Functional characterisation of neuropeptides that act as ligands for both calcitonin-type and pigment-dispersing factor-type receptors in a deuterostome

The calcitonin (CT) family of related peptides exerts diverse physiological effects in mammals via two G-protein coupled receptors: CTR and the CTR-like receptor CLR. Phylogenetic analysis of CT-type signaling has revealed the presence of CT-type peptides and CTR/CLR-type proteins in both deuterostome and protostome invertebrates. Furthermore, experimental studies have demonstrated that in the protostome Drosophila melanogaster the CT-like peptide DH31 can act as a ligand for a CTR/CLR-type receptor and a pigment-dispersing factor (PDF) receptor. Here we investigated the signaling mechanisms and functions of CT-type neuropeptides in a deuterostome invertebrate, the sea cucumber Apostichopus japonicus (phylum Echinodermata). In A. japonicus, a single gene encodes two CT-type peptides (AjCT1 and AjCT2) and both peptides act as ligands for a CTR/CLR-type receptor (AjCTR) and two PDF-type receptors (AjPDFR1, AjPDFR2), but with differential activation of downstream cAMP/PKA, Gq/Ca2+/PKC and ERK1/2 signaling pathways. AjCT1/AjCT2-encoding transcripts were detected in the central nervous system and a variety of organ systems and neuropeptide expression was visualized immunohistochemically using an antiserum to a starfish CT-type peptide (ArCT). In vitro pharmacological experiments demonstrated that AjCT1 and/or AjCT2 cause dose-dependent relaxation of longitudinal muscle and intestine preparations. Furthermore, in vivo pharmacological experiments, combined with gain-and loss-of-function experiments, revealed a potential physiological role for AjCT2/AjPDFR2 signaling in promoting feeding and growth in A. japonicus. To our knowledge, this is the first study to obtain evidence that CT-type peptides can act as ligands for both CTR/CLR-type and PDF-type receptors in a deuterostome. Moreover, it provides the first evidence for appetite-stimulating and growth-promoting effects of CT-type neuropeptides in bilaterians. Given the economic importance of A. japonicus as a foodstuff, the discovery of CT-type peptides as potential regulators of feeding and growth in this species may offer novel strategies for aquaculture applications. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=128 SRC="FIGDIR/small/610391v3_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@663885org.highwire.dtl.DTLVardef@108bea3org.highwire.dtl.DTLVardef@11bd14dorg.highwire.dtl.DTLVardef@14a2008_HPS_FORMAT_FIGEXP M_FIG C_FIG A schematic showing proposed molecular mechanisms by which CT-type signaling regulates feeding and growth in A. japonicus. AjCT1 and AjCT2 are represented by purple and yellow circles, respectively. For cell signaling, black arrows indicate that this process is activated, whilst the pink arrow indicates that ERK1/2 is activated in the AjCT1/AjPDFR1, AjCT2/AjPDFR1 and AjCT2/AjPDFR2 signaling pathways.