Dominant Negative Mutations of the Guanylyl Cyclase-A Receptor

Thompson, Dana K.; Garbers, David L.

doi:10.1074/jbc.270.1.425

Cited by 72 publications

(34 citation statements)

References 31 publications

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“…These findings mirror data published by Chinkers and Wilson (21) demonstrating a lack of interaction between the extracellular domains of NPR-A and NPR-B. Furthermore, a dominant-negative mutant of NPR-A as well as an alternative splice variant of NPR-B acting in a dominant-negative manner have previously been used to study ANP and CNP physiology, providing evidence for the use of dominant-negative mutants in cell culture and transgenic animal models as a powerful method to study the specific function of NPRs (19,(22)(23)(24).…”

Section: Discussionmentioning

confidence: 99%

Cardiac hypertrophy in transgenic rats expressing a dominant-negative mutant of the natriuretic peptide receptor B

Langenickel

Buttgereit

Pagel-Langenickel

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

115

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Natriuretic peptides (NP) mediate their effects by activating membrane-bound guanylyl cyclase-coupled receptors A (NPR-A) or B (NPR-B). Whereas the pathophysiological role of NPR-A has been widely studied, only limited knowledge on the cardiovascular function of NPR-B is available. In vitro studies suggest antiproliferative and antihypertrophic actions of the NPR-B ligand C-type NP (CNP). Because of the lack of a specific pharmacological inhibitor, these effects could not clearly be attributed to impaired NPR-B signaling. Recently, gene deletion revealed a predominant role of NPR-B in endochondral ossification and development of female reproductive organs. However, morphological abnormalities and premature death of NPR-B-deficient mice preclude detailed cardiovascular phenotyping. In the present study, a dominant-negative mutant (NPR-B⌬KC) was used to characterize CNP-dependent NPR-B signaling in vitro and in transgenic rats. Here we demonstrate that reduced CNP-but not atrial NP-dependent cGMP response attenuates antihypertrophic potency of CNP in vitro. In transgenic rats, NPR-B⌬KC expression selectively reduced NPR-B but not NPR-A signaling. NPR-B⌬KC transgenic rats display progressive, blood pressure-independent cardiac hypertrophy and elevated heart rate. The hypertrophic phenotype is further enhanced in chronic volume overload-induced congestive heart failure. Thus, this study provides evidence linking NPR-B signaling to the control of cardiac growth.C-type natriuretic peptide ͉ knockdown

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Section: Discussionmentioning

confidence: 99%

Cardiac hypertrophy in transgenic rats expressing a dominant-negative mutant of the natriuretic peptide receptor B

Langenickel

Buttgereit

Pagel-Langenickel

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

115

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“…This generated a cDNA fragment coding for a cytoplasmic domain of NPRA containing the catalytic fragment of the guanylate cyclase domain. It has been shown that this fragment codes for a soluble cytoplasmic protein that has constitutive guanylate cyclase activity in transfected COS cells (16,17). This cDNA fragment was cloned downstream of a 5.8-kb fragment of the ␣-myosin heavy chain gene promoter (generous gift from J. Robbins, University of Cincinnati) and upstream of a portion of the rabbit ␤-globin gene containing an intron and a polyadenylation signal.…”

Section: Methodsmentioning

confidence: 99%

Expression of Constitutively Active Guanylate Cyclase in Cardiomyocytes Inhibits the Hypertrophic Effects of Isoproterenol and Aortic Constriction on Mouse Hearts

