptive plasticity) (e.g, West-Eberhard 2003).

Others (Stephen J Gould, e.g.) have argued that rather than an argument for plasticity, this means that evolution is constrained, contingent on what is already there: the embryo couldn't develop properly if these pathways to nowhere were to change.  Some argue that hidden variation is not responsible for evolutionary novelty, though, as Diogo and Wood suggest, it can explain the reappearance of traits.

In essence, Dollo's "law" is a principle that something genetically complex is difficult to undo because mutation will remove order if not opposed by some form of selection.  The more steps removed, the more 'canalized' a trait would tend to be and the less flexible.  Indeed, 'canalized' is a word used early in the 20th century by CH Waddington for the persistence of fundamental traits.   Reversals may also be apparent rather than real: new mechanisms might bring about a similar appearance at the trait level without being a literal 'reversal'.  The nature of evolution is to avoid being put in a box out of which organisms couldn't evolve.

So whatever the explanation for these specific instances of violations of Dollo's Law, it's clear yet again that any evolutionary law is made to be broken.

Reversibility: Dollo's 'law' in development and evolution

We recently posted on reports of the re-evolution of traits that had long been lost in evolutionary time.  This seemed to violate Dollo's "law" that evolution was a one-way train that couldn't back up.

When there are people in a population with or without a particular trait, say, eye color, and their children have a different version, we are not perplexed. Contingencies of gene expression or genotype or alleles (genetic variants) in the population can make this happen.  Darwin tried to fit his idea of inheritance with these ideas, mainly by hand-waving.  Now armed with concepts like multi-gene control and recessiveness of alleles, we have no problem understanding how these things happen.

But there are reasons to think that true reversibility of complex traits can't often happen over evolutionary time.  The basis of the argument is that too many genetic changes are required for a complex trait to be constructed and if the trait is 'erased' by mutation (and that is supported in the face of selection), then over time too many other genetic changes (mutations, gene duplications, other uses of genes, etc.) will make it impossible to back-track.

From this point of view, the 'new' version of the trait is physically similar to the old, or to that in a widely distant species, but is due to selection for the same trait that happens to pick up different genes and alleles to get the job done.  But if a pathway has been conserved because it's used for other things in the organism, it may be that simple genetic changes can reactivate that pathway in a context in which it was active long ago.

There has been a similar kind of no-going-back dogma, a kind of Dollo's "law" in developmental genetics.  Stem cells can differentiate into anything, but once that happens, the differentiated cells simply cannot go back to being stem cells.  We now know that this is not accurate. Even a small number of genetic changes in experimental systems can restore various stem-cell states.  This can happen even if the cell being manipulated is highly differentiated.  Is this as surprising or inexplicable as reversals in evolution?

The answer is that it is far less surprising, as a generality.  With some few notable exceptions, all the cells in your body have the same genome. This means that while each cell is of a particular type largely because it uses a specific set, but not all, of the genes in the genome.  There are, so to speak, 'blood' genes, 'stomach' genes, and so on. Gene expression is based on the physical packaging of chromosome regions and the presence of proteins specific to the cell type, that bind to DNA in regions near to, and that cause the expression of the specifically used genes.  But since with few exceptions all the genes still exist in all cells, if one changed these regulatory traits (packaging and so on of DNA, presence of regulatory proteins), one could make the cell do something else.  There may be too many changes in expression needed to make a stomach cell into a lung or muscle cell on its own, but we're looking at cells from the outside, and cells can be engineered to redifferentiate or dedifferentiate by experimentally imposing required sets of change. And in a sense it's why in some instances it only takes about 4 genes being manipulated to bring cells back to a very primitive stem cell type of state.

Evolution is different,  because once a species is committed to a particular direction, its genes themselves as well as their usage have changed, by virtue of mutation and frequency change induced by chance or natural selection.  Thus, spiders and grasshoppers no longer have the same genes so that only the expression pattern would need to be changed to let spiders hop or grasshoppers spin webs.  That is why evolution rarely truly reverses. Sometimes only a few changes would be needed, if basic pathways still exist but have been mutationally inactivated.

