It’s just a stage. A phylotypic stage. Part III: Fish and more

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ResearchBlogging.orgGiven that disputes over the existence and meaning of the phylotypic stage and the hourglass model have simmered in various forms for a century and a half, the remarkable correspondence between the hourglass model and gene expression divergence discovered by Kalinka and Varga and colleagues would be big news all by itself. But amazingly, that issue of Nature included two distinct reports on the underpinnings of the phylotypic stage. The other article involved work in another venerable model system in genetics, the zebrafish.

The report is titled "A phylogenetically based transcriptome age index mirrors ontogenetic divergence patterns" and is co-authored by Tomislav Domazet-Loso and Diethard Tautz. To understand how their work has shed light on the phylotypic stage and the evolution of development, we’ll need to look first at an approach to the analysis of evolutionary genetics that these two scientists pioneered: phylostratigraphy.

The authors first described phylostratigraphy in 2007 and have since used the approach to examine genes that cause human genetic disease and cancer. They define it as:

a statistical approach for reconstruction of macroevolutionary trends based on the principle of founder gene formation and punctuated emergence of protein families.

The idea is that every gene has a birthday, a point at which it is first identifiable in evolutionary history. Some genes are ancient, having arisen before there were even complex cells, and others are relative juveniles, having arisen much more recently. Genes present today, then, can (in principle) be assigned an "age." Domazet-Loso and Tautz represent the "age" of a gene by the evolutionary "epoch" in which it appeared, by analogy with the identification of the appearance of biological lineages with stratigraphic epochs in earth’s history. So for example, some genes appear with the development of true animals (metazoa), and so these genes are assigned to that "stratum" of biological history. In fact, the authors call each epoch a ‘phylostratum’ to reinforce that metaphor.

So how does this work? To do phylostratigraphic analysis, you need two major sets of tools. First, you need a pretty solid phylogeny, or family tree, of your organism(s) of interest. Second, you need complete or nearly-complete genome sequences of the organism of interest and of organisms that can represent the major branch points (or nodes) in the family tree. The procedure from there seems clear enough: using a well-known alignment program, you search through the family tree for each of the genes in your organism of interest, to see where it is first recognizable in the phylogeny. That point is the phylostratum from which that gene arises. With that data, you could look at the contributions of various phylostrata to various body parts or processes. Or conversely, you could look at the relative age of the sets of genes associated with those body parts or processes. Or you could look at the relative age of the sets of genes associated with different stages of development. And that’s what Domazet-Loso and Tautz did in their Nature paper on the hourglass model.

Specifically, the authors took their phylostratigraphic data and merged it with expression data at various stages of zebrafish development; they called the resulting parameter the transcriptome age index (TAI). Basically, they calculated a relative age of the genes that are turned on at each stage of development, corrected for the extent to which particular genes are being used at those stages. Then they mapped the TAI onto the timeline of zebrafish development. And this is what they saw.

PhylostratigraphicHourglassFig1a-700px.gif

Does that look familiar? Like, say, half an hourglass? In the earliest stages of development, active genes are young-ish, as they are in the juvenile and the adult. In between, the genes that are active are older – a lot older. And the low point, where genes are oldest? It’s the end of segmentation and the beginning of the pharyngula stage. That’s the stage that is considered the phylotypic stage in vertebrates. (What this has to do with godless liberalism, I have no idea.) And so we see that hourglass again, this time traced out by the evolutionary age of the genes that are active during the phylotypic stage.

As you look at the graph, you might notice some other interesting periods in the life of a fish. There’s a prominent peak of gene youthfulness at 6 hours of development; this corresponds to gastrulation, that wonderful time in your life when you established yourself as a three-layered animal. That peak is due to the activation of a lot of animal-specific genes, namely those that date to the metazoan phylostratum. This includes genes that control cell-cell interactions, certainly a hallmark of animal-building. Those might seem like incredibly basic functions, but they’re relatively young compared to even more basic cellular processes, and the genes that control those processes are the ones that predominate during the later phylotypic stage. (The authors showed, in fact, that extremely ancient genes are active uniformly throughout development, whereas the younger gene sets display the hourglass pattern: high-low-high.)

