Gi-1/Gi-2-dependent signaling by single-transmembrane natriuretic peptide clearance receptor

Abstract

Single-transmembrane natriuretic peptide clearance receptor (NPR-C), which is devoid of a cytoplasmic guanylyl cyclase domain, interacts with pertussis toxin (PTx)-sensitive G proteins to activate endothelial nitric oxide synthase (eNOS) expressed in gastrointestinal smooth muscle cells. We examined the ability of NPR-C to activate other effector enzymes in eNOS-deficient tenia coli smooth muscle cells; these cells expressed NPR-C and NPR-B but not NPR-A. Atrial natriuretic peptide (ANP), the selective NPR-C ligand cANP-(4–23), and vasoactive intestinal peptide (VIP) inhibited125I-ANP and 125I-VIP binding to muscle membranes in a pattern indicating high-affinity binding to NPR-C. Interaction of VIP with NPR-C was confirmed by its ability to inhibit125I-ANP binding to membranes of NPR-C-transfected COS-1 cells. In tenia muscle cells, all ligands selectively activated Gi-1 and Gi-2; VIP also activated Gs via VIP2 receptors. All ligands stimulated phosphoinositide hydrolysis, which was inhibited by ANP-(1–11), PTx, and antibodies to phospholipase C-β3 (PLC-β3) and Gβ. cANP-(4–23) contracted tenia muscle cells; contraction was blocked by U-73122 and PTx and by antibodies to PLC-β3 and Gβ in intact and permeabilized muscle cells, respectively. VIP and ANP contracted muscle cells only after inhibition of cAMP- and cGMP-dependent protein kinases. ANP and cANP-(4–23) inhibited forskolin-stimulated cAMP in a PTx-sensitive fashion. We conclude that NPR-C is coupled to activation of PLC-β3 via βγ-subunits of Gi-1 and Gi-2 and to inhibition of adenylyl cyclase via α-subunits.