Identification and functional analysis of the estradiol 17β-dehydrogenase gene on the shell hardness of Scylla paramamosain during the molting cycle

Estradiol 17 β-dehydrogenase (17E2DH) is an oxidoreductase gene involved in the metabolism of androgen and estrogen in mammals and birds, and is also associated with molting in crustaceans. However, neither the full-length estradiol 17 β-dehydrogenase nor its molecular mechanism during molting has been reported in crabs. In this study, the cDNA encoding 17-beta-dehydrogenase was cloned from the hepatopancreas of Scylla paramamosain and named Sp17E2DH. The full length Sp17E2DH consists of 922 bp with an open reading frame encoding 255 amino acids. Phylogenetic analysis showed that the predicted Sp17E2DH protein is conserved and clustered with the 17E2DH from other crustacean species. RT-qPCR and immunohistochemistry analyses indicated that the expression level of Sp17E2DH was lowest before molting (TQ), up-regulated at 3 h after molting (TH3h), and highest at 48 h after molting (TH48h) in the hepatopancreas and muscles. An RNA interference experiment showed that the mRNA expression level of Sp17E2DH was significantly down-regulated (P < 0.05) and the hardness of crab shells was significantly decreased (P < 0.05) at TH48h in the hepatopancreas and muscles after injection of Sp17E2DH-targeted dsRNA. Genes such as gastrolith protein (P = 0.045), cuticular protein 15 (P = 0.021), chitin synthase (P = 0.047) and troponin C2 (P = 0.013) were significantly suppressed at TH48h after Sp17E2DH dsRNA injection compared to the control groups, which were potentially regulated by Sp17E2DH to control shell hardness of S. paramamosain. In summary, the results suggest that Sp17E2DH plays a vital role in the molting cycle and shell hardening of the mud crab, and the dsRNA interference of Sp17E2DH may be used to prolong the marketing shelf life of live soft-shelled mud crabs.