Potential role of Methoprene-tolerant (Met) in methyl farnesoate-mediated vitellogenesis in the Chinese mitten crab (Eriocheir sinensis).

Abstract

Methoprene-tolerant (Met) belongs to the basic helix-loop-helix (bHLH)-Per-Arnt-Sim (PAS) family of nuclear transcriptional regulators and is a leading candidate receptor for juvenile hormone (JH III) in insects. Methyl farnesoate (MF) is a de-epoxide form of JH III that regulates many developmental processes in crustaceans, including reproduction, molting, and morphogenesis, much like JH III in insects. In this study, the full-length cDNA for Met was cloned from the Chinese mitten crab (Eriocheir sinensis) (EsMet). The amino acid sequence of EsMet contains three conserved domains (bHLH, PAS-A, and PASB) characteristic of the bHLH-PAS family, having six conserved amino acid residues specifically responsible for JH or MF binding. Tissue distribution analysis revealed that EsMet mRNA is highly expressed in the hepatopancreas. In addition, EsMet and EsVg expression in the hepatopancreas were found to be significantly increased in early endogenous vitellogenic oocytes (stage II) during ovarian development, and the hemolymph MF titer was significantly increased in late exogenous vitellogenic oocytes (stage III), indicating that EsMet is involved in vitellogenesis regulation. In vitro, MF addition markedly upregulated EsMet and EsVg expression in hepatopancreatic tissue, but only EsVg was induced in ovarian tissue. In vivo, EsMet and EsVg expression in the hepatopancreas were both significantly and synchronously increased after MF injection, but not in the ovaries. In addition, EsMet and EsVg expression were upregulated in the hepatopancreas after eyestalk ablation, while only EsVg expression was induced in the ovaries. Thus, our results indicate that Met may act as a receptor for MF in MF-mediated vitellogenesis in crustaceans.