Putting the cart before the… snake?

Time to reexamine some assumptions (again)! And also, talk about Hox genes, because do I even need a reason?

Hox genes often come up when we look for explanations for various innovations in animal body plans – the digits of land vertebrates, the limbless abdomens of insects, the various feeding and walking and swimming appendages of crustaceans, the strongly differentiated vertebral columns of mammals, and so on.

Speaking of differentiated vertebral columns, here’s one group I’d always thought of as having pretty much the exact opposite of them: snakes. Vertebral columns are patterned, among other things, by Hox genes. Boundaries between different types of vertebrae such as cervical (neck) and thoracic (the ones bearing the ribcage) correspond to boundaries of Hox gene expression in the embryo – e.g. the thoracic region in mammals begins where HoxC6 starts being expressed.

In mammals like us, and also in archosaurs (dinosaurs/birds, crocodiles and extinct relatives thereof), these boundaries can be really obvious and sharply defined – here’s Wikipedia’s crocodile skeleton for an example:

In contrast, the spine of a snake (example from Wikipedia below) just looks like a very long ribcage with a wee tail:

Snakes, of course, are rather weird vertebrates, and weird things make us sciencey types dig for an explanation.

Since Hox genes appear to be responsible for the regionalisation of vertebral columns in mammals and archosaurs, it stands to reason that they’d also have something to do with the comparative lack of regionalisation (and the disappearance of limbs) seen in snakes and similar creatures. In a now classic paper, Cohn and Tickle (1999) observed that unlike in chicks, the Hox genes that normally define the neck and thoracic regions are kind of mashed together in embryonic pythons. Below is a simple schematic from the paper showing where three Hox genes are expressed along the body axis in these two animals. (Green is HoxB5, blue is C8, red is C6.)


As more studies examined snake embryos, others came up with different ideas about the patterning of serpentine spines. Woltering et al. (2009) had a more in-depth look at Hox gene expression in both snakes and caecilians (limbless amphibians) and saw that there are in fact regions ruled by different Hoxes in these animals, if a little fuzzier than you’d expect in a mammal or bird – but they don’t appear to translate to different anatomical regions. Here’s their summary of their findings, showing the anteriormost limit of the activity of various Hox genes in a corn snake compared to a mouse:


Such differences aside, both of the above studies operated on the assumption that the vertebral column of snakes is “deregionalised” – i.e. that it evolved by losing well-defined anatomical regions present in its ancestors. But is that actually correct? Did snakes evolve from more regionalised ancestors, and did they then lose this regionalisation?

Head and Polly (2015) argue that the assumption of deregionalisation is a bit stinky. First, that super-long ribcage of snakes does in fact divide into several regions, and these regions respect the usual boundaries of Hox expression. Second, ordinary lizard-shaped lizards (from which snakes descended back in the days of the dinosaurs) are no more regionalised than snakes.

The study is mostly a statistical analysis of the shapes of vertebrae. Using an approach called geometric morphometrics, it turned these shapes from dozens of squamate (snake and lizard) species into sets of coordinates, which could then be compared to see how much they vary along the spine and whether the variation is smooth and continuous or clustered into different regions. The authors evaluated hypotheses regarding the number of distinct regions to see which one(s) best explained the observed variation. They also compared the squamates to alligators (representing archosaurs).

The results were partly what you’d expect. First, alligators showed much more overall variation in vertebral shape than squamates. Note that that’s all squamates – leggy lizards are nearly (though not quite) as uniform as their snake-like relatives. However, in all squamates, the best-fitting model of regionalisation was still one with either three or four distinct regions in front of the hips/cloaca, and in the majority, it was four, the same number as the alligator had.

Moreover, there appeared to be no strong support for an evolutionary pattern to the number of regions – specifically, none of the scenarios in which the origin of snake-like body plans involved the loss of one or more regions were particularly favoured by the data. There was also no systematic variation in the relative lengths of various regions; the idea that snakes in general have ridiculously long thoraxes is not supported by this analysis.

In summary, snakes might show a little less variation in vertebral shape than their closest relatives, but they certainly didn’t descend from alligator-style sharply regionalised ancestors, and they do still have regionalised spines.

Hox gene expression is not known for most of the creatures for which vertebral shapes were analysed, but such data do exist for mammals (mice, here), alligators, and corn snakes. What is known about different domains of Hox gene activation in these three animals turns out to match the anatomical boundaries defined by the models pretty well. In the mouse and alligator, Hox expression boundaries are sharp, and the borders of regions fall within one vertebra of them.

In the snake, the genetic and morphological boundaries are both gradual, but the boundaries estimated by the best model are always within the fuzzy boundary region of an appropriate Hox gene expression domain. Overall, the relationship between Hox genes and regions of the spine is pretty consistent in all three species.

To finish off, the authors make the important point that once you start turning to the fossil record and examining extinct relatives of mammals, or archosaurs, or squamates, or beasties close to the common ancestor of all three groups (collectively known as amniotes), you tend to find something less obviously regionalised than living mammals or archosaurs – check out this little figure from Head and Polly (2015) to see what they’re talking about:


(Moving across the tree, Seymouria is an early relative of amniotes but not quite an amniote; Captorhinus is similarly related to archosaurs and squamates, Uromastyx is the spiny-tailed lizard, Lichanura is a boa, Thrinaxodon is a close relative of mammals from the Triassic, and Mus, of course, is everyone’s favourite rodent. Note how alligators and mice really stand out with their ribless lower backs and suchlike.)

Although they don’t show stats for extinct creatures, Head and Polly argue that mammals and archosaurs, not snakes, are the weird ones when it comes to vertebral regionalisation. For most of amniote evolution, the norm was the more subtle version seen in living squamates. It was only during the origin of mammals and archosaurs that boundaries were sharpened and differences between regions magnified. Nice bit of convergent/parallel evolution there!



Cohn MJ & Tickle C (1999) Developmental basis of limblessness and axial patterning in snakes. Nature 399:474-479

Head JJ & Polly PD (2015) Evolution of the snake body form reveals homoplasy in amniote Hox gene function. Nature 520:86-89

Woltering JM et al. (2009) Axial patterning in snakes and caecilians: evidence for an alternative interpretation of the Hox code. Developmental Biology 332:82-89

Wherein scientists DON’T spill blood over a Precambrian animal

Having gone through much of my backlog, I was going to post about pretty blue limpet shells, then I saw that people have been arguing over Haootia. You remember Haootia? It’s that Precambrian fossil with probable muscle impressions that looks kind of like a modern-day staurozoan jellyfish (living staurozoan Haliclystus californiensis by Allen Collins, Encyclopedia of Life; Haootia quadriformis reconstruction from Liu et al., 2014):


It’s pretty much a law of Precambrian palaeontology that no interpretation of a fossil can ever remain uncontested, and Haootia is no exception. Nonetheless, this might be the tamest debate anyone ever had about a Precambrian fossil, and it gives me all kinds of warm feels.

Good news: Miranda et al. (2015) don’t dispute that the fossils show muscle impressions. They don’t even dispute that they belong to a cnidarian-grade creature. However, they question some of the details of the muscular arrangement, which could have implications for what this creature was and how it functioned.

They don’t have much of an issue with the muscles that run along the stalk and arms. The main point of contention, as far as I can tell, is that the muscles that run around the body (called coronal muscles in modern jellies) are not that big in living staurozoans. Those are the muscles that regular jellyfish use to contract their bells while swimming, but staurozoans don’t swim and therefore don’t need huge coronal muscles.

