Identification and Sexually Dimorphic Expression of a FoxL2-like Gene in Mud Crab (Scylla paramamosain): Potential Roles in Male Differentiation and Development

Monosex breeding technology has received enormous attention for theScylla paramamosain, an economic crustacean species with distinctive sexual dimorphism. However, the molecular mechanism of gonadal development and sexual differentiation still remain inconclusive. In this study, we reported the cloning, characterization of aFoxL2-likegene with male-biased expression in the mud crab (SpFoxL2). TheSpFoxL2encoded a putative forkhead-box protein with 352 amino acids. Phylogenetic tree revealed theSpFoxL2was clustered with the invertebratesFoxL2homologues, most closed toE. sinensis. By quantitative real-time PCR, it was shown that theSpFoxL2was expressed in intestine, ovary and testis, with the highest expression level in testis. During gonadal development, the expression level ofSpFoxL2in ovary was lower than that in testis at all stages. The expression level ofSpFoxL2constantly declined from stage I to stage V in ovary. While, there is a rising tendency from stage I to stage IV in testis. Notably, this male-biased expression pattern could be detected as early as the juvenile crab stage I (CI stage) before morphological sexual differentiation in the mud crab. By fluorescence in-situ hybridization,SpFoxL2mRNA was found located in both somatic and germ cells in gonads. In testis, it was most strongly localized in the meiosis germ cells (spermatocytes). While in ovary, the signal became weaker in the meiosis germ cells (oocytes). The results imply that theSpFoxl2may play an important role in male differentiation and development, laying a foundation for future research on sexual differentiation of the species.