Internalization of EGF and transferrin measured as the rate of uptake of 1251labeled ligands was compared in the cell line CCL39 and a mutant derivative, PS120, lacking the Na+/H+ antiport system. N o significant alteration was detected between the two cell lines. In contrast, pretreatment of the mutant cells PS-I20 with 20 mM NH4CI for 30 min to decrease persistently intracellular pH resulted in an increase in 1251-EGF and 1251-transferrin uptake by 60% and 25%, respectively. However, similar NH4CI pretreatment of the parental cell line, CCL-39, which only affected intracellular pH very transiently did not cause an increase of ligand uptake. The binding of '251-EGF to CCL-39 and PS-120 cells with or without NH4CI pretreatment showed that NH4CI pretreatment did not affect EGF binding in either CCL-39 or PS-120 cells. Since cells regulate intracellular pH by ion transport systems, we also examined the role of Na+, K+-ATPase. Ouabain, an inhibitor of Na+, K+-ATPases, showed no effect on 1251-ECF uptake in either of the cell types with or without NH4CI pretreatment. Taken together, these results suggest that the plasma mem- brane-bound Na+/H + antiport, a major pHi-regulating system in vertebrates, indirectly plays a role in ligand internalization through regulation of intracellular pH.
Many ligands, after binding to cell surface receptors, CCL-39 cell line and a mutant, PS-120, derived from are rapidly internalized through coated pits into endo-CCL-39, which lacks the Naf/H+ exchange activity cytic vesicles in a process termed receptor-mediated en- (L'Allemain et al., 1984;Pouyssegur et al., 1984).