Did cephalopod reflective proteins originate by horizontal transfer?

Not too long ago, I posted a thread on Twitter (url in references), exploring the claims of a recent paper in Current Biology (Guan et al. 2017) that reflective proteins of cephalopod skin originated via Horizontal Gene Transfer (HGT) from bacteria. If true, this would be really amazing.

Demonstrating HGT relies on phylogenetics, which is not well done in this paper. Still, and even though some of the language of the paper is imprecise, I think the evidence for HGT is enticing. The first evidence for HGT in the paper is the similarity between 24 base pairs found in the protein reflectin and a plasmid in Vibrio fisheri, the light-producing symbiont of some cephalopods. The gene similar to reflectin in Vibrio is called TnlD. The authors found traces of it in a completely sequenced Octopus genome! They also report evidence of transposition, a 12 bp inverted repeat (IR) sequence in that Octopus genome! So, I find that to be good evidence of transposition. But what direction?

As the authors acknowledge, V. fisheri might have picked up a cephalopod gene, rather than vice versa, a question that can be answered with phylogenetic methods. Unfortunately, here is where the phylogenetics falls down. In their supplement they provide two phylogenies, both using distance based NJ, which are very fast, but often less accurate than Maximum Likelihood and Bayesian methods. Even worse, the authors do not state how they choose an outgroup sequence. The claim of HGT relies critically on which gene(s) are set as the outgroup(s).

In one tree (Fig S1C) the Vibrio sequence is nested within cephalopod sequences, which would mean the horizontal transfer went TO Vibrio FROM Octopus, assuming S1C is a rooted tree, which goes unstated. What they really needed to do is find a bunch of sequences in OTHER Vibrionaceae. I did a quick BLAST search, and such sequences do exist, in multiple genera of Vibrionacea! This is consistent with origin in Cephalopoda by HGT, but would require better phylogenetic analysis and an explicit definition of the outgroup genes.

Here is where we get to the imprecise language. The paper suggests that Vibrio fisheri is the source of horizontal gene transfer. But cephalopods are old. Coleoids are something like 250MY (Tanner et al. 2017)! So, if HGT is the source of coleoid reflectin, unless HGT happened a bunch of times it happened hundreds of millions of years ago because all coleiods (living cephalopods except Nautilus) probably have reflectins. I very much doubt Vibrio fisheri, living today, is that old. However, if Vibrionacea were around then, and had the TnL genes, origin in cephalopods by HGT is possible. Since I found evidence of TnL genes (similarity) in multiple Vibrionacea,those could be old, inherited from the ancestor of those Vibrionacea. If it’s indeed old, HGT from vibrionacea to cephalopods is possible. However, this would require a much better phylogenetic analysis. In that case a phylogenetic analysis of Vibrionacea genes and cephalopod reflectins, should show the cephalopod genes nested within Vibrionacea genes. Furthermore, rooting gene trees is difficult. Maybe midpoint rooting is the best way to go, although there can be some signal about the root in non-reversible models of molecular evolution.

So, in conclusion -- HGT by transposition seems to me quite likely. However, I remain agnostic about directionality. That said, even HGT TO Vibrio is quite interesting, as is transposition of reflectin, as is photonics of core “protosequence”!

References

https://twitter.com/UCSB_OakleyLab/status/907657070366699520

Guan Z., Cai T., Liu Z., Dou Y., Hu X., Zhang P., Sun X., Li H., Kuang Y., Zhai Q., Ruan H., Li X., Li Z., Zhu Q., Mai J., Wang Q., Lai L., Ji J., Liu H., Xia B., Jiang T., Luo S.-J., Wang H.-W., Xie C. 2017. Origin of the Reflectin Gene and Hierarchical Assembly of Its Protein. Curr. Biol. 27:2833–2842.e6.

Tanner A.R., Fuchs D., Winkelmann I.E., Gilbert M.T.P., Pankey M.S., Ribeiro Â.M., Kocot K.M., Halanych K.M., Oakley T.H., da Fonseca R.R., Pisani D., Vinther J. 2017. Molecular clocks indicate turnover and diversification of modern coleoid cephalopods during the Mesozoic Marine Revolution. Proc. Biol. Sci. 284.