Prolonged atrial natriuretic peptide exposure stimulates guanylyl cyclase-a degradation.
Natriuretic peptide receptor-A (NPR-A), also known as guanylyl cyclase-A, is a transmembrane receptor guanylyl cyclase that is activated by the cardiac hormones atrial natriuretic peptide and B-type natriuretic peptide. Although ligand-dependent NPR-A degradation (also known as down-regulation) is widely acknowledged in human and animal models of volume overload, down-regulation in cultured cells is controversial. Here, we examined the effect of ANP exposure on cellular NPR-A levels as a function of time. Relative receptor concentrations were estimated using guanylyl cyclase and immunoblot assays in a wide variety of cell lines that endogenously or exogenously expressed low or high numbers of receptors. ANP exposures of 1 h markedly reduced hormone-dependent but not detergent-dependent guanylyl cyclase activities in membranes from exposed cells. However, 1-h ANP exposures did not significantly reduce NPR-A concentrations in any cell line. In contrast, exposures of greater than 1 h reduced receptor concentrations in a time-dependent manner. The time required for half of the receptors to be degraded (t(1/2)) in primary bovine aortic endothelial and immortalized HeLa cells was approximately 8 h. In contrast, a 24-h exposure of ANP to 293T cells stably overexpressing NPR-A caused less than half of the receptors to be degraded. To our knowledge, this is the first report to directly measure NPR-A down-regulation in endogenously expressing cells. We conclude that down-regulation is a universal property of NPR-A but is relatively slow and varies with receptor expression levels and cell type.
Flora
Darcy R
DR
University of Minnesota Twin Cities, Department of Biochemistry, Molecular Biology, and Biophysics, 6-155 Jackson Hall, 321 Church Street, South East Minneapolis, Minnesota 55455, USA.
Potter
Lincoln R
LR
D000818:Animals; D009320:Atrial Natriuretic Factor / Q000494:pharmacology*; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D006162:Guanylate Cyclase / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D034741:RNA, Small Interfering; D011863:Radioimmunoassay; D051381:Rats; D017461:Receptors, Atrial Natriuretic Factor / Q000235:genetics / Q000378:metabolism
D034741:RNA, Small Interfering; D009320:Atrial Natriuretic Factor; D006162:Guanylate Cyclase; D017461:Receptors, Atrial Natriuretic Factor; C091715:atrial natriuretic factor receptor A
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