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Phylogeny of melanopsin in C. milii and other vertebrates.Phylogenetic analyses based on a codon-matched nucleotide alignment of elephant shark opn4 cDNA sequences compared to the melanopsin sequences of representative chordates and the published visual and non-visual photosensory pigments of the zebrafish (Danio rerio), showing the relative positioning of elephant shark opn4m1, opn4m2 and opn4x photopigments (arrow) within the two main clades (m-class (purple) and x-class (blue)) of melanopsin. (A) A Bayesian Probabilistic Inference (BPI) method, performed with a Metropolis Markov chain Monte Carlo (MCMC) algorithm [43], [44] and incorporating a general time-reversal (GTR) model [45] with posterior probability values (represented as a percentage) indicated at the base of each node. (B) Maximum Composite Likelihood (MCL) [47] and (C) Kimura 2-Parameter (K2P) substitution [48] matrix methods, both generating bootstrapped, Neighbour-Joining (NJ) phylogenetic trees [49], with the degree of internal branching expressed as a percentage. The scale bar indicates the number of nucleotide substitutions per site. The human GPR21 (GenBank accession number: NM005294) and GPR52 (GenBank accession number: NM005684) nucleotide sequences were used as outgroups. The opsin sequences and their GenBank accession numbers used for generating the tree are as follows: (i) exorhodopsin (exorh): zebrafish (Danio rerio), NM131212; (ii) rod opsin (rh1): zebrafish (Danio rerio), NM131084 (rh1.1), HM367062 (rh1.2); (iii) rod opsin-like 2 (rh2): zebrafish (Danio rerio), NM131253 (Rh2.1), NM182891 (Rh2.2), NM182892 (Rh2.3), NM131254 (Rh2.4); (iv) short-wavelength-sensitive 2 (sws2): zebrafish (Danio rerio), NM131192; (v) short-wavelength-sensitive 1 (sws1): zebrafish (Danio rerio), NM131319; (vi) long-wavelength-sensitive/middle-wavelength-sensitive (lws/mws): zebrafish (Danio rerio), NM131175 (lws1), NM001002443 (lws2); (vii) vertebrate ancient (va) opsin: zebrafish (Danio rerio), AB035276 (va1), AY996588 (va2); (viii) panopsin (opn3): zebrafish (Danio rerio), NM001111164; (ix) teleost multiple tissue (tmt) opsin: zebrafish (Danio rerio), BC163681; (x) retinal pigment epithelium-specific rhodopsin homolog (rrh) (peropsin): zebrafish (Danio rerio), NM001004654; (xi) retinal G protein-coupled receptor (rgr): zebrafish (Danio rerio), NM001017877; (xii) mammalian-like melanopsin (opn4m): human (Homo sapiens), NM033282 (OPN4V1); cat (Felis catus), AY382594; mouse (Mus musculus), EU303118 (Opn4mlong); rat (Rattus norvegicus), NM138860; hamster (Phodopus sungorus), AY726733; mole-rat (Spalax ehrenbergi), AM748539; dunnart (Sminthopsis crassicaudata), DQ383281; chicken (Gallus gallus), EU124632 (OPN4Mlong); catfish (Ictalurus punctatus), FJ839437 (opn4m1), FJ839438 (opn4m2); roach (Rutilus rutilus), AY226847; cichlid (Astatotilapia burtoni), EU523855; zebrafish (Danio rerio), GQ925715 (opn4m1), GQ925716 (opn4m2), GQ925717 (opn4m3); elephant shark (Callorhinchus milii), JQ172797 (opn4m1), JQ172798 (opn4m2); (xiii) xenopus-like melanopsin (opn4x): chicken (Gallus gallus), EU124630 (OPN4Xlong); lizard (Podarcis siculus), DQ013043; African clawed frog (Xenopus laevis), AF014797; cod (Gadus morhua), AF385823 (opn4xa), AY126448 (opn4xb); zebrafish (Danio rerio), GQ925718 (opn4x1), GQ925719 (opn4x2); elephant shark (Callorhinchus milii), JQ172799 (opn4xlong); (xiv) chordate melanopsin (opn4): lancelet (Branchiostoma belcheri), AB205400; and (xv) outgroup (not shown): human (Homo sapiens), NM005294 (GPR21), NM005684 (GPR52). The gene nomenclature used follows the guidelines adopted by the Entrez Gene database (www.ncbi.nlm.nih.gov/sites/entrez?db=gene). In brief, the genes of all terrestrial species are in uppercase, except for the mouse and rat, where only the first letter is capitalized. The genes of all aquatic species, including amphibians, are in lowercase.

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