NaPO (4) cotransport type III (PiT1) expression in human embryonic kidney cells and regulation by PTH
Fernandes, I.; Béliveau, R.; Friedlander, G.; Silve, C.
American Journal of Physiology 277(4): F543-F551
1999
ISSN/ISBN: 0002-9513 PMID: 10516278 Document Number: 498433
The aim of the present study was to characterize the type(s) of NaPO4 cotransporter expressed in the human renal cell line HEK-293 and its regulation by parathyroid hormone (PTH) in wild-type cells and in cells transfected by the PTH/PTH-related protein (PTHrP) receptor. The results showed that human embryonic kidney HEK-293 cells expressed NaPO4 cotransporter type III (PiT1) mRNA and protein. In contrast, type I (NPT1) or II (NPT2) cotransporter mRNA were not expressed. Na+-dependent phosphate uptake followed a Michaelis-Menten model (apparent maximal transport rate and affinity constant: 23.32 ± 0.69 nmol PO4.mg protein-1.10 min-1 and 0.147 ± 0.014 mM KH2PO4, respectively), was stimulated by phosphate deprivation (maximal increase 24.5 ± 0.8%, P < 0.001, after 15 h of phosphate deprivation), and was inhibited by increasing pH (3.6 ± 0.2-fold decrease at pH 8.5, P < 0.0001). It was inhibited in a time- and concentration-dependent fashion by PTH in HEK-293 cells stably transfected by PTH/PTHrP receptors but not in parental HEK-293 cells. Maximal inhibition of Na+-dependent phosphate transport was observed at 30 min after the addition of 72 nM PTH-(1-34) (31.5 ± 2.4% inhibition, P < 0.01). PTH inhibition of phosphate transport was maintained in phosphate-deprived cells and reversed by both GF109203X (10-6 M) or staurosporine (5.5 nM), two protein kinase C inhibitors. Na+-dependent phosphate uptake was also significantly inhibited by phorbol 12-myristate 13-acetate (20.9 ± 3.9% inhibition, P < 0.001) but not by dibutyril-cAMP (10-4 M) or forskolin (50 mM). The physiological role played by type III NaPO4 cotransport expression in the overall renal regulation of phosphate homeostasis remains to be established. Reprinted by permission of the publisher.