Forced homodimerization by site-directed mutagenesis alters guanylyl cyclase activity of natriuretic peptide receptor B

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Item Type:	Article
Title:	Forced homodimerization by site-directed mutagenesis alters guanylyl cyclase activity of natriuretic peptide receptor B
Creators Name:	Langenickel, T., Buttgereit, J., Pagel, I., Dietz, R., Willenbrock, R. and Bader, M.
Abstract:	Natriuretic peptides mediate their physiologic effects through activation of membrane-bound, guanylyl cyclase-coupled receptors (NPRs). Receptor dimerization is an important feature of signal transduction. This study was aimed at characterizing structurally important residues of the extracellular ligand-binding domain of NPR-B for receptor dimerization and cGMP generation. Deletion mutagenesis was used to replace cysteine residues at positions 53 (C53S), 417 (C417S), and 426 (C426S) by serine. Receptor expression, dimerization, whole-cell cGMP response, and guanylyl cyclase activity of membrane fractions were determined in stably transfected COS-7 cells. C53S, C417S, and C426S mutants were expressed and found to form disulfide-bridged covalent dimers. In contrast to NPR-B and C53S, C417S and C426S mutants displayed constitutive activity in whole cells (C417S, 146±12%, P<0.01; C426S, 153±7% of ligand-independent NPR-B cGMP generation, P<0.01). The cGMP response of C417S and C426S mutants in whole cells was dose dependent and ≈4 times lower than that in NPR-B, whereas it was blunted in C53S-transfected cells (1 μmol/L CNP, NPR-B 2868±436%; C53S, 206±16% of control, P<0.001 vs NPR-B, C417S, and C426S). Guanylyl cyclase assay in transfected cells confirmed the constitutive activity of C417S and C426S mutants. These data suggest that receptor dimerization by covalent disulfide bridges alters ligand-independent as well as ligand-dependent receptor activity. Localization of the crosslink in relation to the cell membrane is important for configuration of the extracellular domain and the consecutive signal transduction.
Keywords:	Cyclic GMP, Natriuretic Peptides, Natriuretic Peptides Receptors, Signal Transduction, Animals, Rats
Source:	Hypertension
ISSN:	0194-911X
Publisher:	American Heart Association
Volume:	43
Number:	2
Page Range:	460-465
Date:	February 2004
Official Publication:	https://doi.org/10.1161/01.HYP.0000110907.33263.0b
PubMed:	View item in PubMed

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