HYPOTONIC-STIMULATED TAURINE EFFLUX IN SKATE ERYTHROCYTES - REGULATION BY TYROSINE PHOSPHATASE-ACTIVITY

Treatment of skate erythrocytes with FCCP, dinitrophenol, or sodium az ide lowers ATP levels and inhibits Na+-independent taurine uptake afte r hypotonic volume expansion. Inside-out vesicles isolated from hypoto nic volume-expanded cells demonstrate greater Na+-independent taurine uptake, and pretreatment of cells with FCCP abolishes this stimulation . Addition of ATP to the vesicles does not restore stimulated taurine uptake, suggesting that ATP does not act as a ligand modulator on the transporter. Therefore the role of protein phosphorylation was investi gated. Because known protein kinase inhibitors have previously been fo und to have little effect on taurine fluxes in skate erythrocytes, we focused on the effects of protein phosphatase inhibition. When volume- expanded cells were returned to isotonic medium, taurine flux returned to basal values more slowly after treatment with the tyrosine phospha tase inhibitor pervanadate, suggesting that dephosphorylation may regu late inactivation. A similar effect of phosphatase inhibitors was obse rved in the inside-out vesicles from volume-expanded cells: the revers al of stimulated taurine uptake takes place more slowly in vesicles pr epared from cells that had been incubated with pervanadate. Band 3, a major protein involved in the taurine transport pathway, shows increas ed tyrosine phosphorylation after hypotonic volume expansion. Pervanad ate treatment of the cells potentiates and prolongs the increased phos phorylation. Therefore tyrosine phosphorylation of band 3 may play an important role in the activation of taurine fluxes after volume expans ion.