And notice that gene youthfulness declines during aging (after adulthood). Now why would that be? The authors propose that the most recent innovations (facilitated by relatively young genes) are likely to have resulted from adaptation, and so:

The fact that ageing animals revert to older transcriptomes is in line with the notion that animals beyond the reproductive age are not ‘visible’ to natural selection and can therefore not be subject to specific adaptations any more.

There’s a lot more: the study found differences between males and females (look at the dotted lines in the figure), for example. But they also extended their analysis to other animals with known genomes: fruit fly, roundworm and mosquito. In every case they saw the same pattern: young-old-young. Their fly graph displays a pattern strikingly similar to that in the fish, and nicely dovetails with the distinct analysis done by Pavel Tomancak’s group:

PhylostratigraphicHourglassFig3aFly-700px.gif

Look at the low point, where the genes are the oldest. It’s the germband elongation stage – the recognized phylotypic stage for insects, and the same point singled out in the fly paper. Remarkable.

So to summarize, the two papers, reported separately but simultaneously, strongly support the hourglass model of development, in which embryos are seen to converge on an evolutionarily-ancient form, after diverse beginnings and followed by radical divergence into the wonderful variety of animals seen today and in the past. Domazet-Loso and Tautz explain how these new results make sense of the hourglass:

These consistent overall patterns across phyla, as well as the detailed analysis within zebrafish, suggest that there is a link between evolutionary innovations and the emergence of novel genes. Adaptations are expected to occur primarily in response to altered ecological conditions. Juvenile and adults interact much more with ecological factors than embryos, which may even be a cause for fast postzygotic isolation. Similarly, the zygote may also react to environmental constraints, for example, via the amount of yolk provided in the egg. In contrast, mid-embryonic stages around the phylotypic phase are normally not in direct contact with the environment and are therefore less likely to be subject to ecological adaptations and evolutionary change.

And as they note, Darwin himself made this connection, reflecting on von Baer’s earlier observations. Ideas, like genes, can have a long and productive history.

[Cross-posted at Quintessence of Dust.]

———-

Domazet-Lošo, T., & Tautz, D. (2010). A phylogenetically based transcriptome age index mirrors ontogenetic divergence patterns. Nature, 468 (7325), 815-818. DOI: 10.1038/nature09632

17 Comments

I’m putting together an overview lecture of basic embryology ideas and how they represent yet another line of evidence for evolution over winter break. What wonderful data to have to incorporate. Right up there with the Neanderthal Genome papers.

There seem to be two explanations around. One is the one you quote from Domazet-Loso and Tautz about the phylotypic stages being those that are in less contact with the environment, hence less modified by natural selection. The other attributes the paucity of modifications of that stage to the multiple effects of modifications early in development, so that these tend to disrupt development more than later modifications. Thus, the argument goes, most modifications of the embryo by natural selection will tend to be late ones.

Is there anything in these recent papers that argues for one of these explanations or the other?

Joe Felsenstein said:

There seem to be two explanations around. One is the one you quote from Domazet-Loso and Tautz about the phylotypic stages being those that are in less contact with the environment, hence less modified by natural selection. The other attributes the paucity of modifications of that stage to the multiple effects of modifications early in development, so that these tend to disrupt development more than later modifications. Thus, the argument goes, most modifications of the embryo by natural selection will tend to be late ones.

Is there anything in these recent papers that argues for one of these explanations or the other?

Well, first of all, it’s the hourglass that needs to be explained, and the “don’t-mess-with-early-development” answer is therefore incomplete, since very early development is evidently more malleable than the later phylotypic stage. But yes, both papers advance a version of that hypothesis. Here is the final sentence by Domazet-Loso and Tautz:

the phylotypic phase can be defined as the ontogenetic progression during which the oldest gene set is expressed, either because this is the phase with the lowest opportunity for lineage-specific adaptations, or because it is internally so constrained that newly evolved genes cannot become integrated.

They earlier refer to this as the “constraint hypothesis,” and I think it’s the competing explanation to which you refer. Kalinka and Varga et al. document selective constraint during the phylotypic period, but they don’t seem to connect this explicitly to contact with the environment, and instead discuss increased global interactions and gene regulatory network topology. See their final paragraph and/or the supplemental discussion (final section of the supplemental material, freely available online).

