Identification and pathway analysis of immediate hyperosmotic stress responsive molecular mechanisms in tilapia (Oreochromis mossambicus) gill

Salinity is a major environmental factor that strongly influences cellular and organismal function. We have used the euryhaline fish Oreochromis mossambicus to identify and annotate immediate hyperosmotic stress responsive molecular mechanisms and biological processes in gill epithelial cells. Using a suppression subtractive hybridization (SSH) approach, we have identified and cloned 20 novel immediate early genes whose mRNAs are induced in gill epithelial cells 4 h after transfer of fish from freshwater (FW) to seawater (SW). Full-length or partial sequences of open reading frames (ORFs) were obtained using the rapid amplification of cDNA ends (RACE) technique. Kinetics of induction was analyzed for all hyperosmotic stress-induced genes. Most genes show a robust transient increase in mRNA abundance characteristic of immediate early stress response genes with peak levels observed between 2 and 8 h after seawater transfer. The newly identified genes were classified according to their sequence similarity with other vertebrate homologs and based on their predicted functions. Pathway analysis revealed that more than half of the identified immediate hyperosmotic stress genes interact closely within a cellular stress response signaling network. Moreover, the 20 genes cluster together in six molecular processes that are rapidly activated in tilapia gills upon salinity transfer. These processes are (1) stress response signal transduction, (2) compatible organic osmolyte accumulation, (3) energy metabolism, (4) lipid transport and cell membrane protection, (5) actin-based cytoskeleton dynamics, and (6) protein and mRNA stability. Our identification and analysis of a set of novel osmo-responsive tilapia genes provides insight into critical physiological processes and pathways constituting the hyperosmotic stress adaptation program in gill epithelial cells of euryhaline fishes.