Characterization of an evolutionarily conserved calcitonin signaling system in a lophotrochozoan, the Pacific oyster (Crassostrea gigas)

Abstract

In Protostoma, the diuretic hormone 31 (DH31) signaling system was long considered as the orthologue of the chordate calcitonin (CT) signaling system. Using the Pacific oyster (Crassostrea gigas) transcriptomic database GigaTON (http://ngspipelines-sigenae.toulouse.inra.fr/), we characterized seven G-protein-coupled receptors (GPCRs) named Cragi-CTR1/7 and phylogenetically related to chordate CT receptors (CTRs) and to protostome DH31 receptors. Two CT Precursors (Cragi-CTP1 and Cragi-CTP2) containing two CT-type peptides and encoded by two distinct genes with a similar organization were also characterized. These oyster neuropeptides (Cragi-CT1/2) exhibit the two N-terminal paired cysteine residues and except CTP2 derived peptide (Cragi-CTP2dp) the C-terminal proline-amide motif typical of deuterostome CT-type peptides. All mature Cragi-CTs but Cragi-CTP2dp were detected in visceral ganglion (VG) extracts using mass spectrometry. Cell-based assays revealed that the formerly characterized oyster receptors Cg-CTR and Cragi-CTR2 were specifically activated by Cragi-CT1b and Cragi-CT2, respectively. This activation does not require the co-expression of receptor activity-modifying proteins (RAMPs). Thus, the oyster CT signaling appears functionally more closely related to the vertebrate CT/CTR signaling than to the (Calcitonin Gene Related Peptide) CGRP/CLR signaling. Gene expression profiles in different adult tissues and in oysters acclimated to brackish water suggest the potential implication of both Cg-CT-R/Cragi-CT1b and Cragi-CTR2/Cragi-CT2 in water and ionic regulations, though with apparently opposite effects. The present study represents the first comprehensive characterization of a functional CT-type signaling system in a protostome and provides evidence for its evolutionarily ancient origin and its early role in osmotic homeostasis.