Yet again another independent data set and independent type of analysis and yet another stunning confirmation of the predictions of evolutionary theory. This is one of the most remarkable results that I can remember seeing for avery long time. Also another beautiful example of evo/devo in the age of genomics, coupled with phylogenetics.

Now of course the creationists have no explanation whatsoever for any of these observations. Nor did they do any of the research that led to these discoveries. If they thought that this pattern would not be found, why didn’t they try to disprove it themselves. This is yet another data set that can instantly be incorporated in the fight against creationism. I’m sure the authors would be proud to see it used for that purpose.

Now there is still an obnoxious concern troll who is going around derailing threads. He apparently cannot stand for real science to be discussed by anyone. He will no doubt show up on this thread and whine and moan about how no one is discussing the science, or some such equally transparent nonsense. Until Steve can purge the thread of this type of pollution, everyone should ignore the troll and let him starve on a diet of his own vomit.

It would seem that the simplest way to explain the hourglass, is that the relative difficulty of modifying early development is essentially correct. The reason that the earliest development is divergent, is because you have to close the loop so to speak. As animals diverge as adults and their reproductive strategies and environments diverge the starting points for development become more divergent themselves, and what you see then is selection acting to funnel that variability back into the “standard” program. At a first pass that explanation would suggest that the width of the bottom of the hourglass would correlate well with the speciation rate for the taxon, while if it was the environment in general it would tend to correlate somewhat better with absolute divergence time. Whether or not we have a model system at this moment that could give enough data to distinguish that fine a level of detail is another matter.

Another interesting thing about this article is the method for determining the age of genes. Apparently new genes arise all of the time, undoubtedly through a process of gene duplication followed by random mutation and natural selection. This of course causes an increase in information, which creationists claim cannot happen. As Joe Felsenstein correctly points out, gene duplication is not necessary in order for information to increase, it simply represents an incontrovertible example with which to confront creationist nonsense.

Now why would god create everything in six days and then keep adding genes periodically every few years. Doesn’t seem to make much sense for her to do it that way.

This comment has been moved to The Bathroom Wall.

This comment has been moved to The Bathroom Wall.

DS said:

Another interesting thing about this article is the method for determining the age of genes. Apparently new genes arise all of the time, undoubtedly through a process of gene duplication followed by random mutation and natural selection. This of course causes an increase in information, which creationists claim cannot happen. As Joe Felsenstein correctly points out, gene duplication is not necessary in order for information to increase, it simply represents an incontrovertible example with which to confront creationist nonsense.

Now why would god create everything in six days and then keep adding genes periodically every few years. Doesn’t seem to make much sense for her to do it that way.

DS, you’re right that this is a major focus of work by Domazet-Loso and Tautz in general, and I’ll write soon about that topic by itself. Ideas about gene origins are indeed a way to answer creationist objections to evolution. But your last little comment there is pretty dumb: for one thing, a deity could do anything he/she likes and certainly might add genes anytime she/he wants. It seems that the comment is troll-bait (demonstrably effective, to my annoyance), and you can take that to another thread. (Where, clearly, the silliness originated.)

Steve Matheson said:

Joe Felsenstein said:

There seem to be two explanations around. One is the one you quote from Domazet-Loso and Tautz about the phylotypic stages being those that are in less contact with the environment, hence less modified by natural selection. The other attributes the paucity of modifications of that stage to the multiple effects of modifications early in development, so that these tend to disrupt development more than later modifications. Thus, the argument goes, most modifications of the embryo by natural selection will tend to be late ones.

Is there anything in these recent papers that argues for one of these explanations or the other?

Well, first of all, it’s the hourglass that needs to be explained, and the “don’t-mess-with-early-development” answer is therefore incomplete, since very early development is evidently more malleable than the later phylotypic stage.

Agreed, and I misstated the hyothesis, which I represented more correctly in my comment on Part I of your posts on the phylotypic stage work.

But yes, both papers advance a version of that hypothesis.

They earlier refer to this as the “constraint hypothesis,” and I think it’s the competing explanation to which you refer.

Yes.