On the other hand, most traits have many paths and most genes have many uses, so that there can be many different paths by which some absent trait--or its likeness!--can reappear.  Natural selection and chance could activate some suitable set of genes to make this happen, and how likely it is depends on what environmental constraints are.  And since many developmental genes are highly conserved over long time periods, there can easily be similarities in the genetic basis of reappearance.

We know that some traits, such as complete vs incomplete metamorphosis in some species of amphibians (i.e., whether or not they go through a larval stage), or the pattern of ocelli (middle eyes) in insects, have re-evolved.  And we know that some genes from mammals can induce similar effects even in insects, by replacing or over activating their corresponding insect gene.

So, reversals are of many types due to many causes.  How likely they are, and how genetically they are brought about, are statistical and context-specific questions.  But there are no real mysteries about whether or not they are possible.

Lost and found -- Breaking Dollo's Law

It has long been thought that once a species loses a trait, that trait is gone forever.  The loss can't be reversed.  This is so well-accepted that it's risen to the status of a law of nature, Dollo's Law.  Dollo was a Belgian paleontologist working around the turn of the last century, and as he put it,  "An organism is unable to return, even partially, to a previous stage already realized in the ranks of its ancestors."  So, for example, according to Dollo's Law, we have lost the tail our ancestors had, and we won't regain it, reptiles that have made the transition from egg-laying to giving birth to live offspring can't go back, and so on.

From the BBC

But laws are made to be broken, and the idea that life follows 'laws' the way gravity and chemistry do is tenuous at best.  In this case, Dollo's 'Law' is currently under challenge.  A paper published online in Evolution on Jan 27, and discussed on the BBC website, describes the re-evolution of mandibular teeth in frogs after more than 200 million years.  Recent reports of exceptions to the irreversibility principle -- wings lost and regained in stick insects, the re-evolution of coiling in snail shells, re-evolved ocelli in cave crickets, wings in water spiders, and so forth -- have been called into question for methodological reasons, but the re-evolution of frogs teeth and other examples look pretty solid.  As Wiens concludes,

The results presented here offer an incontrovertible phylogenetic example of trait re-evolution, showing that mandibular teeth were lost in the ancestor of all living frogs and then re-evolved in the hemiphractid species G. guentheri. The alternate hypothesis, that mandibular teeth were lost independently in each of the dozens of lineages leading up to G. guentheri, is statistically unsupported and seems incredibly unlikely. Although the hypothesis that G. guentheri re-evolved mandibular teeth may be unsurprising to experts in amphibian anatomy, this compelling example has been ignored in the recent literature on Dollo's law. Further, this example is made remarkable by the application of a time scale for this event: mandibular teeth were absent for at least 225 million years (and likely much longer) before being regained.

Another example of an exception to Dollo's Law is described by Lynch and Wagner in the Jan 2010 issue of Evolution.  They present evidence for the re-evolution of oviparity, or egg laying, in "Old World sand boas in the genus Eryx nearly 60 million years after the initial boid transition to viviparity" based on a phylogenetic analysis of genetic data from boid snakes and other related groups.  In addition to the statistical support for re-evolution, they note that morphological evidence includes the fact that, like live born boas, the hatchlings of the "oviparous Eryx lack an egg-tooth providing independent evidence that oviparity is a derived state in these species."  And, the shells of the boas that have re-evolved oviparity are extremely thin compared with the shells of other boas, suggesting that the structure of the eggshell re-evolved as well.

The argument over whether Dollo's Law can be violated may primarily reflect the way we classify species and think about phylogenies.  As Wiens says,

...the observation that most well-documented cases of trait re-evolution occur after a period of trait loss of greater than 15 million years (Table 2) may also reflect methodological bias. A complex structure that re-evolves may need to be absent for tens of millions of years before its re-acquisition can be confidently distinguished from multiple losses (e.g., given typical diversification rates, this period of time may be needed for a group to diversify enough to have species nested deep in the phylogeny). Yet, if the gain of lost traits is possible, a consideration of the underlying genetics suggests that it should be much more likely soon after the trait is lost (Marshall et al. 1994). Thus, trait re-evolution may actually be hardest to detect under the conditions when it is most likely to occur, raising the question of whether trait re-evolution might be more widespread but frequently undetected due to methodological biases.