By Liu et al.‘s (2014) reconstruction (see above), Haootia has pretty massive coronal muscles. Miranda et al. (2015) wonder whether this was really the case, or the deformation of the fossils combined with the subconscious influence of regular jellyfish misled the original authors. They offer an alternative reconstruction, in which most of the body musculature runs up and down rather than around the body wall:


However, they also entertain the possibility that Liu et al.‘s reconstruction is correct – in which case, they note, Haootia must have done something with those muscles. Did jellyfish-like pulsations somehow form part of its feeding method? Could this even be a precursor to the jellyfish way of swimming? Who knows!

Liu et al. (2015) gave the most amazing response – much of their short reply to Miranda et al.‘s comments is basically thanking them for all the extra information and insight. They seem really pleased that biologists who study living cnidarians are taking an interest in their fossils, and enthusiastic about fruitful discussions in the future. (I concur. Biologists and palaeontologists need to talk to each other!)

They did take another, closer look at Haootia and maintain that they still see a large amount of musculature running around the body. So perhaps this peculiar Precambrian animal was doing something peculiarly Precambrian that has few or no parallels in modern seas. “We must keep in mind,” they write,  “that some, or maybe most, Ediacaran body plans and feeding strategies may have been specifically adapted to Ediacaran conditions.”

Either way, the whole exchange makes me very warm and fuzzy – I love to see scientists having constructive debates and learning from each other. (I also love that Miranda et al. thank Alex Liu in their acknowledgements; they were so obviously not out to tear one another down.) Plus both teams agree that we DO have a cnidarian-type creature from the Precambrian, and we DO have lovely lovely muscle impressions. Here’s to nice people, and to the slowly sizzling fuse of the Cambrian explosion! :)



Liu AG et al. (2014) Haootia quadriformis n. gen., n. sp., interpreted as a muscular cnidarian impression from the Late Ediacaran period (approx. 560 Ma). Proceedings of the Royal Society B 281:20141202

Liu AG et al. (2015) The arrangement of possible muscle fibres in the Ediacaran taxon Haootia  quadriformis. Proceedings of the Royal Society B 282:20142949

Miranda LS et al. (2015) Is Haootia quadriformis related to extant Staurozoa (Cnidaria)? Evidence from the muscular system reconsidered. Proceedings of the Royal Society B 282:20142396

Finally, that sponge ParaHox gene

ParaHox genes are a bit like the underappreciated sidekicks of Hox genes. Or little sisters, as the case may be, since the two families are closely related. Hox genes are probably as famous as anything in evo-devo. Being among the first genes controlling embryonic development to be (a) discovered, (b) found to be conserved between very distantly related animals, they are symbolic of the late 20th century evo-devo revolution.

ParaHoxes get much less attention despite sharing some of the most exciting properties of Hox genes. Like those, they are involved in anteroposterior patterning – that is, partitioning an embryo along its head to tail axis. Also like Hox genes, they are often neatly clustered in the genome, and when they are, they tend to be expressed in the same order (both in space and time) in which they sit in the cluster*. Their main ancestral roles for bilaterian animals seem to be in patterning the gut and the central nervous system (Garstang and Ferrier, 2013).

There are three known types of ParaHox gene, which are generally thought to be homologous to specific Hox subsets of Hox genes – by the most accepted scheme, Gsx is the closest sister of Hox1 and Hox2, Xlox is closest to Hox3, and Cdx to Hox9 and above. It is abundantly clear that Hoxes and ParaHoxes are closely related, but there has been a bit of debate concerning the number of genes in the ancestral gene cluster that gave rise to both – usually called “ProtoHox” (Garcia-Fernàndez, 2005).

Another big question about these genes is precisely when they originated, and in this regard, ParaHox genes are proving much more interesting than Hoxes. You see, there are plenty of animals with both Hox and ParaHox genes, which is what you’d expect given the ProtoHox hypothesis, but there are also animals with only ParaHoxes. If there really was a ProtoHox gene/cluster that then duplicated to give rise to Hoxes and ParaHoxes, then lone ParaHoxes (or Hoxes for that matter) shouldn’t happen – unless the other cluster was lost along the way.

So a suspiciously Gsx-like gene in the weird little blob-creature Trichoplax, which has nothing remotely resembling a Hox gene, was a big clue that (a) Hox/ParaHox genes might go back further in animal evolution than we thought, (b) the loss of the entire Hox or ParaHox cluster is totally possible**, despite how fundamental these genes appear to be for correctly building an animal.

I wrote (at length) about a study by Mendivil Ramos et al. (2012), which revealed that while Trichoplax had no Hox genes and only one of the three types of ParaHox gene, it preserved the more or less intact genomic neighbourhoods in which Hox and ParaHox clusters are normally situated. One of the more interesting results of that paper was that the one sponge genome available at the time – that of Amphimedon queenslandica, which had no trace of either Hox or ParaHox genes – also contained statistically significant groupings of Hox and ParaHox neighbour genes, as if it had a Hox neighbourhood and a ParaHox neighbourhood, but the Hoxes and ParaHoxes themselves had moved out.

That study thus pointed towards an intriguing hypothesis, previously championed by Peterson and Sperling (2007) based solely on gene phylogenies: sponges once did have Hox and ParaHox genes/clusters, which at least some of them later lost. This would essentially mean that the two gene clusters go straight back to the origin of animals if not further***, and we may never find any surviving remnant of the ancestral ProtoHox cluster, since the closest living relatives of animals have neither the genes nor their neighbourhoods (that we know of).

Hypotheses are nice, but as we know, they do have a tendency to be tragically slain by ugly facts. Can we further test this particular hypothesis about sponges? Are there facts that could say yay or nay? (Of course there are. I wouldn’t be writing this otherwise ;) )

I keep saying that we should always be careful when generalising from one or a few model organisms, that we ignore diversity in the animal kingdom at our own peril, and that “distantly related to us” = “looks like our distant ancestors” is an extremely dodgy assumption. Well, here’s another lesson in that general vein: unlike Amphimedon, some sponges have not just the ghosts of vanished ParaHox clusters, but intact, honest to god ParaHox genes!

It’s calcareous sponges again. Sycon ciliatum and Leucosolenia complicata, two charming little calcisponges, recently had their genomes sequenced (alas, they weren’t yet public last time I checked), and since then, there’s been a steady stream of “cool stuff we found in calcisponge genomes” papers from Maja Adamska’s lab and their collaborators. I’ve discussed one of them (Robinson et al., 2013), in which the sponges revealed their rather unhelpful microRNAs, and back in October (when I was slowly self-destructing from thesis stress), another study announced a couple of delicious ParaHoxes (Fortunato et al., 2014).

(Exciting as it is, the paper starts by tickling my pet peeves right off the bat by calling sponges “strong candidates for being the earliest extant lineage(s) of animals”… I suppose nothing can be perfect… *sigh*)

The study actually covers more than just (Para)Hox genes; it looks at an entire gene class called Antennapedia (ANTP), which includes Hoxes and ParaHoxes plus a handful of related families I’m far less interested in. Sycon and Leucosolenia don’t have a lot of ANTP genes – only ten in the former and twelve in the latter, whereas a typical bilaterian like a fruit fly or a lancelet has several times that number – but from phylogenetic analyses, these appear to be a slightly different assortment of genes from those present in Amphimedon, the owner of the first sequenced sponge genome. This picture is most consistent with a scenario in which all of the ANTP genes in question were present in our common ancestor with sponges, and each sponge lineage lost some of them independently. (You may not realise this until you start delving into the history of various gene families, but genes come and go a LOT in evolution.)

Sadly, many of the branches on these gene trees are quite wonky, including the one linking a gene from each calcisponge to the ParaHox gene Cdx. However, somewhat fuzzy trees are not the only evidence the study presents. First, the putative sponge Cdxes possess a little motif in their protein sequences that is only present in a handful of gene families within the ANTP class. If you take only these families rather than everything ANTP and make trees with them, the two genes come out as Cdx in every single tree, and with more statistical support than the global ANTP trees gave them. Another motif they share with all Hoxes, ParaHoxes and a few of their closest relatives, but not with other ANTP class families.