Kalinka and Varga et al. document selective constraint during the phylotypic period, but they don’t seem to connect this explicitly to contact with the environment, and instead discuss increased global interactions and gene regulatory network topology. See their final paragraph and/or the supplemental discussion (final section of the supplemental material, freely available online).

So I guess the overall answer to the question I raised is no, they don’t present any data that is capable of choosing between these hypotheses?

Steve: Kalinka and Varga et al. document selective constraint during the phylotypic period, but they don’t seem to connect this explicitly to contact with the environment, and instead discuss increased global interactions and gene regulatory network topology. See their final paragraph and/or the supplemental discussion (final section of the supplemental material, freely available online).

Joe: So I guess the overall answer to the question I raised is no, they don’t present any data that is capable of choosing between these hypotheses?

No, they don’t, and they both note this. Domazet-Loso and Tautz make it clear in their final paragraph quoted above, and here’s the last sentence from Kalinka and Varga et al.:

Future studies will also need to address the mode and strength of selection acting on gene expression with greater resolution by coupling interspecific expression divergence with intraspecific variation during embryogenesis.

Do you think that Kalinka and Varga et al. are right about this? Or are there other ways to address the question?

Steve Matheson said:

me: So I guess the overall answer to the question I raised is no, they don’t present any data that is capable of choosing between these hypotheses?

No, they don’t, and they both note this. Domazet-Loso and Tautz make it clear in their final paragraph quoted above, and here’s the last sentence from Kalinka and Varga et al.:

Future studies will also need to address the mode and strength of selection acting on gene expression with greater resolution by coupling interspecific expression divergence with intraspecific variation during embryogenesis.

Do you think that Kalinka and Varga et al. are right about this? Or are there other ways to address the question?

Well, no. This starts to connect with evolutionary quantitative genetics and what can be inferred from what, and it happens that I am working in this area now.

Basically, from between-species changes combined with within-species variation, you can make some (quite noisy) inferences about which directions selection has acted in. But that does not help you tell whether changes in late embryogenesis occurred because they was less resistance to them because they were less constrained by interactions with other genes, or because there was more selective pressure on them because the embryo was more in contact with the environment. Or even both.

(I tend to be skeptical of the latter, at least for mammals, because the embryos are quite isolated from the environment until birth).

Joe: Basically, from between-species changes combined with within-species variation, you can make some (quite noisy) inferences about which directions selection has acted in. But that does not help you tell whether changes in late embryogenesis occurred because they was less resistance to them because they were less constrained by interactions with other genes, or because there was more selective pressure on them because the embryo was more in contact with the environment. Or even both.

(I tend to be skeptical of the latter, at least for mammals, because the embryos are quite isolated from the environment until birth).

I surmise that this is where the phylostratigraphy approach could help a lot. Genes that date to the origin of the placenta (the eutherian phylostratum) might be expected to depart from the hourglass pattern (of expression or expression divergence) the way the earlier phylostrata do (Figure 1b of the Domazet-Loso and Tautz paper).

(I tend to be skeptical of the latter, at least for mammals, because the embryos are quite isolated from the environment until birth).

Aside from the mother’s diet?

Henry J said:

me:

(I tend to be skeptical of the latter, at least for mammals, because the embryos are quite isolated from the environment until birth).

Aside from the mother’s diet?

That of course is an outside influence, but is it greater late in embryonic development than in the middle of embryonic development? I would guess not.

Of course your guess is as good as mine, until we have some empirical evidence.

That fly graph seems to show a big difference in the age of the genes used by adult males (young) and females (old). Do they give an explanation for this?

Using the quote right about the fly chart as a jumping-off point, I would tentatively think that this implies that adult males undergo much stronger selection (maybe not natural selection - could be sexual by females, or some other sort???) than females. But I could easily be getting that wrong.

Yet again another independent data set and independent type of analysis and yet another stunning confirmation of the predictions of evolutionary theory. This is one of the most remarkable results that I can remember seeing for avery long time. Also another beautiful example of evo/devo in the age of genomics, coupled with phylogenetics.

IANAS and may be excused for not having been aware of the prediction.

But I find it a beautiful example of the continuing process of revealing the intriguing complexity of the simple, easily accessible theory that Darwin founded 150 years ago!

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This page contains a single entry by Steve Matheson published on December 18, 2010 1:56 AM.

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