Expression and localization analysis of ncc and nkcc genes in gill tissues of Lateolabrax maculatus during salinity adaptation

The ncc gene encodes Na⁺-Cl⁻ synergistic transporter, while the nkcc gene encodes Na⁺-K⁺-2Cl⁻synergistic transporter. Both of them belong to the solute carrier family12 (SLC12), and play key roles in the osmotic regulation mechanism of fish. In order to explore the roles of ncc and nkcc in the osmotic regulation of spotted sea bass in this study, whole genome identification, multiple sequence alignment, phylogenetic tree construction and protein structure prediction were performed to identify the ncc genes of spotted sea bass and analyse their sequence structure. The expression levels of ncc and nkcc genes in gill tissues were detected by qRT-PCR, and the sites of expression of ncc2 and nkcc1a in gills of spotted sea bass in freshwater and seawater were determined by in situ hybridization (ISH). The results showed that two ncc genes, ncc1 and ncc2, were identified in spotted sea bass. Their CDSs length were 2 691 and 3 120 bp, encoding 896 and 1 039 amino acids, respectively. The expression of ncc2 gene in gill tissue of freshwater fish was significantly higher than that in seawater, while the expression of nkcc1a in gill tissue of seawater fish was significantly higher than that in freshwater. Moreover, the expression of ncc2 in gill was gradually up-regulated during freshwater adaptation, while the expression of nkcc1a was gradually down-regulated. Meanwhile, the opposite expression trend was showed in seawater adaptation process. In addition, the ISH results showed that ncc2 and nkcc1a genes were located in the epithelium between adjacent gill lamellae in freshwater and seawater, respectively. The above results indicate that ncc2 and nkcc1a genes encode important Na⁺-Cl⁻ transporter subtypes in the gills of spotted sea bass in freshwater and seawater, respectively. They can be used as specific markers to distinguish the spotted sea bass in freshwater and seawater, and play an important role in osmotic regulation and salinity adaptation.