RNAi Analysis of Potential Functions of Cyclin B3 in Reproduction of Male Oriental River Prawns (Macrobrachium nipponense)

SIMPLE SUMMARY: The rapid gonad reproduction of hatchlings restricts the sustainable development of Macrobrachium nipponense. Thus, it is urgently required to establish an artificial technique to regulate the process of gonad reproduction in M. nipponense. A previous study predicted that Cyclin B3 (CycB3) may perform crucial functions in the regulation of male reproduction in M. nipponense. In the present study, we aimed to investigate the potential roles of CycB3 in the male reproduction of this species. qPCR analysis results suggested that CycB3 was involved in the process of spermiogenesis, oogenesis, and embryogenesis in M. nipponense. RNA interference analysis showed that CycB3 affected the expression of insulin-like androgenic gland hormone and inhibited testis reproduction in M. nipponense. Taken together, these findings suggest that CycB3 plays essential roles in the regulation of male reproduction in M. nipponense, promoting the studies of the regulation of testis reproduction. ABSTRACT: Cyclin B3 (CycB3) is involved in the metabolic pathway of the cell cycle, playing essential roles in the regulation of cell proliferation and mitosis. CycB3 is also predicted to be involved in the reproduction of male oriental river prawns (Macrobrachium nipponense). In this study, the potential functions of CycB3 in M. nipponense were investigated using quantitative real-time PCR, RNA interference, and histological observations. The full-length DNA sequence of CycB3 in M. nipponense was 2147 base pairs (bp) long. An open reading frame of 1500 bp was found, encoding 499 amino acids. A highly conserved destruction box and two conserved cyclin motifs were found in the protein sequence of Mn-CycB3. Phylogenetic tree analysis revealed that this protein sequence was evolutionarily close to that of CycB3s of crustacean species. Quantitative real-time PCR analysis results suggested that CycB3 was involved in the process of spermiogenesis, oogenesis, and embryogenesis in M. nipponense. RNA interference analysis showed that CycB3 had a positive regulatory relationship with insulin-like androgenic gland hormone (IAG) in M. nipponense. In addition, sperm were rarely observed in the testis of double-stranded CycB3-injected prawns after 14 days of treatment, and sperm abundance was dramatically lower than that in the double-stranded GFP-injected prawns on the same day. This result indicated that CycB3 can regulate the testis reproduction in M. nipponense through inhibiting the IAG expressions. Overall, these results indicated that CycB3 plays essential roles in the regulation of male reproduction in M. nipponense, which may promote the studies of male reproduction in other crustacean species.