So, how is the re-evolution of a trait explained genetically?  Weins suggests that the fact that G. Guentheri still have teeth in their upper jaw facilitates the repositioning of teeth to the lower jaw.  The structural genes are there, they just need to be recruited for expression in a different place.  Generally, this is not very perplexing.  Single gene expression changes can lead to signaling environments that, in relevant tissues, can induce cascades of patterning-gene interactions that produce structures like teeth.

Indeed, the normal tissue is not required.  There are mouse and human examples in which hair or teeth grow where the other normally does.  Ovarian teratomas are disorganized tumor-like structures of disorganized embryos that, as was known even in the early 1800's (and cited by Darwin), developed hair and teeth (even chicks do not have enamel genes, but teeth can be induced to form by genetically similar processes as hair or feathers, and transgenic expression of some environment-preparing genes in embryonic chick jaws can lead to tooth-like structures).

Dental formulas (numbers and shapes of teeth in upper and lower jaws) and hair or coloration patterns can come and go for similar reasons.  In many cases, the genetic change can be a modification of existing genes and tissues.  Similar dental formulas have evolved numerous times in vertebrates.  In others, there are different pathways to similar outcomes.  This is what happens in some selection experiments, and seems, if we recall correctly, to be the case with ocelli in insects.  But the overall pathways are presumably rather simple, or the different activations activate similar pathways that are conserved because they are still doing something.  In the frogs, they're making teeth in the upper jaw, a tissue histologically similar to lower jaws.

The point here is not that reversals are trivial or uninteresting, but they are explicable when they occur.  However, highly organized or complex structures are unlikely to recur after too long of divergence.  If we re-evolved, say, swimming anatomy it would not likely be the same as what is found in fish.  Indeed, that is what whales did, and they swim by up and down motion rather than the side to side motion of fish, because their mechanism is anatomically different.

Again, these instances are interesting, and perhaps most interesting is that even before the genetic age anyone should have suggested, much less canonized, 'laws' about how evolution works.  At most they are statistical generalizations that, in genetic terms, relate to the likelihood of the same genetic mechanism re-evolving.

Evolution to and fro

Dollo's Law
In the last bit of tidying up after our trip, Holly asked us to comment on a recent paper in Nature (An epistatic ratchet constrains the direction of glucocorticoid receptor evolution, Bridgham et al., 461, 515-519, 24 September 2009), related to the question of whether evolution can, or does, ever go backwards. This is a long-standing question among evolutionary theorists; indeed, in 1890, Belgian paleontologist Louis Dollo addressed it by proposing what is now known as Dollo's Law: "An organism is unable to return, even partially, to a previous stage already realized in the ranks of its ancestors."

But, what does it mean to ask whether evolution can reverse itself? As Bridgham et al. themselves point out, there are many ways to make a given trait, not all genetically equivalent. So, is evolutionary reversal the replication of an older form, which wouldn't necessarily have to involve the same genes as the original iteration? Or, does it mean replication of the same form, produced by the same genes? The first, reproduction of an ancestral form in a novel way, is much more plausible than the second, given the apparent strength of contingency and cooperation in development; what gets made next depends on what's here now, and that depends on what was just here. Thus, to reproduce an ancestral form would require that each step in the evolution of a protein be reversed, along with each step in the evolution of the proteins with which it works. And, because, say, a receptor cooperates with a ligand, although imprecise receptor/ligand binding often works, neither can change too drastically too quickly, lest the signal telling the cell what to do next won't get sent. So, reversal of form and DNA sequences would require reversal of each of the co-evolutionary steps that led to the modern form, which seems highly implausible.

In addition, reversal to a previous form, via the same genes or not, would require the return of the environment in which that form was once successful, and that makes it that much more implausible, since environments are always changing. As Bridgham et al. say, "The past is difficult to recover because it was built on the foundation of its own history, one irrevocably different from that of the present and its many possible futures." So, because of so much that is known about biology, we would a priori agree with Dollo's Law.

