Een lost in the common ancestor to mammals. In addition, two
Een lost in the common ancestor to mammals. In addition, two copies of the Type IIa-2 GnRH receptor are present in teleosts, indicating the retention of a duplicate copy in a teleost ancestor. A third Type IIa-2 receptor is unique to pufferfish, which indicates at least one additional lineage specific duplication event (Figure 5). The Type IIa-3 receptor is present only in coelacanths, amphibians, reptiles, and some mammals. The relatively recent common ancestry between the Type IIa-2 and IIa-3 receptors combined with the nested taxonomic distribution for Type IIa-3 receptors suggests it is the youngest subfamily. Topological position and taxonomic distribution indicate the Type IIa-3 subfamily arose through duplication of a Type IIa-2 receptor in early sarcopterygians. The absence of Type IIa-3 receptors in birds and multiple clades of mammals indicates additional lineage specific losses of this receptor subtype. Finally, the Type IIb GnRH receptor subfamily is present in all vertebrate clades surveyed, with the exception of mammals. As observed with the Type IIa-2 PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/25681438 receptors, a pair of Type IIb GnRH receptor genes was present in both zebrafish and pufferfish. Ancestral teleosts appear to have harbored only the Type IIa-2 and IIb subfamilies. Both subfamilies are typically single copy, but each was found as a pair of duplicate genes in teleosts, consistent with the teleost genome duplication event. The single copy genes for the Type IIb subfamily in medaka and gourami could be the result of independent gene loss. However, many teleost Type IIb receptors were identified on small contigs that lacked syntenic neighboring genes, so gene absence could be a technical artifact of genes missing from each genome database. Additional results from BLAST searches of the chimaera, spotted gar (Lepisosteus oculatus), little skate, and lamprey (Petromyzon marinus) genomes also resulted in the identification of partial sequences (less than 60 amino acids) for putative homologs. The small size of these sequences precluded their use in MK-5172 site phylogenetic analyses. These four species are found at important topological positions with respect to the origin of GnRH receptor clades, but the presence or absence of homologs based only on sequences used for phylogenetic analyses constituted a potentially biased sample. Apparent gene absence could be due sub-sampling during targetedWilliams et al. BMC Evolutionary Biology 2014, 14:215 http://www.biomedcentral.com/1471-2148/14/Page 4 ofFigure 2 cDNA PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/28914615 and deduced amino acid sequence of the GnRH receptor IIa-3 gene in axolotls. Analysis and formatting as described in Figure 1.cloning using degenerate PCR primers. Apparent absence could also be a technical artifact from low sequence coverage, consistent with the observation that the genome assemblies for these taxa largely comprise small contigs. To address this issue, we used probabilistic methods to determine sequence homology for short peptide sequences. Specifically, we constructed Hidden Markov Models (HMM) corresponding to four different sequence regions nested within a canonical GnRH receptor, where the size and location of each region corresponded with peptides consistently identified in BLAST searches (termed TM1-4, TM4-5, TM6 and TM6-7 in Table 1, Additional file 2: Table S2 and Additional file 3: Table S3). The HMM profiles were further refined into `types’ based on the taxonomic subset of input GnRH receptor sequences used for HMM construction with HMMER v. 3.1.