Characteristics of phosphate uptake by Ehrlich ascites tumor cells

The characteristics of Pi transport across the plasma membrane have been studied in intact cells of the rapidly proliferating Ehrlich ascites tumor cell line. Uptake appears to proceed via a single carrier-mediated process displaying Michaelis-Menten kinetics. The observed uptake is insensitive to sulfhydryl group reagents and, therefore, distinct from mitochondrial Pi transport systems. Moreover, inhibitors of other plasma membrane anion transport systems (i.e. those for lactate, Cl^-, or SO₄^2-) are without effect on Pi uptake. Phosphate analogs with either one or two negative charges (AsO₄^-, FPO₃^2-) markedly inhibit uptake. Pi uptake rates do not appear to depend directly on cellular ATP levels. Moreover, the absence of notable effects of valinomycin, nigericin, or ouabain on Pi uptake indicate that neither a transmembrane electrical potential nor a Na⁺ chemical gradient is required for normal rates of Pi uptake. However, small amounts of Na⁺ (or Li⁺) are required in the external medium, and the protonophoric uncoupling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone greatly stimulates Pi uptake. In addition, dicyclohexylcarbodiimide, an inhibitor of H⁺ translocation, markedly suppresses Pi uptake. The rate of glycolysis in Ehrlich cells is stimulated only slightly by the addition of Pi in the physiological range. Consistent with this observation, the Pi analog arsenate has no effect on the rate of glycolysis under conditions where Pi uptake is substantially inhibited. These results suggest that Pi uptake by Ehrlich ascites tumor cells occurs exclusively via an independent carrier-mediated process which may involve co-transport of both H⁺ and Na⁺. They indicate also that the rate of Pi uptake by this carrier does not limit the rate of glycolysis under physiological conditions.