Second, at least the gene in Sycon appears to have the right neighbours (Leucosolenia was not analysed for this). Since the Sycon genome sequence is currently in pieces much smaller than whole chromosomes, only four or so of the genes flanking ParaHox clusters in other animals are clearly linked to the putative Cdx in the sponge. However, when the researchers did the same sort of simulation Mendivil Ramos et al. (2012) did for Amphimedon, testing whether Hox neighbours and ParaHox neighbours found across all fragments of the genome are (a) close to other Hox/ParaHox neighbours or randomly scattered (b) mixed or segregated, they once again found cliques of genes with little overlap, indicating the once-existence of separate Hox and ParaHox clusters.

Fortunato et al. (2014) also examined the expression of their newfound Cdx gene, and found it no less intriguing than its sequence or location in the genome, although their description in the paper is very limited (no doubt because they’re trying to cram results on ten genes into a four-page Nature paper). The really interesting activity they mention and picture is in the inner cell mass of the young sponge in its post-larval stages – the bit that develops into the lining of its feeding chambers. Which, Adamska’s team contend, may well be homologous to our gut lining. In bilaterians, developing guts are one of the major domains of Cdx and ParaHox genes in general!

So at least three different lines of evidence – sequence, neighbours and expression – make this picture hang together quite prettily. It’s incredibly cool – the turning on their heads of long-held assumptions is definitely the most exciting part of science, I say! On the other hand, it’s also a little disheartening, because now that everyone in the animal kingdom except ctenophores has definitive ParaHox genes and at least the empty seats once occupied by Hox genes, are we ever going to find a ProtoHox thingy? May it be that it’ll turn up in one of the single-celled beasties people like Iñaki Ruiz-Trillo are sequencing? That would be cool and weird.

The coolest twist on this story, though, would be to discover traces of ProtoHoxes in a ctenophore, since solid evidence for ProtoHox-wielding ctenophores would (a) confirm the strange and frankly quite dubious-sounding idea that ctenophores, not sponges, are the animal lineage farthest removed from ourselves, (b) SHOW US A FREAKING PROTOHOX CLUSTER. (*bounces* >_> Umm, * cough* OK, maturity can suck it :D ) However, given how horribly scrambled at least one ctenophore genome is (Ryan et al., 2013), that’s probably a bit too much to ask…



*Weirdly, the order of expression in time is the opposite of that of the Hox cluster. In both clusters, the “anterior” gene(s), i.e. Hox1-2 or Gsx, are active nearest the front of the embryo, but while anterior Hox genes are also the earliest to turn on, in the ParaHox cluster the posterior gene (Cdx) wakes up first. /end random trivia

**Of course we’ve long known that losing a Hox cluster is not that big a deal, but previously, all confirmed losses occurred in animals with more than one Hox cluster to begin with – a fish has plenty of Hox genes left even after chucking an entire set of them.

***With the obligatory ctenophore caveat



Fortunato SAV et al. (2014) Calcisponges have a ParaHox gene and dynamic expression of dispersed NK homeobox genes. Nature 514:620-623

Garcia-Fernàndez J (2005) The genesis and evolution of homeobox gene clusters. Nature Reviews Genetics 6:881-892

Garstang M & Ferrier DEK (2013) Time is of the essence for ParaHox homeobox gene clustering. BMC Biology 11:72

Mendivil Ramos O et al. (2012) Ghost loci imply Hox and ParaHox existence in the last common ancestor of animals. Current Biology 22:1951-1956

Peterson KJ & Sperling EA (2007) Poriferan ANTP genes: primitively simple or secondarily reduced? Evolution and Development 9:405-408

Robinson JM et al. (2013) The identification of microRNAs in calcisponges: independent evolution of microRNAs in basal metazoans. Journal of Experimental Zoology B 320:84-93

Ryan JF et al. (2013) The genome of the ctenophore Mnemiopsis leidyi and its implications for cell type evolution. Science 342:1242592

Hi, real world, again!

The Mammal has emerged from a thesis-induced supermassive black hole and a Christmas-induced food coma, only to find that in the month or so that she spent barely functional and buried in chapters covered in the supervisor’s dreaded Red Pen, things actually happened in the world outside. This, naturally, manifested in thousands of items feeling thoroughly neglected in RSS readers and email inboxes. (Jesus. How many times have I vowed never to neglect my RSS feed again? Oh well, it’s not like unemployment is such a busy occupation that I can’t deal with a measly two and a half thousand articles :-P )

… earlier tonight, the paragraph here said I wasn’t doing a proper post yet, “just pointing out” a couple of the cooler things I’ve missed. Then somehow this thing morphed into a 1000+ word post that goes way beyond “pointing things out”. It’s almost like I’ve been itching to write something that isn’t my thesis. >_>

So the first cool thing I wanted to “point out” is the genome paper of the centipede Strigamia maritima, which is a rather nondescript little beast hiding under rocks on the coasts of Northwest Europe. This is the first sequenced genome of a myriapod – the last great class of arthropods to remain untouched by the genome sequencing craze after many genomes from insects, crustaceans and chelicerates (spiders, mites and co.).  The genome sequence itself has been available for years (yay!), but its “official” paper (Chipman et al., 2014) is just recently out.

Part of the appeal of Strigamia – and myriapods in general – is that they are considered evolutionarily conservative for an arthropod. In some respects, the genome analysis confirms this. Compared to its inferred common ancestor with us, Strigamia has lost fewer genes than insects, for example. Quite a lot of its genes are also linked together similarly to their equivalents in distantly related animals, indicating relatively little rearrangement in the last 600 million years or so. But this otherwise conservative genome also has at least one really unique feature.

Specifically, this centipede – which is blind – has not only lost every bit of DNA coding for known light-sensing proteins, but also all known genes specific to the circadian clock. In other animals, genes like clock and period mutually regulate one another in a way that makes the abundance of each gene product oscillate in a regular manner (this is about the simplest graphical representation I could find…). The clock runs on a roughly daily cycle all by itself, but it’s also connected to external light via the aforementioned light-sensing proteins, so we can constantly adjust our internal rhythms according to real day-night cycles.

There are many blind animals, and many that live underground or otherwise find day and night kind of irrelevant, but even these are often found to have a functioning circadian clock or keep some photoreceptor genes around. However, based on the genome data, our favourite centipede may be the first to have completely lost both. The authors of the genome paper hypothesise that this may be related to the length of evolutionary time the animals have spent without light. Things like mole rats are relatively recent “inventions”. However, the geophilomorph order of centipedes, to which Strigamia belongs, is quite old (its most likely sister group is known from the Carboniferous, so they’re probably at least that ancient). Living geophilomorphs are all blind, so chances are they’ve been that way for the last 300+ million years.

Nonetheless, the authors also note that geophilomorphs are still known to avoid light – the question now is how the hell they do it… And, of course, whether Strigamia has a clock is not known – only that it doesn’t have the clock we’re used to. We also have no idea at this point how old the gene losses actually are, since all the authors know is that one other centipede from a different group has perfectly good clock genes and opsins.

In comparison with fruit flies and other insects, the Strigamia genome also reveals some of the ways in which evolutionary cats can be skinned in multiple ways. There is an immune-related gene family we share with arthropods and other animals, called Dscam. The product of this gene is involved in pathogen recognition among other things, and in flies, Dscam genes are divided into roughly 100 chunks or exons, most of which are are found in clusters of variant copies. When the gene is transcribed, only one of these copies is used from each such cluster, so in practical terms the handful of fruit fly Dscam genes can encode tens of thousands of different proteins, enough to adapt to a lot of different pathogens.

