Cloning and RNA interference indicate that the Na/K-ATPase gene helps Hong Kong oysters (Crassostrea hongkongensis) resist short-term hypersalinity stress

Na/K-ATPase is a carrier protein embedded in the cell membrane and its main function is to maintain the osmotic pressure balance inside and outside the cell. To investigate the role of Na/K-ATPase under high-salt stress in the Hong Kong oyster (Crassostrea hongkongensis), full-length cloning and expression analysis of Na/K-ATPase were performed, and the function of the gene was verified using RNA interference (RNAi) technology. The full-length sequence was 1,230 bp, including a 5' untranslated region (UTR) of 25 bp; a 3’ UTR of 263 bp; and an open reading frame of 942 bp, encoding a protein of 313 amino acids, with a molecular weight 35.452 KDa and a theoretical isoelectric point 5.55. Reverse transcription-quantitative polymerase chain reaction results showed that Na/K-ATPase is expressed in all tissues of the Hong Kong oyster, with the highest relative expression level in the blood sinus, followed by the gills, and the lowest level of expression in the labellum. Under conditions of acute high-salt stress, the expression level of Na/K-ATPase in the gills increased and then decreased. Using RNAi technology, it was found that Na/K-ATPase enzyme activity in the gills decreased significantly after 72, 96, and 120 h of high-salinity stress, and the survival rate of the RNAi group was lower than that of the blank control group and the negative control group with only 75%. These results indicated that Na/K-ATPase is closely related to osmoregulation in Hong Kong oysters and plays an important role in osmotic pressure regulation and resistance to salinity stress.