Receptor deorphanization in an echinoderm reveals kisspeptin evolution and relationship with SALMFamide neuropeptides

Abstract

Background Kisspeptins are neuropeptides that regulate reproductive maturation in mammals via G-protein-coupled receptor-mediated stimulation of gonadotropin-releasing hormone secretion from the hypothalamus. Phylogenetic analysis of kisspeptin-type receptors indicates that this neuropeptide signaling system originated in a common ancestor of the Bilateria, but little is known about kisspeptin signaling in invertebrates. Results Contrasting with the occurrence of a single kisspeptin receptor in mammalian species, here, we report the discovery of an expanded family of eleven kisspeptin-type receptors in a deuterostome invertebrate — the starfish Asterias rubens (phylum Echinodermata). Furthermore, neuropeptides derived from four precursor proteins were identified as ligands for six of these receptors. One or more kisspeptin-like neuropeptides derived from two precursor proteins (ArKPP1, ArKPP2) act as ligands for four A. rubens kisspeptin-type receptors (ArKPR1,3,8,9). Furthermore, a family of neuropeptides that act as muscle relaxants in echinoderms (SALMFamides) are ligands for two A. rubens kisspeptin-type receptors (ArKPR6,7). The SALMFamide neuropeptide S1 (or ArS1.4) and a ‘cocktail’ of the seven neuropeptides derived from the S1 precursor protein (ArS1.1-ArS1.7) act as ligands for ArKPR7. The SALMFamide neuropeptide S2 (or ArS2.3) and a ‘cocktail’ of the eight neuropeptides derived from the S2 precursor protein (ArS2.1-ArS2.8) act as ligands for ArKPR6. Conclusions Our findings reveal a remarkable diversity of neuropeptides that act as ligands for kisspeptin-type receptors in starfish and provide important new insights into the evolution of kisspeptin signaling. Furthermore, the discovery of the hitherto unknown relationship of kisspeptins with SALMFamides, neuropeptides that were discovered in starfish prior to the identification of kisspeptins in mammals, presents a radical change in perspective for research on kisspeptin signaling.

Discovery and functional characterization of neuropeptides in crinoid echinoderms

“…The occurrence of kisspeptin-type precursors in echinoderms was first reported with the identification of a protein in A. rubens (ArKPP), which comprises two kisspeptin-like peptides ( Semmens et al, 2016 ), and orthologs of this protein have been identified in other echinoderms ( Zandawala et al, 2017 ; Chen et al, 2019 ). Recently, a second kisspeptin-type precursor, also comprising two kisspeptin-like peptides, was identified in A. rubens and other echinoderms ( Escudero Castelán et al, 2022 ). Therefore, these two precursor types are now referred to as KPP1 and KPP2, respectively, and here we report their identification in crinoids.…”

“…The A. mediterranea kisspeptin-type precursor 1 (AmKPP1) is a 189-residue protein that comprises two putative kisspeptin-like peptides (AmKP1.1, AmKP1.2), both of which are predicted to be C-terminally amidated ( Figure 2C ). Accordingly, kisspeptin-type precursors in eleutherozoans also comprise two putative C-terminally amidated kisspeptin-type peptides (e.g., ArKP1.1 and ArKP1.2 in A. rubens ; Semmens et al, 2016 ; Escudero Castelán et al, 2022 ). Alignment of crinoid KP1.1-type peptides with eleutherozoan KP1.1-type peptides revealed a conserved C-terminal Arg-X-Leu-Pro-Phe-NH 2 (RXLPFamide, where X is variable) motif and a conserved Asn (N) residue in the core of the peptide sequences ( Figure 5 ).…”

“…Two kisspeptin-type precursors (KPP1 and KPP2) have been identified in eleutherozoan echinoderms and both precursors comprise two kisspeptin-like peptides ( Semmens et al, 2016 ; Escudero Castelán et al, 2022 ). Here we report for the first time the identification KPP1-type and KPP2-type precursors in crinoids.…”

“…Here we report for the first time the identification KPP1-type and KPP2-type precursors in crinoids. Analysis of the sequences of KPP1-type precursors in eleutherozoans has revealed the presence of two predicted neuropeptides, which we refer to as KP1.1 and KP1.2 ( Escudero Castelán et al, 2022 ). KP1.1-type peptides are preceded by an amino acid sequence that contains two cysteine residues, which was originally thought to form the N-terminal region of the mature bioactive peptide ( Semmens et al, 2016 ).…”

“…The A. mediterranea kisspeptin-type precursor 2 (AmKPP2) is a 153-residue protein that comprises a single putative kisspeptin-like peptide (AmKP2), which is predicted to be C-terminally amidated (Figure 2D). This contrasts with kisspeptin-type precursors in eleutherozoans that comprise two putative C-terminally amidated kisspeptin-type peptides (e.g., ArKP2.1 and ArKP2.2 in A. rubens; Semmens et al, 2016;Escudero Castelán et al, 2022). Sequence alignment revealed that crinoid KP2-type peptides have a C-terminal Gly-Met-X-NH 2 motif (where X is variable) and a histidine (H) residue in the core of the peptides that is also a feature of one or more eleutherozoan KP2.1-type peptides.…”