A similar arrangement is seen in the closely related crustaceans, although with fewer potential alternative products. In other groups – the paper uses vertebrates, echinoderms, nematodes and molluscs for comparison – the Dscam family is pretty boring with at most one or two members and none of these duplicated exons and alternative splicing business. However, it looks like insects+crustaceans are not the only arthropods to come up with a lot of DSCAM proteins. Strigamia might also make lots of different ones (“only” hundreds in this case), but it achieved this by having dozens of copies of the whole gene instead of performing crazy editing feats on a small number of genes. Convergent evolution FTW!

Before I paraphrase the entire paper in my squeeful enthusiasm (no, seriously, I’ve not even mentioned the Hox genes, and the convergent evolution of chemoreceptors, and I think it’s best if I shut up now), let’s get to something else that I can’t not “point out” at length: a shiny new vetulicolian, and they say it’s related to sea squirts!

Vetulicolians really deserve a proper discussion, but in lieu of a spare week to read up on their messiness, for now, it’s enough to say that these early Cambrian animals have baffled palaeontologists since day one. Reconstructions of various types look like… a balloon with a fin? Inflated grubs without faces? I don’t know. Drawings below (Stanton F. Fink, Wikipedia) show an assortment of the beasts, plus Yunnanozoon, which may or may not have something to do with them. Here are some photos of their fossils, in case you wondered.

Vetulicolians from Wiki

They’re certainly difficult creatures to make sense of. Since their discovery, they’ve been called both arthropods and chordates, and you can’t get much farther than that with bilaterian animals (they’re kind of like the Nectocaris of old, come to think of it…).

The latest one was dug up from the Emu Bay Shale of Australia, the same place that yielded our first good look at anomalocaridid eyes. Its newest treasure has been named Nesonektris aldridgei by its taxonomic parents (García-Bellido et al., 2014), and it looks something like this (Diego García-Bellido’s reconstruction from the paper):


In other words, pretty typical vetulicolian “life but not as we know it”, at first glance. Its main interest lies in the bit labelled “nc” in the specimens shown below (from the same figure):


This chunky structure in the animal’s… tail or whatever is a notochord, the authors contend. Now, only one kind of animal has a notochord: a chordate. (Suspicious annelid muscle bundles notwithstanding. Oh yeah, I also wanted to post on Lauri et al. 2014. Oops?) So if this thing in the middle of Nesonektris’s tail is a notochord, then at the very least it is more closely related to chordates than anything else.

Why do they think it is one? Well, there are several long paragraphs devoted to just that, so here goes a summary:

1. It’s probably not the gut. A gut would be the other obvious ID, but it doesn’t fit very well in this case. Structures interpreted as guts in other vetulicolians – which sometimes contain stuff that may be half-digested food – (a) start in the front half of the body, where the mouth is, (b) constrict and expand and coil and generally look much floppier than this, (c) don’t look segmented, (d) sometimes occur alongside these tail rod-like thingies, so probably aren’t the same structure.

2. It positively resembles modern half-decayed notochords. The notochords of living chordates are long stacks of (muscular or fluid-filled) discs, which fall apart into big blocks as the animal decomposes after death. Here’s what remains of the notochord of a lamprey after two months for comparison (from Sansom et al. (2013)):


This one isn’t as regular as the blockiness in the fossils, I think, but that could just be the vetulicolians not being quite as rotten.

There is, of course, a but(t). To be precise, there are also long paragraphs discussing why the structure might not be a notochord after all. It’s much thicker than anything currently interpreted as such in reasonably clear Cambrian chordates, for one thing. Moreover, it ends right where the animal does, in a little notch that looks like a good old-fashioned arsehole. By the way, the paper notes, vetulicolian tails in general don’t go beyond their anuses by any reasonable interpretation of the anus, and a tail behind the anus is kind of a defining feature of chordates, though this study cites a book from the 1970s claiming that sea squirt larvae have a vestigial bit of proto-gut going all the way to the tip of the tail. (I suspect that claim could use the application of some modern cell labelling techniques, but I’ve not actually seen the book…)

… and there is a phylogenetic analysis, in which, if you interpret vetulicolians as deuterostomes (which impacts how you score their various features), they come out specifically as squirt relatives whether or not you count the notochord. I’m never sure how much stock to put in a phylogenetic analysis based on a few bits of anatomy gleaned from highly contentious fossils, but at least we can say that there are other things – like a hefty cuticle – beyond that notochord-or-not linking vetulicolians to a specific group of chordates.

Having reached the end, I don’t feel like this paper solved anything. Nice fossils either way :)

And with that, I’m off. Maybe next time I’ll write something that manages to be about the same thing throughout. I’ve been thinking that I should try to do more posts about broader topics rather than one or two papers (like the ones I wrote about ocean acidification or homology versus developmental genetics), but I’ve yet to see whether I’ll have the willpower to handle the necessary reading. I’m remarkably lazy for someone who wants to know everything :D

(Aside: holy crap, did I ALSO miss a fucking Nature paper about calcisponges’ honest to god ParaHox genes? Oh my god, oh my GOD!!! *sigh* This is also a piece of incredibly exciting information I’ve known for years, and I miss it when it actually comes out in a journal bloody everyone reads. You can tell I’ve been off-planet!)


Chipman AD et al. (2014) The first myriapod genome sequence reveals conservative arthropod gene content and genome organisation in the centipede Strigamia maritima. PLoS Biology 12:e1002005

García-Bellido DC et al. (2014) A new vetulicolian from Australia and its bearing on the chordate affinities of an enigmatic Cambrian group. BMC Evolutionary Biology 14:214

Lauri A et al. (2014) Development of the annelid axochord: insights into notochord evolution. Science 345:1365-1368

Sansom RS et al. (2013) Atlas of vertebrate decay: a visual and taphonomic guide to fossil interpretation. Palaeontology 56:457-474

Because I couldn’t not post about Dendrogramma

And the deep sea surprises us yet again (photos of the type specimen of Dendrogramma enigmatica from Just et al. [2014]).

I totally ignored the original hype about these beasties. I saw them pop up on I Fucking Love Science the other day, read the headline, decided it was probably another annoyingly sensationalised news story about a moderately strange new species and went on with my life. (The fact that they kinda look like weird flatworms didn’t help) Well, now that I’ve seen the paper, I… nah, I don’t regret the decision to ignore the news story, because hyperbole like that headline about rewriting the tree of life drives me up the wall, but I am glad that I finally checked what the hype was all about.

It’s really cool, after all these years of humanity cataloguing the living world, to find something so weird that basically all we can say about it is that it’s an animal. At this point it’s not clear to me how much of that is genuine weirdness and how much is simply down to the lack of data. The organisms were found in bulk seafloor samples brought up from depths of 400 and 1000 m somewhere off Tasmania nearly thirty years ago, and they are apparently quite poorly preserved. There’s no DNA, though commenters on the PLoS article seem to think it might be possible to get some out of the specimens. (That would be nice!)

According to the authors’ description, the general organisation of Dendrogramma species can be discerned and is much like a cnidarian or a ctenophore – two basic germ layers with thick jelly in between, and a blind gut – but they appear to lack anything that would clearly identify them as a member of either group, such as comb rows or stinging cells. Because they appear to have only two germ layers, the authors conclude they are probably not bilaterians, but because they don’t have diagnostic features of any other kind of animal, and because there’s so much more we don’t know about them, they don’t feel brave enough to place them beyond that.

