Cholera toxin action on rabbit renal brush-border membranes inhibits phosphate transport

Cholera toxin was used to enhance ADP-ribosylation of rabbit renal brush-border membranes. Treatment of brush-border membrane sheets with cholera toxin in the presence of NAD resulted in a specific inhibition of the initial phase of Na+-dependent Pi uptake, compared to controls incubated with NAD alone. The Pi uptake was determined after conversion of the membrane sheets to vesicles. The equilibrium uptake of Pi, the Na+-independent uptake of Pi, the Na+-dependent uptake of L-proline and the activities of several brush-border membrane enzymes were not changed. The inhibition of Pi transport was dependent on the presence of both NAD and cholera toxin. Incubation of membrane sheets with [3H]NAD produced acid-stable binding of radioactivity to the membranes and the binding was increased 5-fold by the presence of cholera toxin. The use of [32P]NAD and autoradiography confirmed that the bound radioactivity was associated with several different membrane proteins, and that cholera toxin increased binding to these proteins including three that were not labelled in the absence of the toxin. The specific inhibitory action of cholera toxin on Na+/Pi cotransport is probably mediated by ADP-ribosylation of membrane proteins, suggesting that the Pi transport system can be regulated by ADP-ribosylation, at least in vitro.