Tyrosine phosphorylation selectively regulates renal cellular phosphate transport. Evidence that it mediates the stimulatory effect of insulin-like growth factor-1.

Abstract

The signal transduction mechanism responsible for the stimulation of the transport of inorganic phosphate (Pi) in response to mitogens is not known. In the present study, the changes in both Pi transport and tyrosine phosphorylation activity were determined in response to orthovanadate (VO4), an insulin-like agent and genistein, a specific inhibitor of tyrosine kinase activity. The results indicate that in opossum kidney epithelia, VO4 stimulated and genistein inhibited Pi transport dose dependently. The characteristics of the VO4 effect were quite similar to those described for insulin-like growth factor-1 (IGF-1) in terms of time course, selectivity (no effect on the Na-alanine transport), and dependency of the de novo synthesis of proteins. The effects of VO4 and IGF-1 on Pi transport, when tested at submaximal and maximal concentrations, respectively, were not additive suggesting that these two agents act through a common regulatory mechanism. As previously shown with IGF-1, the VO4 effect on Pi transport was additive to that of the maximal effect of Pi deprivation. Changes in tyrosine phosphorylation activity were tested in purified plasma membrane isolated from confluent OK cells using the polymer Glu:Tyr (4:1) as exogenous substrate. VO4 markedly enhanced whereas genistein inhibited the tyrosine kinase activity. The VO4 effect was dose dependent (0.1-1.0 mM), a concentration range similar to that eliciting the Pi transport response. The change in Pi transport was highly correlated with the variation in the tyrosine kinase activity induced by either vanadate (R = 0.969 P less than 0.01) or genistein (R = 0.992, P less than 0.01). In conclusion, VO4, an insulin-like agent and genistein, a specific inhibitor of tyrosine kinase activity selectively altered cellular Pi transport. These changes in Pi transport were associated with a dose-related alteration in tyrosine kinase activity measured in purified plasma membrane. The characteristics of the vanadate effects on Pi transport are similar to those reported for IGF-1 suggesting an important role of this signal transduction mechanism in mediating changes in Pi transport in response to mitogenic factors such as IGF-1.