The beasties are made of a stalk and a flat disc; the mouth opens at the tip of the stalk and the gut extends into the disc, where it bifurcates repeatedly to form dozens of branches. Two comments on the PLoS website point out that this arrangement is a bit like a flatworm – many of which have a long pharynx that they can poke out to feed, and a highly branched intestine occupying most of the body (a lovely diagram and photo can be found in the bottom half of this page).

Superficially at least, it sounds possible that Dendrogramma‘s “stalk” is an extended pharynx. However, flatworms are bilaterians, and between their skin and their gut wall they are full of the tissues of the mesoderm, the third germ layer – muscles, simple kidneys, reproductive organs and quite a lot of cell-rich connective tissue. Just et al.‘s description of Dendrogramma states that the equivalent space in these creatures is filled with mesogloea, i.e. jelly with few or no cells. If Dendrogramma really lacks mesodermal tissues, then it wouldn’t make a very good flatworm! (The paper itself doesn’t discuss the flatworm option at all, presumably for similar reasons.)

Of course, the thing that piqued my interest in Dendrogramma is its supposed resemblance to certain Ediacaran fossils, specifically these ones. It would be awesome if we could demonstrate that the living and the fossil weirdos are related, since then determining what Dendrogramma is would also classify the extinct forms, but I’m not holding my breath on this count. The branching… whatevers in the fossils in question may look vaguely like the branching gut of Dendrogramma, but, as discussed above, so do flatworm guts. The similarity to the fossils may well have nothing to do with actual phylogenetic relatedness, which the authors sound well aware of.

Nature, helpful as always. >_>

It seems all we can do for the moment is wait for more material to come along, hopefully in a good enough state to make detailed investigations including genetic studies. My inner developmental biologist is also praying for embryos, but the gods aren’t generally kind enough to grant me these sorts of wishes :-P

I do quite like the name, though. Mmmmm, Dendrogramma. :)



Just J et al. (2014) Dendrogramma, new genus, with two new non-bilaterian species from the marine bathyal of southeastern Australia (Animalia, Metazoa incertae sedis) – with similarities to some medusoids from the Precambrian Ediacara. PLoS ONE 9:e102976

Precambrian muscles??? Oooooh!

Okay, consider this a cautious squee. I wish at least some of those Ediacaran fossils were a little more obvious. I mean, I might love fossils, but I’m trained to squirt nasty chemicals on bits of dead worm and play with protein sequences, not to look at faint impressions in rock and see an animal.

Most putative animals from the Ediacaran period, the “dark age” that preceded the Cambrian explosion, are confusing to the actual experts, not just to a lab/computer biologist with a fondness for long-dead things. The new paper by Liu et al. (2014) this post is about lists a “but see” for pretty much every interpretation they cite. The problem is twofold: one, as far as I can tell, most Ediacaran fossils don’t actually preserve that much interpretable detail. Two, Ediacaran organisms lived at a time when the kinds of animal body plans we’re familiar with today were just taking shape. The Ediacaran is the age of ancestors, and it would be more surprising to find a creature we can easily categorise (e.g. a snail) than a weird beastie that isn’t quite anything we know.

Having said that, Liu et al. think they are able to identify the new fossil they named Haootia quadriformis. Haootia comes from the well-known Fermeuse Formation of New Foundland, and is estimated to be about 560 million years old. The authors say its body plan – insofar as it can be made out on a flat image pressed into the rock – looks quite a lot like living staurozoan jellyfish, with a four-part symmetry and what appear to be branching arms or tentacles coming off the corners of its body. The most obvious difference is that Haootia seems to show the outline of a huge circular holdfast that’s much wider than usual for living staurozoans.

However, the most exciting thing about this fossil is not its shape, but the fact that most of it is made up of fine, highly organised parallelish lines – what the authors interpret as the impressions of muscle fibres. The fibres run in different directions according to their position in the body; for example, they seem to follow the long axes of the arms.

(Below: the type specimen of Haootia with some of the fibres visible, and various interpretive drawings of the same fossil. Liu et al. is a free paper, so anyone can go and look at the other pictures, which include close-ups of the fibres and an artistic reconstruction of the living animal.)

If the lines do indeed come from muscle fibres, then regardless of its precise affinities, Haootia is certainly an animal, and it is probably at least related to the group called eumetazoans, which includes cnidarians like jellyfish and bilaterians like ourselves (and maybe comb jellies, but let’s not open that can of jellies just now). Non-eumetazoans – sponges and Trichoplax – do not have muscles, and unless comb jellies really are what some people think they are, we can be almost certain that the earliest animals didn’t either.

Finding Ediacaran muscles is also interesting because it gives us further evidence that things capable of the kinds of movement attributed to some Ediacaran fossils really existed back then. Of course, it would have been nicer to find evidence of muscle and evidence of movement in the same fossils, but hey, this is the Precambrian. You take what you get.

(P.S.: Alex Liu is cool and I heart him. OK, I saw him give one short talk, interviewing for a job at my department that he didn’t get *sniffles*, so maybe I shouldn’t be pronouncing such fangirlish judgements, but that talk was awesome. As I’ve said before, my fangirlish affections are not very hard to win :) )



Liu AG et al. (2014) Haootia quadriformis n. gen., n. sp., interpreted as a muscular cnidarian impression from the Late Ediacaran period (approx. 560 Ma). Proceedings of the Royal Society B 281:20141202

A bit of Hox gene nostalgia

I had the most random epiphany over my morning tea today. I don’t even know what got me thinking about the Cambrian explosion (as if I needed a reason…). Might have been remembering something from the Euro Evo Devo conference I recently went to. (I kind of wanted to post about that, because I saw some awesome things, but too much effort. My brain isn’t very cooperative these days.)


I was thinking about explanations of the Cambrian explosion and remembering how the relevant chapter in The Book of Life (otherwise known as the book that made me an evolutionary biologist)  tried to make it all about Hox genes. It’s an incredibly simplistic idea, and almost certainly wrong given what we now know about the history of Hox genes (and animals)*. At the time, and for a long time afterwards, I really wanted it to be true because it appeals to my particular biases. But I digress.

Then it dawned on me just how new and shiny Hox genes were when this book was written. I thought, holy shit, TBoL is old. And how far evo-devo as a field has come since!

The Book of Life was first published in 1993. That is less than a decade after the discovery of the homeobox in fruit fly genes that controlled the identity of segments (McGinnis et al., 1984; Scott and Weiner, 1984), and the finding that homeoboxes were shared by very distantly related animals (Carrasco et al., 1984). It was only four years after the recognition that fly and vertebrate Hox genes are activated in the same order along the body axis (Graham et al., 1989; Duboule and Dollé, 1989).

This was a HUGE discovery. Nowadays, we’re used to the idea that many if not most of the genes and gene networks animals use to direct embryonic development are very ancient, but before the discovery of Hox genes and their clusters and their neatly ordered expression patterns, this was not at all obvious. What were the implications of these amazing, deep connections for the evolution of animal form? It’s not surprising that Hox genes would be co-opted to explain animal evolution’s greatest mysteries.

It also occurred to me that 1993 is the year of the zootype paper (Slack et al., 1993). Slack et al. reads like a first peek into a brave new world with limitless possibilities. They first note the similarity of Hox gene expression throughout much of the animal kingdom, then propose that this expression pattern (their “zootype”) should be the definition of an animal. After that, they speculate that just as the pattern of Hox genes could define animals, the patterns of genes controlled by Hoxes could define subgroups within animals. Imagine, they say, if we could solve all those tough questions in animal phylogeny by looking at gene expression.

As always, things turned out More Complicated, what with broken and lost Hox clusters and all the other weird shit developmental “master” genes get up to… but it was nice to look back at the bright and simple childhood of my field.