All that said, Bridgham et al. actually put the question to a test. They engineered a hormone receptor protein to mimic its ancestral state and the steps it took to become the modern receptor it is, and evaluated its function along the way. In its ancestral state, the receptor was 'promiscuous', able to bind with different classes of hormones, but over 40 million years, and with an estimated 37 different amino acid changes, it lost that ability and became specific to a single class of hormone. The authors found that only 2 amino acid changes were necessary for the receptor's evolved specificity but that subsequent amino acid changes that "optimized the new specificity of the glucocorticoid receptor, also destabilized elements of the protein structure that were required to support the ancestral conformation." Thus, building in just the 2 original changes wasn't sufficient to allow them to resurrect the receptor's ancestral function. They conclude that too many changes are required for successful reversal of the receptor to its ancestral function. Does this prove that evolutionary reversal is impossible? No, but it does suggest that contingency and cooperation are indeed foundational principles in development and evolution, and are important reasons why reversal is unlikely.

How unlikely?
Perhaps the points can be more clearly and immediately seen if we ask how probable rather than plausible they are. A mutation replacing an A with a T in DNA can be reversed, the T being replaced with an A--changes that do occur. But because in general mutation is very improbable at any given spot in DNA, the same specific mutation is even less probable. And even if that chance were, say, 1/1000 (much much greater than is actually the case), if 100 of these reversals were needed, the chance would be 1/1000 to the hundredth power, infinitessimally small. And this doesn't take account of the order, viability of intermediate stages in the reversal process, etc. So while most things like this might be possible, they are too unlikely to take seriously if we're thinking about anything at all complicated.

And we can seal the 'no' deal in two other ways. First, we already know that things can reverse. People who have a certain height can have shorter children but their children could again be taller. Much of the time this will be due to different genotypes.

And many mutations involve deletions of DNA, sometimes of chunks many nucleotides long. It would take a fairy godmother to wave a wand to reverse this and somehow conjure up the exact chunk to be re-inserted some time later.

Darwinism as natural law
Here's another way to think of it. Evolutionary theory, since Darwin, has attempted to be a natural science based on natural law. Darwin was very Newtonian in this, suggesting that natural selection was a kind of 'force', rhetoric often used today even though we have a strong sense of probabilism (including both mendelian sampling of genes from parent to offspring, and genetic drift in which the frequency of genetic variants changes in a population strictly by chance).

In a perfectly Newtonian world, nature is predictable and retrodictable. If you know the state today you can predict tomorrow, or tomorrow you can predict today (e.g., by changing the sign of the equation from plus to minus, so to speak). This was an ideal until the 20th century, roughly speaking, when disturbances such as quantum theory showed that things were not so uniformly homogeneous in time.

Largely stimulated by Darwinian thinking, even physicists began to realize that time had a direction. If change is probabilistic, we can go from today to some state tomorrow, but tomorrow we can't tell what today's state had been.

In principle, if we compare DNA sequences between species, we see that evolution diverges forward in time, as mutational changes occur in different descendant copies of a gene from generation to generation, producing a branching or tree-like structure of sequence relationships. These are presumably related to traits, like fingers and leaves, and natural selection (and drift) produce differentiated, adaptive organisms over evolutionary time.

In a sense, this would not seem to be reversible. Once you can make a limb (on a tree or on your body), it is so complex a process that you can destroy it but you can't go back to the state before it was a limb.

However, if you just look at the nucleotides, as we said above, traits of organisms involve many different genes, and the probability of everything being exactly reversed is trivially small, even if mathematically possible.

If you want to be a stickler for exactness then you have the answer: reversal is technically possible but in practice impossible. But if you look at traits or function, then evolution certainly reverses itself. That's what happened in the evolution of flightless birds. Different lineages of insects have repeatedly evolved, or de-evolved, similar states related to the number of ocelli (small central eyes), and different lineages of amphibians have gained or lost tadpole stages in their development. Some of these may involve reverse mutations in the same genes, but there are undoubtedly different genetic pathways 'forward' as well as 'backward' in this phenotypic sense.

As so often is the case, the answer depends on the question. Dollo's principle seems reasonable (if not a 'law' in the cosmic sense) in regard to complex adaptive traits. Different versions of what seem to be the 'same' trait usually have at least some genetic differences. In that purist sense, you really can't go home again.

-Ken and Anne