Intracellular pH regulation of SV-40 virus transformed human MRC-5 fibroblasts and cell membrane cholesterol.

Abstract

Alterations in membrane cholesterol could affect the activity of various membrane transporters, including the Na(+)-H+ antiport. The effect of cellular cholesterol depletion (with phosphatidylcholine liposomes) and enrichment (with cholesterol and phosphatidylcholine liposomes) on cellular pH regulation was studied in SV-40 virus transformed human MRC-5 fibroblasts. Cellular cholesterol depletion led to activation of the Na(+)-H+ antiport by an increased maximal velocity (Vmax) of the transporter, with no changes in the apparent dissociation constant (Kd) or Hill coefficient for intracellular H+. Cholesterol enrichment had no effect on the activation of the Na(+)-H+ antiport by intracellular acidosis. However, activation of the Na(+)-H+ antiport by an osmotic stimulus was enhanced in cholesterol-depleted cells and reduced in cholesterol-enriched cells. Liposomes that had no effect on cellular cholesterol did not alter the activation of Na(+)-H+ antiport activity by intracellular acidosis or an osmotic stimulus. Thus in situ modification of cellular cholesterol altered Na(+)-H+ antiport activity differently depending on the type of activating stimulus.