Assessment of the function of gonad-specific PmAgo4 in viral replication and spermatogenesis in Penaeus monodon

Abstract Argonaute family is phylogenetically subdivided into Ago and Piwi subfamilies that operate either transcriptional or post-transcriptional regulation in association with particular types of small RNAs. Among the four members of Ago subfamily (PmAgo1-4) found in black tiger shrimp Penaeus monodon, PmAgo4 exhibits gonad-restricted expression and takes part in transposon repression as the Piwi subfamily. While PmAgo1-3 participate in RNA interference (RNAi)-based mechanism, the role of PmAgo4 in RNAi is still mysterious, and was therefore investigated in this study. The results showed that knockdown of PmAgo4 in shrimp testis did not have a significant effect on the potency of PmRab7 silencing by dsPmRab7. In addition, replication of YHV as well as YHV-induced cumulative mortality in PmAgo4-knockdown shrimp are comparable to the control shrimp, suggesting the irrelevant association of PmAgo4 with RNAi-mediated gene silencing and antiviral immunity. Since PmAgo4 did not function in common with the Ago subfamily, its potential function in gametogenesis of male shrimp was further investigated. The reduction of PmAgo4 transcript levels in male shrimp revealed significant defect in testicular maturity as measured by Testicular Index (TI). Moreover, the numbers of mature sperm in spermatophore of PmAgo4-knockdown shrimp were significantly decreased comparing with the control shrimp. Our studies thus suggest a distinctive role of PmAgo4 that is not consistent with a dsRNA-mediate gene regulation and virus replication, but has a key function in controlling spermatogenesis in P. monodon.