Acute down-regulation of sodium-dependent phosphate transporter NPT2a involves predominantly the cAMP/PKA pathway as revealed by signaling-selective parathyroid hormone analogs

Abstract

The parathyroid hormone (PTH)/PTH-related peptide (PTHrP) receptor (PTHR1) in cells of the renal proximal tubule mediates the reduction in membrane expression of the sodium-dependent Pi co-transporters, NPT2a and NPT2c, and thus suppresses the re-uptake of Pi from the filtrate. In most cell types, the liganded PTHR1 activates GαS/adenylyl cyclase/cAMP/PKA (cAMP/PKA) and Gαq/11/phospholipase C/phosphatidylinositol 1,4,5-trisphosphate (IP3)/Ca2+/PKC (IP3/PKC) signaling pathways, but the relative roles of each pathway in mediating renal regulation Pi transport remain uncertain. We therefore explored the signaling mechanisms involved in PTH-dependent regulation of NPT2a function using potent, long-acting PTH analogs, M-PTH(1-28) (where M = Ala1,12, Aib3, Gln10, Har11, Trp14, and Arg19) and its position 1-modified variant, Trp1-M-PTH(1-28), designed to be phospholipase C-deficient. In cell-based assays, both M-PTH(1-28) and Trp1-M-PTH(1-28) exhibited potent and prolonged cAMP responses, whereas only M-PTH(1-28) was effective in inducing IP3 and intracellular calcium responses. In opossum kidney cells, a clonal cell line in which the PTHR1 and NPT2a are endogenously expressed, M-PTH(1-28) and Trp1-MPTH(1-28) each induced reductions in 32P uptake, and these responses persisted for more than 24 h after ligand wash-out, whereas that of PTH(1-34) was terminated by 4 h. When injected into wild-type mice, both M-modified PTH analogs induced prolonged reductions in blood Pi levels and commensurate reductions in NPT2a expression in the renal brush border membrane. Our findings suggest that the acute down-regulation of NPT2a expression by PTH ligands involves mainly the cAMP/PKA signaling pathway and are thus consistent with the elevated blood Pi levels seen in pseudohypoparathyroid patients, in whom Gαs-mediated signaling in renal proximal tubule cells is defective. © 2011 by The American Society for Biochemistry and Molecular Biology, Inc.