(And my bright and simple childhood. I read The Book of Life in 1998 or 1999, not entirely sure, and in between Backstreet Boys fandom, exchanging several bookfuls of letters with my BFF and making heart-shaped eyes at long-haired guitar-playing teenage boys, I somehow found true, eternal, nerdy love. *nostalgic sigh*)


*Caveat: it’s been years since I last re-read the book, and my copy is currently about 2500 km from me, so the discussion of the Cambrian explosion might be more nuanced than I remember. Also, my copy is the second edition, so I’m only assuming that the Hox gene thing is there in the original.



Carrasco AE et al. (1984) Cloning of an X. laevis gene expressed during early embryogenesis coding for a peptide region homologous to Drosophila homeotic genes. Cell 37:409-414

Duboule D & Dollé P (1989) The structural and functional organization of the murine HOX gene family resembles that of Drosophila homeotic genes. The EMBO Journal 8:1497-1505

Graham A et al. (1989) The murine and Drosophila homeobox gene complexes have common features of organization and expression. Cell 57:367-378

McGinnis W et al. (1984) A conserved DNA sequence in homoeotic genes of the Drosophila Antennapedia and bithorax complexes. Nature 308:428-433

Scott MP & Weiner AJ (1984) Structural relationships among genes that control development: sequence homology between the Antennapedia, Ultrabithorax, and fushi tarazu loci of Drosophila. PNAS 81:4115-4119

Slack JMW et al. (1993) The zootype and the phylotypic stage. Nature 361:490-492

Ctenophore nervous systems redux

… and reasons I suddenly find myself liking Joseph Ryan.

Ryan was the first author on the first ctenophore genome paper, published last December, though I’d known his name long before that thanks to his developmental genetic work on jelly creatures of various kinds. As is clear from the genome study, he leans quite strongly towards the controversial idea that ctenophores represent the sister lineage to all other animals.

And here’s reason one that my eyes suddenly have little cartoon hearts pulsing in their irises upon reading his short perspective paper in Zoology (Ryan, 2014). Throughout the paper, not once does he refer to ctenophores as “the” basal animal lineage. Instead, he uses phrases like “most distant relative to all other animals” or “the sister group to the rest of the animals”.

In other words, he’s scrupulously avoiding my giantest pet peeve, and I’m sure he doesn’t do it to please an obscure blogger, but gods, that’s even better. I don’t want to be pleased, I want evolutionary biology to get rid of stupid anthropocentric ladder-thinking nonsense.

Anyway, the little paper isn’t actually about animal phylogeny, it’s about nervous systems.

Both ctenophore genome papers argued that the ancestors of these pretty beasties might have evolved nervous systems independently of ours. The second one seemed positively convinced of this, but, as Ryan’s review points out, there are other possibilities even assuming that the placement of ctenophores outside the rest of the animals is correct.

While it’s possible that nerve cells and nervous systems evolved twice among the animals – it is equally possible that they have been lost twice (i.e. in sponges and blobby little placozoans). Full-fledged nerve cells wouldn’t be the first things that sponges and blobs have lost.

And Ryan basically wrote this short piece just to point that out. The argument that ctenophore nervous systems are their own invention is based on the absence or strange behaviour of many “conserved” nervous system-related genes. Ctenophores appear to completely lack some common neurotransmitters such as dopamine, as well as a lot of genes/proteins that are necessary for nerve synapses to work in us. Other genes that are “neural” in other animals are present but not associated with the nervous system in ctenophores.

BUT, Ryan cautions, there are also commonalities that shouldn’t be dismissed. While ctenophores can’t make dopamine, they do possess several other messenger molecules common in animal nervous systems. Same goes for the proteins involved in making synapses. Likewise, while they completely lack some of the genes responsible for defining various types of nerve cells (see: Hox genes), other genes involved in the same kind of stuff are definitely there.

The key thing, he says, is to take a closer look at more of these genes and find out what they do by manipulating them. Since there are clearly both similarities and differences, we must assess their extent.

And that, my friends, is the question at the heart of every homology argument ever. How similar is similar enough? Greater minds than mine have struggled with the answer, and I imagine they’ll continue to struggle until we invent time machines or find fossils of every single stage in the evolution of everything.

Until then, I’ll leave you with the closing lines of Ryan’s paper. I may not agree that we’ve “revealed” the position of ctenophores, but I’m absolutely on board with the excitement :)

One thing is quite clear: something remarkable happened regarding the evolution of the nervous system very early in animal evolution. Either a nervous system existed in the ancestor and was lost in certain lineages, or ctenophores invented their own nervous system independently (Fig. 1). Either possibility is quite extraordinary. The revelation that ctenophores are the sister group to the rest of animals has sparked a truly exciting debate regarding the evolutionary origins of the nervous system, one that will continue as additional genomic and functional data come to the fore.


Ryan JF (2014) Did the ctenophore nervous system evolve independently? Zoology in press, available online 11/06/2014, doi: 10.1016/j.zool.2014.06.001

About X-frogs and failing at regeneration

Not the usual mad squee, but here’s a neat little system for studying regeneration that I quite liked today. I normally think about regeneration in terms of amputated limbs, mutilated hearts, decapitated flatworms. But you can induce a kind of “regeneration” in a less drastic and rather more tricksy way, at least in some animals. In newts and salamanders, you can create a small, superficial wound on the side of a limb, then manipulate a nearby nerve into it and add some skin from the other side of the limb.

The poor hurt limb then decides you’ve actually cut something off and tells the wound to grow a new limb. If you don’t add skin, regeneration begins but doesn’t progress very far; if you don’t add a nerve, nothing happens. IIRC you can also make extra heads in some worms in a similar way, but I digress. The figure below from Endo et al. (2004) illustrates just how well the procedure can work. The top row shows stages in the development of the extra limb, while D shows the stained skeletons of the original and new limbs. I’d say that’s a pretty good looking forearm and hand!



That this trick works is in itself a very interesting insight into the nature of regeneration, as it helps us figure out exactly what it is that triggers various steps of regeneration as opposed to a simple healing process (Endo et al., 2004).

Clawed frogs (Xenopus) have been staples of embryology for a long time, but they are also quite fascinating from a regeneration point of view. One, they can regrow their limbs while they are tadpoles, but mostly lose the ability as they mature. They also have a really weird thing going on with their tadpole tails, which they can regenerate early on, then can’t, then can again (Slack et al., 2004). Huh? O.o

Two, their adult limb regeneration ability is not totally absent: it’s somewhere between salamanders’ (oh, whatever, fine, I can do that!) and ours (uh… as long as I’m a baby and it’s just a fingertip?). In a frog, an amputated arm or leg doesn’t simply heal over, but the… thing that grows out of the stump is just a simple cartilaginous spike with no joints or muscles. It’s as if the system was trying very hard to remember how to form a limb but kind of got distracted.

We are obviously interested in creating superhumans with mad regeneration skillz, which also makes us interested in how and why animals lose this seemingly very useful ability*. (Bely (2010) wrote a lovely piece on this not at all simple question.) So: Xenopus yay!

Now, Mitogawa et al. (2014) have devised a skin wound + nerve deviation system to grow little extra limb buds in adult frogs. As you might expect, it doesn’t work nearly as well as it does in axolotls: you need three nerves rather than one, and it only induces a bud about half the time, but it works well enough for research purposes.

The bud (technically, a blastema when you’re talking about regeneration) looks like a good regeneration blastema: it’s got the seemingly undifferentiated cells inside, it’s got the thickened epidermis at the tip that teams up with the nerves to give developmental instructions to the rest of the thing, and it expresses a whole bunch of genes that are turned on in normal limb blastemas.

(Totally random aside: thanks to Chrome’s spell checker, I have discovered that “blastema” is an anagram for “lambaste”.)

