Molecular evidence for farnesoic acid O-methyltransferase (FAMeT) involved in the biosynthesis of vitellogenin in the Chinese mitten crab Eriocheir sinensis

Abstract

Sesquiterpenoid methyl farnesoate (MF), a crustacean equivalent of insect juvenile hormone (JH III), has essential functions in regulating physiological processes in crustaceans, including reproduction and vitellogenesis. Farnesoic acid O-methyltransferase (FAMeT) is a key rate-limiting enzyme catalyzing the conversion of farnesoic acid (FA) to JH/MF in insects and crustaceans. In this study, a full-length cDNA of EsFAMeT from Eriocheir sinensis was isolated and characterized. The deduced EsFAMeT amino acid sequence indicated there were two conserved Methyltransf-FA domains characteristic of FAMeT family proteins. With use of sequence alignment analysis procedures, there was an indication that FAMeT proteins are highly conserved among crustaceans and FAMeT is more closely related to crustacean FAMeT than to insect FAMeT. Results from quantitative real-time PCR analysis revealed there was ubiquitous EsFAMeT in all tissues examined, with greater abundances of mRNA transcripts in the ovary. The transcription of EsFAMeT indicated there were stage-specific patterns in the hepatopancreas and ovary during ovarian development, with the greatest abundance during ovarian development Stages II and III, respectively. To investigate functions of EsFAMeT in vitellogenin biosynthesis in E. sinensis, RNA interference-mediated gene knockdown was used in vitro and in vivo. Injection of EsFAMeT dsRNA resulted in a marked decrease in EsVg (encoding vitellogenin) transcripts in the ovary and hepatopancreas both in vitro and in vivo. Results from the present study indicated EsFAMeT is involved in vitellogenin biosynthesis in the ovary and hepatopancreas of E. sinensis, providing a new resource to study modulatory effects of the FAMeT family of enzymes in crustacean reproduction.