Small cytoplasmic domain peptides of natriuretic peptide receptor-C attenuate cell proliferation through Giα protein/MAP kinase/PI3-kinase/AKT pathways

Abstract

The present studies were undertaken to investigate the effect of C-atrial natriuretic peptide (ANP)4–23and several peptide fragments containing 12 amino acids from different regions of the cytoplasmic domain of natriuretic peptide receptor (NPR)-C on cell proliferation in the absence or presence of angiotensin (ANG) II, endothelin (ET)-1, and arginine vasopressin (AVP) in A-10 vascular smooth muscle cells (VSMC). The peptide fragments used have either complete Giactivator sequences K461-H472(peptide 1) and H481-H492(peptide 3) or partial Giactivator sequences R469-K480(peptide 2) and I465-H472(peptide Y) with truncated COOH or NH2terminus, respectively. The other peptide used had no structural specificity (Q473-K480, peptide X) or was the scrambled peptide control for peptide 1 (peptide Z). ANG II, ET-1 and AVP significantly stimulated DNA synthesis in these cells as determined by [3H]thymidine incorporation that was inhibited by peptides 1, 2, and 3 and not by peptides X, Y, and Z in a concentration-dependent manner, with an apparent Kibetween 1 and 10 nM. In addition, C-ANP4–23, which interacts with NPR-C, also inhibited DNA synthesis stimulated by vasoactive peptides; however, the inhibition elicited by C-ANP4–23was not additive with the inhibition elicited by peptide 1. On the other hand, basal DNA synthesis in these cells was not inhibited by C-ANP4–23or the peptide fragments. Furthermore, vasoactive peptide-induced stimulation of DNA synthesis was inhibited by PD-98059 and wortmannin, and this inhibition was potentiated by peptide 1. In addition, peptide 1 also inhibited vasoactive peptide-induced phosphorylation of ERK1/2 and AKT and enhanced expression of Giα proteins. These data suggest that C-ANP4–23and small peptide fragments containing 12 amino acids irrespective of the region of the cytoplasmic domain of NPR-C inhibit proliferative responses of vasoactive peptides through Giα protein and MAP kinase/phosphatidylinositol 3-kinase/AKT pathways.