Cloning and regulation of expression of the Na+–Cl––taurine transporter in gill cells of freshwater Japanese eels

Abstract

Our previous studies have demonstrated the hypertonic-induced expression of osmotic stress transcription factor and the regulatory volume increase (RVI) response in gill cells isolated from freshwater eels. In this study, we aimed to clone one of the organic osmolyte transporters, the Na+-Cl--taurine transporter (TauT), and to characterize its expression in anisosmotic conditions, using both in vivo and in vitro approaches. A cDNA clone encoding TauT was isolated from gill tissues of Japanese eels, Anguilla japonica. The deduced amino acid sequence shows 88-90% identity to other reported piscine TauT sequences. Our data indicated that TauT mRNA was detectable in both freshwater and seawater fish gills. The expression level of TauT mRNA increased in gills of seawater-acclimating fish. A high abundance of TauT protein was found to be localized in seawater gill chloride cells. Using primary gill cell culture, expression of the gene was induced when the ambient osmolarity was raised from 320 to 500 mosmol l(-1). Hypertonic treatment of the culture caused an increase of F-actin distribution in the cell periphery. Treatment of the cells with colchicine or cytochalasin D significantly reduced TauT transcript level following hypertonic exposure. The inhibition of myosin light chain (MLC) kinase by ML-7 had a significant additive effect on hypertonic-induced TauT expression. Collectively, the data of this study reveal, for the first time, the regulation of TauT expression in gill cells of euryhaline fish. We have demonstrated the involvement of ionic strength, the cytoskeleton and MLC kinase in the regulation of TauT expression. The results shed light on the osmosensing and hyperosmotic adaptation in fish gills.