Identification and functional characterisation of the relaxin-like gonad-stimulating peptide receptor in sea...

Relaxin-like gonad-stimulating peptide (RGP) regulates final gamete maturation and spawning in starfish through a specific gonadal receptor, the RGP receptor (RGPR). Homologous RGPs have since been identified in sea cucumbers and shown to induce spawning, supporting a conserved reproductive role across echinoderms. However, the molecular basis of RGP-RGPR signalling in sea cucumbers remained unknown, as the sea cucumber RGPR had not previously been identified. Therefore, we systematically searched for G protein-coupled receptors similar to starfish RGPRs using comparative sequence analysis and identified candidate RGPRs in five sea cucumber species: Holothuria leucospilota, Holothuria glaberrima, Holothuria scabra, Apostichopus japonicus, and Isostichopus badionotus. Receptor sequences from Holothuria species shared 77 to 85% identity, while similarity to Apostichopus and Isostichopus sequences was lower, at about 60%. Structural modelling revealed the expected features of relaxin family peptide receptors, including seven transmembrane helices, a large extracellular domain with multiple leucine-rich repeats, and an N-terminal LDLa module. Functional cell-based assays demonstrated that the candidate H. scabra RGPR responded specifically and dose-dependently to recombinant H. scabra RGP, but not to human H2 relaxin, providing the first functional evidence of an RGP receptor in a sea cucumber. An RGPR homologue was detected in the H. leucospilota ovary across all reproductive stages, although expression did not differ significantly among stages. Together, these findings support the conservation of relaxin-like reproductive signalling in echinoderms, while direct functional characterisation of the H. leucospilota RGP-RGPR pair remains necessary.