Zahabi

Picard

Fortin

et al. 2003

Journal of Biological Chemistry

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Evidence from several rodent models has suggested that a reduction of either atrial natriuretic peptide or its receptor in the heart affects cardiac remodeling by promoting the onset of cardiac hypertrophy. The atrial natriuretic peptide receptor mediates signaling at least in part via the generation of intracellular cyclic GMP. To directly test whether accumulation of intracellular cyclic GMP conveys protection against cardiac hypertrophy, we engineered transgenic mice that overexpress a catalytic fragment of constitutively active guanylate cyclase domain of the atrial natriuretic peptide receptor in a cardiomyocyte-specific manner. Expression of the transgene increased the intracellular concentration of cyclic GMP specifically within cardiomyocytes and had no detectable effect on cardiac performance under basal conditions. However, expression of the transgene attenuated the effects of the pharmacologic hypertrophic agent isoproterenol on cardiac wall thickness and prevented the onset of the fetal gene expression program normally associated with cardiac hypertrophy. Likewise, expression of the transgene inhibited the hypertrophic effects of abdominal aortic constriction, since it abolished its effects on ventricular wall thickness and greatly attenuated its effects on cardiomyocyte size. Altogether, our results suggest that cyclic GMP is a cardioprotective agent against hypertrophy that acts via a direct local effect on cardiomyocytes. Left ventricular hypertrophy (LVH)1 results from the activation of multiple signaling pathways by either mechanical or neurohumoral stimuli (1, 2). A great number of studies have used animal models of transgenesis or gene inactivation to test the possible roles of these pathways in the induction of LVH in vivo. However, the contributions of possible negative regulators of LVH have so far received much less attention. Recently, several lines of evidence have suggested that atrial natriuretic peptide (ANP) and endothelial nitric-oxide synthase may act as such negative regulators, since 1) blood-pressure-independent LVH is present in mice with either general (3) or cardiomyocyte-restricted (4) inactivation of natriuretic peptide receptor A (NPRA); 2) cardiomyocyte-specific expression of NPRA partially rescues the cardiac hypertrophic phenotype seen in NPRA-null mice (5); 3) we have shown that cardiac mass and ventricular expression of ANP were both associated (in an inverse fashion) with a naturally occurring allele of natriuretic peptide precursor A (i.e. the gene that codes for ANP) (6 -8); and 4) overexpression of an endothelial nitric-oxide synthase transgene attenuates isoproterenol-induced LVH (9). The common denominator between ANP and endothelial nitric-oxide synthase is that many (if not most) of their biologic effects are mediated by cGMP (10, 11). Interestingly, it has also been shown that cGMP protects cultured neonatal cardiomyocytes against the effects of hypertrophic agents in vitro (12-14). However, it remains to be proven that cGMP has similar actions in vivo.To add...

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“…Other studies using gel filtration instead of co-immunoprecipitation concluded that the intracellular portion is also involved in receptor dimerization (18). Furthermore, it was shown that this interaction is mediated by a hinge region located between the KHD and GC regions (18,19).…”

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confidence: 99%

A Disulfide-bridged Mutant of Natriuretic Peptide Receptor-A Displays Constitutive Activity

Labrecque¹,

Nicoll

Marquis³

et al. 1999

Journal of Biological Chemistry

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Natriuretic peptide receptor-A (NPR-A), a particulate guanylyl cyclase receptor, is composed of an extracellular domain (ECD) with a ligand binding site, a transmembrane spanning, a kinase homology domain (KHD), and a guanylyl cyclase domain. Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), the natural agonists, bind and activate the receptor leading to cyclic GMP production. This receptor has been reported to be spontaneously dimeric or oligomeric. In response to agonists, the KHD-mediated guanylate cyclase repression is removed, and it is assumed that ATP binds to the KHD. Since NPR-A displays a pair of juxtamembrane cysteines separated by 8 residues, we hypothesized that the removal of one of those cysteines would leave the other unpaired and reactive, thus susceptible to form an interchain disulfide bridge and to favor the dimeric interactions. Here we show that NPR-A C423S mutant, expressed mainly as a covalent dimer, increases the affinity of pBNP for this receptor by enhancing a high affinity binding component. Dimerization primarily depends on ECD since a secreted NPR-A C423S soluble ectodomain (ECD C423S ) also documents a covalent dimer. ANP binding to the unmutated ECD yields up to 80-fold affinity loss as compared with the membrane receptor. However, the ECD C423S mutation restores a high binding affinity. Furthermore, C423S mutation leads to cellular constitutive activation (20 -40-fold) of basal catalytic production of cyclic GMP by the fulllength mutant. In vitro particulate guanylyl cyclase assays demonstrate that NPR-A C423S displays an increased sensitivity to ATP treatment alone and that the effect of ANP ؉ ATP joint treatment is cumulative instead of synergistic. Finally, the cellular and particulate guanylyl cyclase assays indicate that the receptor is desensitized to agonist stimulation. We conclude the following: 1) dimers are functional units of NPR-A guanylyl cyclase activation; and 2) agonists are inducing dimeric contact of the juxtamembranous region leading to the removal of the KHD-mediated guanylyl cyclase repression, hence allowing catalytic activation.

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Dominant Negative Mutations of the Guanylyl Cyclase-A Receptor

Cited by 72 publications

References 31 publications

Cardiac hypertrophy in transgenic rats expressing a dominant-negative mutant of the natriuretic peptide receptor B

Cardiac hypertrophy in transgenic rats expressing a dominant-negative mutant of the natriuretic peptide receptor B

Expression of Constitutively Active Guanylate Cyclase in Cardiomyocytes Inhibits the Hypertrophic Effects of Isoproterenol and Aortic Constriction on Mouse Hearts

A Disulfide-bridged Mutant of Natriuretic Peptide Receptor-A Displays Constitutive Activity

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