Evolutionary conservation and divergence of CXCL17 orthologs: functional evidence in reptiles and loss in the avian lineage

The mucosal chemoattractant C-X-C motif chemokine ligand 17 (CXCL17) was recently identified as a ligand for the orphan G protein-coupled receptor 25 (GPR25). Although our recent studies have identified non-mammalian CXCL17 orthologs in fishes and amphibians, their presence in reptiles and birds remains unclear. In this study, we employed bioinformatic searches based on gene synteny and sequence features to identify reptilian and avian CXCL17 orthologs in public databases. We identified intact CXCL17 orthologs in 46 reptilian species, including lizards, snakes, turtles, and alligators. In contrast, we found only CXCL17 gene relics in 22 bird species, suggesting the avian lineage lost functional CXCL17 during evolution. A recombinant reptilian CXCL17 from the loggerhead turtle (Caretta caretta), termed Cc-CXCL17, directly bound to and efficiently activated its corresponding receptor, Cc-GPR25, in a C-terminal fragment-dependent manner. Activation of Cc-GPR25 by recombinant Cc-CXCL17 also induced chemotactic movement of transfected human embryonic kidney (HEK) 293T cells. In cross-species activity assays, recombinant CXCL17s from other species had almost no activity towards Cc-GPR25; Cc-CXCL17 and human CXCL17 had low activity towards mallard GPR25, but all tested CXCL17s had no detectable activity towards chicken GPR25. Our findings demonstrate the presence of functional CXCL17 orthologs in extant reptiles and provide evidence for their evolutionary loss in the avian lineage, offering new insights into the phylogenetic distribution of the newly identified CXCL17-GPR25 signaling system.