One area where this blastema-by-trickery fails is making cartilage, which is one of the few proper limb things the defective spike regenerates in frogs do contain. There’s no simple way of coaxing a complete spike out of these blastemas. The researchers tried the skin graft thing from axolotls (which can already form cartilage without the skin graft), but they still only got a little blastema with no cartilage. To get a skeleton, however crappy,  you need to cut out muscles and crack the underlying bone, which kind of defeats the purpose of the whole exercise IMO. At that point, you might as well just chop off the arm.

Below: the best a frog can do. Development of blastema-like bumps and “spike limbs” on the upper arm from Mitogawa et al. (2014). Compared to the fully formed accessory limbs of axolotls, the things you can see in B-D here are not terribly impressive, but they may be just the “transitional form” we need!

The failure of skin grafts alone at inducing cartilage, however, does hint at the things that go wrong with regeneration in frogs. Mitogawa et al. speculate that newt and axolotl limbs produce factors that frogs can only get from damaged bone. Broken bones even in us form a cartilaginous callus as they begin to heal, and unlike the cartilage in the extra limbs of axolotls, the cartilage in frog spikes is directly attached to the underlying bone.

They also point out that if you add proteins called BMPs to amputated mouse arms, which are otherwise even shitter at regeneration than frog arms, a surprising amount of bone formation occurs. (“BMP” stands for bone morphogenetic protein, which is a big clue to their function.) So it looks like there may be a kind of regeneration gradient from mammals (need bone damage AND extra BMP), through frogs (need bone damage, take care of BMPs themselves) to salamanders (don’t need either).

I should point out that salamanders and frogs are equally closely related to us, so this isn’t a proper evolutionary gradient, but given all the ways in which we and amphibians are fundamentally similar, our loss of regenerative ability may well have evolved through a similar stage to where frogs are now. Neat!

(I just wish they stopped calling us “higher vertebrates”. That phrase annoys me right up the fucking wall, because, and I may have said this before, EVOLUTION IS NOT A GODDAMNED LADDER. The group they are referring to has a perfectly good name that doesn’t imply ladder thinking. Amniotes, people. Or mammals, if you mean mammals, but I think if they’d meant mammals they would have said mammals. End grump.)


*I mean “us” in a very general sense. I think regenerative medicine is the coolest thing in medicine since vaccines and antibiotics, but I personally don’t think that the evolution of regenerative ability needs medical considerations to make it interesting. Whatever. I’m not exactly a practically minded person :-P



Bely AE (2010) Evolutionary loss of animal regeneration: pattern and process. Integrative and Comparative Biology 50:515-527

Endo T et al. (2004) A stepwise model system for limb regeneration. Development 270:135-145

Mitogawa K et al. (2014) Ectopic blastema induction by nerve deviation and skin wounding: a new regeneration model in Xenopus laevis. Regeneration 2:11

Slack JMW et al. (2004) Cellular and molecular mechanisms of regeneration in Xenopus. Philosophical Transactions of the Royal Society B 359:745-751

The ctenophore conundrum, by popular demand

So, a new ctenophore genome has just been published in Nature (Moroz et al., 2014), it makes some extraordinary claims, and my resident palaeontologist/web-buddy Dave Bapst wants my opinion ;)

Given that I already planned to have an opinion about the first ctenophore genome back in December (Ryan et al., 2013) and miserably failed to finish the post… the temptation is just too strong. (That thesis chapter draft in the other window of MS Word wasn’t going to be finished today anyway  >_>)

Whatever I might seem from words on the internet, I’m not some kind of expert on phylogenetics, so I’m going to use a crutch. I had this idea back when I first read Ryan et al. (2013), because I remember thinking that it was written almost as if Nosenko et al. (2013) had never happened, and I’d really liked Nosenko et al. (as you can guess from the word count of this post), so I was mildly indignant about that. The Nosenko paper is going to be my crutch. (No offence to Hervé Philippe and friends, but there are only so many papers I’m going to reread for an out of the blue blog post ;) )

Although I’m obviously not writing a public post specifically for a phylogeny nut, I may get somewhat technical, and I’m definitely going to get verbose.


Ctenophores. Comb jellies, sea gooseberries, Venus girdles. They are floaty, ethereal, mesmerizingly beautiful creatures, and I have it on good authority that they are also complete pains in the arse.

Here’s some pretty pictures before it gets too painful ;) Left: Mnemiopsis leidyi from Ryan et al. (2013); right: Pleurobrachia bachei from Moroz et al. (2014). And a bonus video of a Venus girdle making like an ancient nature spirit. I could watch these beasties all day.


Venus from Sandrine Ruitton on Vimeo.

The problem(s)

And now, the pain. Let’s pull out my trusty old animal phylogeny, because the question marks are once again highly appropriate. (Also, I’m hell-bent on breaking your bandwidth with PICTURES.)


Ryan et al. (2013) helpfully have a figure distilling the ideas people have had about those question marks so far:


Bi = bilaterians, Cn = cnidarians, Ct = ctenophores, Tr = Trichoplax, and Po = sponges (Porifera).

I say “helpfully,” but it’s not all that helpful after all, since pretty much every possible configuration has been proposed. Why is this such a difficult question? Here’s a quick rundown of the problems Nosenko et al.’s study found to affect the question marks:

  1. Fast-evolving protein sequences – these can cause artefacts because too much change overwrites informative changes and creates chance similarities. Excluding faster-evolving sequences from the analysis changes the tree.
  2. Sequence data that don’t conform to the simplifying assumptions of popular evolutionary models – again, this can result in chance similarities and artefacts, and using a poorer model replicates the effects of using less ideal sequences.
  3. Long-branched outgroups – these are the non-animal groups used to place the root of animals. The more distant from animals and less well-sampled the outgroup, the longer the branches it forms, which can attract fast-evolving animal lineages towards the root. In Nosenko et al.’s analyses, even the closest outgroup seemed to cause problems, and removing the outgroup altogether made the conflicts between different models and datasets disappear completely – but this isn’t exactly helpful when you’re looking for the root of the animal tree!

The problem with ctenophores in particular is illustrated by this one of Nosenko et al.’s trees, made from one of their less error-prone datasets:


The ctenophore branch is not only longer overall than pretty much any other in the tree; its length is also very unevenly distributed between the loooong history common to all species and the short unique lineage of each individual species. That is bad news. And it may stay that way forever, because the last common ancestor of living ctenophores may genuinely be very recent, so there’s no way to divide up that long-ass internal branch without a time machine.

Round 1: Nosenko vs. Ryan

In fairness, the Mnemiopsis genome team probably didn’t have a whole lot of time to specifically deal with Nosenko et al.’s points (OTOH, none of those individual points were truly new). The Nosenko paper came out in January 2013, and the Mnemiopsis genome paper was received by Science in July of the same year – I imagine most of the data had been generated way before then, and you can’t just redo all your data analysis and rewrite a paper on short notice.

I’m still going to view Ryan et al. (2013) in the light of Nosenko, because regardless of the genome team’s ability to answer them, some of Nosenko et al.’s points are very relevant to the claims they make. Their biggest claim, of course, being that ctenophores are the sister group to all other animals.

In Nosenko et al.’s experiments, this placement showed up in trees where faster-evolving genes, poorer models or more distant outgroups were used, but not when the slowest-evolving gene set was analysed with the best models and the closest outgroup.

Ryan et al. acknowledge that “supermatrix analyses of the publicly available data are sensitive to gene selection, taxon sampling, model selection, and other factors [cite Nosenko].” Their data are obviously sensitive to such factors. In fact, they behave rather similarly to what I saw in the Nosenko study.

