PTH-related protein activates a G protein-coupled PTH/PTHrP receptor in many cell types and produces diverse biological actions. To study the signal transduction events associated with biological activity of the PTH/PTHrP receptor in vascular smooth muscle, a principal PTHrP-responsive tissue, rat aortic smooth muscle cells (A10) were stably transfected with a plasmid encoding a PTH/PTHrP receptor and tested for ligand binding, PTHrP-(1-34)- induced cAMP levels, inositol phosphate production, and cytosolic calcium transients. Of nineteen G418-resistant lines recovered, all exhibited high affinity binding [~ dissociation constant (K(d) >10-10) of iodinated [Tyr36]hPTHrP(1-36)NH2 and ligand-induced cAMP accumulation (2- to 100- fold), which was directly proportional to PTH/PTHrP receptor number (range 4 x 103 to 7 x 107 sites/cell]. PTHrP had no effect on intracellular calcium or inositol phosphate formation in any cell line regardless of receptor number despite the presence of detectable Gα(q). Transient overexpression of individual Gα(q) proteins (Gα(q), Gα11 or Gα14) into PTH/PTHrP receptor-expressing A10 cells conferred the ability of PTHrP to increase intracellular calcium and inositol phosphate formation. Ligand activation of the recombinant PTH/PTHrP receptor elicited appropriate downstream biological effects in A10 cells including inhibition of DNA synthesis and osteopontin messenger RNA (mRNA) expression. Thus, a single PTH/PTHrP receptor, though capable of coupling to different G proteins, signals exclusively through a cAMP-dependent pathway in vascular smooth muscle.
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