Isolation and functional characterization for oocyte maturation and sperm motility of the oocyte maturation arresting factor from the Japanese scallop, Patinopecten yessoensis

Abstract

In bivalves, serotonin (5-hydroxytriptamine, 5-HT) acts as a major promotional factor in oocyte maturation, sperm motility, and sequential spawning. The previously reported novel neuronal protein, oocyte maturation arresting factor (OMAF) that was found in the central nervous system and hemolymph of the Japanese scallop, Patinopecten yessoensis, has an inhibitory activity in the 5-HT-induced oocyte maturation via a receptor-mediated mechanism, resulting in an arrest of spawning [30]. In this study, OMAF protein was isolated from the supernatant of hemolymph of the scallop using gel and anion-exchange chromatography, and SDS-PAGE. Three digested partial peptides with 4, 11, and 16 amino acid residues were determined through reversed-phase HPLC and amino acid sequencing. The anti-OMAF antibodies generated against the obtained peptides with 11 and 16 amino acid residues were applied to immunohistochemistry and 5-HT-induced spawning and oocyte maturation assays. Fusiform OMAF neurons were localized in the external area of the anterior lobe of the cerebral ganglion, supporting our presumption that OMAF was secreted from the cerebral and pedal ganglia (CPG). Pretreatment with anti-OMAF antibody on three kinds of bivalve species showed a strong in vivo amplification of 5-HT-induced release of egg and sperm, and an in vitro restoration of 5-HT-induced germinal vesicle breakdown (GVBD) from inhibition by the CPG extract, suggesting the release from suppressive activity of OMAF due to the absorption with antibody. These results confirm that the isolated peptides are from OMAF and OMAF acts as an inhibitor of 5-HT-induced oocyte maturation and sperm motility as previously reported.