Identification of potentially novel functions of DNA polymerase zeta catalytic subunit in oriental river prawn, Macrobrachium nipoponense: cloning, qPCR, in situ hybridization and RNAi analysis

Abstract

The goal of this study was to analyze the functions of DNA polymerase zeta catalytic subunit (Rev3) in the oriental river prawn Macrobrachium nipponense (Mn-Rev3) with a focus on its potential roles in sex differentiation and development. The full length of Mn-Rev3 cDNA sequence was 6832 base pairs (bp) with an open reading frame of 6102 bp encoding 2033 amino acids. Mn-Rev3 showed the closest evolutionary relationship with Penaeus vannamei. The highest expression level of Mn-Rev3 occurred in the hepatopancreas and strong signals were observed in hepatopancreas cells, suggesting that Mn-Rev3 played a role in the immune system. Expression levels of Mn-Rev3 also were relatively high in the androgenic gland and testis, suggesting its potential roles in male sexual differentiation and development. During development, expression of Mn-Rev3 was highest on larval day 15 and relatively high from post-larval day 1 (PL1) to PL15, indicating that it played essential roles in promoting metamorphosis and gonad differentiation and development in M. nipponense. Strong Mn-Rev3 signals were detected in spermatids, spermatocytes, and sperm in the testes, and Mn-Rev3 expression was higher in the testes during the reproductive season than in the non-reproductive season. This result indicated that Rev3 promoted whole testis development, and especially sperm development, in M. nipponense. The expression level of Mn-Rev3 was high from ovary V to ovary II stages, indicating that Rev3 may be involved in yolk deposition. The expression level of Mn-insulin-like androgenic gland hormone (Mn-IAG) and the content of testosterone showed the same expression pattern as that of Mn-Rev3 after injection of double-stranded RNA of Mn-Rev3, which indicated that Rev3 had positive effects on male sexual differentiation and development in M. nipponense. The results of this study advance our understanding of male sexual development in M. nipponense and provide the basis for further studies of male sexual differentiation and development in crustaceans.