FUNCTIONAL EXPRESSION OF A STABLY TRANSFECTED PARATHYROID-HORMONE PARATHYROID-HORMONE RELATED PROTEIN-RECEPTOR COMPLEMENTARY-DNA IN CHO CELLS

Chinese hamster ovary (CHO) cells were stably transfected with OK-O co mplementary DNA encoding the parathyroid hormone/parathyroid hormone r elated protein (PTH/PTHrP) receptor derived from opossum kidney (OK) c ells (Juppner et al., 1991). A subclone of transfected CHO cells, CHO- E(2), presented high affinity binding of I-125-labeled [Tyr(36)]chicke nPTHrP(1-36)amide ([I-125]chPTHrP(1-36)) (K-d 1.28 +/- 0.10 nM) simila r to that of wildtype OK cells (K-d 2.23 +/- 0.16 nM) (P < 0.01). Phot oaffinity labeling of the PTH/PTHrP receptors using N-hydroxysuccinimi dyl-4-azidobenzoate modified [I-125]chPTHrP(1-36) revealed the same sp ecifically labeled 90 kDa protein in CHO-E(2) and OK cells. In CHO-cel ls, chPTHrP(1-36) stimulated cyclic AMP accumulation in dose-dependent fashion (EC(50) 0.15 +/- 0.04 nM) and raised peak cytosolic free calc ium concentration (EC(50), 2.90 +/- 0.36 nM) independent of extracellu lar calcium, and stimulated phosphate uptake (EC(50) 0.21 +/- 0.07 nM) . Both, chPTHrP(1-36) and 12-O-tetradecanoylphorbol-13-acetate stimula ted phosphate uptake were suppressed by staurosporine. But, Sp-cyclic adenosine-3',5'-monophosphothioate did not affect phosphate uptake in CHO-E(2) cells. In conclusion, a PTH/PTHrP receptor stably expressed i n CHO cells is linked to stimulation of phosphate uptake. Receptor cou pling presumably occurred through the protein kinase C rather than the protein kinase A pathway.