Ryan et al. used two method/model combinations – one of the models was the preferred CAT model of Nosenko et al., and the other was the OK but not great GTR model that CAT beat by miles in terms of actually fitting Nosenko et al.’s data. (Caveat: in the genome paper, the CAT and GTR models were used with different treebuilding methods, so we can’t blame the models for different results with any certainty.) Also, they analysed the data with three different outgroups.

And guess what – the ctenophores-outside-everything tree was best supported with (1) the GTR model, (2) the more distant outgroups. There is not much testing of the effect of gene choice – there were two different data sets, but they were both these massive amalgamations of everything useable, and they also included totally different samples of species.

However, here comes another nod to Nosenko et al. and all the other people who advocated trying things other than “conventional” sequence comparisons through the years. Provided you can securely identify genes across different organisms, you can also try to deduce evolutionary history based on their presences and absences rather than their precise sequences. This is not a foolproof approach because genes can be (commonly) lost or (occasionally) picked up from other organisms, but it is often regarded as less artefact-prone than sequence-based trees.

But does it help with ctenophores? Like the GTR model-based sequence trees, the tree based on gene presence/absence (you obviously need complete genomes for this!) supports ctenophores being the outsider among animals:


My problem with this? Note what else it supports. The white circles indicate groupings that this method had absolutely no doubt about. And these groupings include things that frankly sound like abject nonsense. Here’s one annelid worm (the leech Helobdella) sitting next to a flatworm, while another annelid worm (Capitella) teams up with a limpet right next to a chordate. If anything, that is more controversial than the placement of ctenophores, because we thought we had it settled!

So if we’re concluding that ctenophores are basal to all other animals, why aren’t we also making a fuss about the explosion of phylum Annelida? Surely, if this method gives us strong enough conclusions to arbitrate between different sequence-based hypotheses about ctenophores, it’s strong enough to make those claims too. The cake can’t quite decide if it’s being eaten, I think.

I’m not sure what to think about the sequence trees. I’m far more confident about the presence/absence one. Maybe I’m just demonstrating the Dunning-Kruger effect here, but I’m not buying that tree for a second.

Overall verdict?

Not convinced. Not by a long shot.

Round 2: Nosenko vs. Moroz

The Pleurobrachia genome took me completely by surprise. I’d known Mnemiopsis was sequenced since Ryan et al. (2010). (Three years. Can you imagine the twitching?) I had no idea this other project was happening, so I nearly fell off my chair when Nature dropped it into my RSS reader yesterday. Another ctenophore genome – and another one that supports ctenophore separatism? (This hypothesis is becoming strangely popular…)

Bonus: it’s not just a genome paper, it also describes the transcriptomes of ten different ctenophores. Transcriptomes, the set of all active genes, are a little bit easier to sequence and assemble than genomes, and if you’re thorough they’ll catch most of the genes the organism has, so they can be almost as good for the analysis of gene content.

Which they kind of don’t do properly. There is a discussion of specific gene families that ctenophores lack – including many immune- and nervous system-related genes – but that’s not exactly saying much given that we know even “important” genes can be lost (case in point: the disappearing (Para)Hox genes of Trichoplax). The fact that ctenophores seem to completely lack microRNAs is interesting, but again, it doesn’t mean they never had them. Sponges do have microRNAs but don’t seem to be nearly as big on them as other animals.

As for the global analysis of gene content – I had to chase down a reference (Ptitsyn and Moroz, 2012) to understand what they actually did. As far as I can tell, there is no phylogenetic analysis involved – they just took a tree they already had, and used this method to map gene gains and losses onto that tree. Which is cool if you’re fairly sure about your tree, but pretty much meaningless when the tree is precisely the question. The Mammal is disappointed.

One of the problems with listing genes that aren’t there or don’t work in the “expected” way in ctenophores is that even if they’re not outside everything else, it’s still a distinct possibility that these guys branched off from our lineage before cnidarians did. For example, the Pleurobrachia paper spends a lot of time on “nervous system-specific” genes like elav missing or not being expressed in neurons, and common neurotransmitters like serotonin not being used by ctenophores.

But, assuming that the tree of animals looks something like (sponges + (ctenophores + (cnidarians + bilaterians))), we wouldn’t expect ctenophore nervous systems to share every property that cnidarians and bilaterians share. Remember: (1) sponges don’t have nervous systems, so they’re not much use as a comparison, (2) cnidarians + bilaterians had a longer common ancestry than either did with ctenophores. Genes possessed by sponges PLUS cnidarians and/or bilaterians but missing from ctenophores are more suggestive, but only if you can demonstrate that they weren’t lost. (We’re kind of going in circles here…)

The other problem is that pesky last common ctenophore ancestor. If it really is very recent, then taking even all living ctenophores to represent ctenophore diversity is like taking my close family to represent human diversity. Just like my family contains pale-skinned, lactose tolerant people, it is entirely possible that this lone surviving ctenophore lineage possesses (or lacks) important traits that aren’t at all typical of ctenophores as a whole. Ryan et al.’s supplementary data are clear that at least the Mnemiopsis genome is horribly scrambled, all trace of conserved gene neighbourhoods erased from it. That’s not exactly promising if you’re hoping for “trustworthy” animals.

The actual phylogenetic trees in Moroz et al. (2014) seem to follow an approach of throwing AAAALLL the genes at the problem. The biggest dataset contains 586 genes, compared to 122 in Nosenko et al.’s largest collection, and there is not much filtering by gene properties other than “we can tell what it is”. I have no idea how the CAT + WAG model they used compares to CAT or WAG or GTR on their own; unfortunately, the Nosenko paper doesn’t test that particular setup and this one doesn’t do any model testing. Moroz et al.’s supplementary methods claim it’s pretty good, cite something, and I’m not gonna chase down that reference. (Sorry, I’ve been poring over this for four hours at this point).

Interestingly, the support for ctenophores being apart from other animals increases when they start excluding distant outgroups. The only time it’s low is when they add all ten ctenophores and use fewer genes. Hmm. This is where I would like to hear some real experts’ opinions, because on the face of it, I can’t pinpoint anything obviously wrong. (Other than saying that chucking more genes at a problem tree is perfectly capable of making the problem worse)

TL;DR version: While I’m generally underwhelmed by the gene content stuff, I literally have no idea what to think about the trees.

I’m banking on the hope that someone will do.


And… I think that is all the opinion I’m going to have about ctenophores for a long time. Lunch was a long time ago, my brain is completely fried, and I’m not sure how much of the above actually makes sense. To be clear, I don’t really have a horse in this race, though I’d really like to know the truth. (Fat chance of that, by the looks of it…) I think I’m going to need a bit more convincing before I stop looking sideways at this idea that ctenophores are further from us than sponges. If anything is clear from recent phylogenomics papers, it’s that what data you analyse and how you analyse them makes a huge difference to the result you get, and this is happening with data and methods where it’s not necessarily easy to dismiss an approach as clearly inferior.

It’s a mess, damn it, and I’m not qualified to untangle it. Urgh.



Moroz LL et al. (2014) The ctenophore genome and the evolutionary origin of neural systems. Nature advance online publication, 21/05/2014; doi: 10.1038/nature13400

Nosenko T et al. (2013) Deep metazoan phylogeny: When different genes tell different stories. Molecular Phylogenetics and Evolution 67:223-233

Ptitsyn A & Moroz LL (2012) Computational workflow for analysis of gain and loss of genes in distantly related genomes. BMC Bioinformatics 13:S5

Ryan JF et al. (2010) The homeodomain complement of the ctenophore Mnemiopsis leidyi suggests that Ctenophora and Porifera diverged prior to the ParaHoxozoa. EvoDevo 1:9

Ryan JF et al. (2013) The genome of the ctenophore Mnemiopsis leidyi and its implications for cell type evolution. Science 342:1242592