HeLa cells express cAMP-inhibitable sodium-dependent phosphate uptake
Receptor-mediated regulation of the sodium-phosphate symporter, and hence sodium-dependent phosphate uptake, typically relates to epithelial cells of renal origin. In this study, we have characterized sodium-dependent phosphate uptake and aspects of its receptor-mediated regulation in the HeLa cell line, a cell line derived from a human epitheloid carcinoma. Phosphate uptake (>90% sodium dependent; V(max) = 4.02 ± 0.24 nmol·mg protein-1·3 min-1 and K(m) = 0.11 ± 0.02 mM phosphate at 140 mM sodium) was kinetically similar to that observed in opossum kidney cells. Incubation with vasoactive intestinal peptide (VIP) resulted in a dose-dependent (50% maximal dose of 8.8 ± 3.6 nM) approximately fivefold increase in basal adenosine 3',5'-cyclic monophosphate (cAMP) levels (basal = 14.6 ± 1.7 pmol·mg protein-1·15 min-1; VIP stimulated = 72.7 ± 13.2 pmol·mg protein-1· 15 min-1), as well as a dose-dependent maximal 32.6 ± 5.5% decrease in sodium-dependent phosphate uptake (50% maximal decrease of 46.2 ± 21.2 nM). The VIP-induced decrease in phosphate uptake was due to decrease in maximal transport (V(max VIP) = 2.78 ± 0.16 nmol·mg protein-1·3 min-1) and not to a change in the affinity of the transporter for phosphate (K(m VIP) = 0.11 ± 0.01 mM phosphate). Preincubation of HeLa cells with forskolin and cholera toxin, which stimulate adenylate cyclase, resulted in dose-dependent decreases in sodium-dependent phosphate uptake. Incubation with 8-bromo-cAMP and dibutyryl cAMP, permeant analogues of cAMP, similarly resulted in a dose-dependent decrease in sodium-dependent phosphate uptake. Maximal inhibition of phosphate uptake by each of these agents was ~35% (forskolin = 37.3 ± 1.4; cholera toxin = 34.5 ± 1.5; dibutyryl-cAMP = 36 ± 2.0; 8-bromo-cAMP = 34.7 ± 2.7%). These results demonstrate that sodium-dependent phosphate uptake in HeLa cells is rapidly inhibited by agents which increase intracellular levels of cAMP.
Raymond, JR; Middleton, JP; Dennis, VW
Volume / Issue
International Standard Serial Number (ISSN)