Phosphate concentration and transport in Ehrlich ascites tumor cells: effect of sodium

The effects of extracellular Pi and Na+ on cellular Pi concentration and transport were studied. Steady-state Pi exchange flux was measured by 32P uptake in the presence and absence of Na+. Model experiments were also conducted to assess the possibility that hydrolysis of organic phosphate esters contributes to the chemically measured intracellular Pi concentration of Ehrlich ascites tumor cells. The results of these experiments indicate that hydrolysis of labile organic phosphate esters does not contribute to the measured intracellular pool of Pi. The Pi transport system exhibits an apparent Ks of 0.115 mM Pi and a maximal flux of 1.73 mmole min-1 (kg dry wt)-1. When incubated in a phosphate-buffered choline chloride medium (5 mM Pi) the intracellular Pi and the Pi influx fall by 65 and 88%, respectively. At 5 mM extracellular Pi, the Na+-dependent component of Pi transport fits Michaelis-Menten kinetics with the maximal flux equal to 2.46 mmole min-1 (kg dry wt)-1 and an apparent Ks of 35.4 mM Na+. In addition, a Na+-independent component of Pi transport, comprising about 12% of the total Pi flux, was identified. The data support the hypothesis that a Pi transport system, dependent on Na+, plays a principal role in the maintenance of intracellular Pi concentration.