Prevention of PKG1α oxidation augments cardioprotection in the stressed heart

Nakamura, Taishi; Ranek, Mark J.; Lee, Dong I.; Hahn, Virginia S.; Kim, Choel; Eaton, Philip; Kass, David A.

doi:10.1172/jci80275

Cited by 66 publications

(78 citation statements)

References 27 publications

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“…Consequently, it is possible that scenario-specific localization of PKG I␣ occurs depending on the subcellular source of cGMP or oxidant, as well as the amounts of each generated. Indeed, it is notable that the ability of PKG I␣ to form a disulfide dimer impacts on its subcellular localization in cardiac myocytes in response to exogenous hydrogen peroxide or those from transaortic constriction (TAC)-stressed hearts (11).…”

Section: Discussionmentioning

confidence: 99%

“…Thus, the zipper in PKG I␣ is integral to the protection that elevation in cGMP by PDE5 inhibitors can provide, and this is because it enables the kinase to bind and phospho-activate RhoA, which ultimately limits apoptosis. The C42S PKG I␣ KI mice are also innately resistant to TAC-induced cardiac dysfunction (11). The disulfide in PKG I␣ occurs between Cys-42 residues directly within the leucine zipper domain required for targeting of the kinase to RhoA.…”

Section: Discussionmentioning

confidence: 99%

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Phosphodiesterase 5 Inhibition Limits Doxorubicin-induced Heart Failure by Attenuating Protein Kinase G Iα Oxidation

Prysyazhna¹,

Burgoyne²,

Scotcher³

et al. 2016

Journal of Biological Chemistry

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Phosphodiesterase 5 (PDE5) inhibitors limit myocardial injury caused by stresses, including doxorubicin chemotherapy. cGMP binding to PKG I␣ attenuates oxidant-induced disulfide formation. Because PDE5 inhibition elevates cGMP and protects from doxorubicin-induced injury, we reasoned that this may be because it limits PKG I␣ disulfide formation. To investigate the role of PKG I␣ disulfide dimerization in the development of apoptosis, doxorubicin-induced cardiomyopathy was compared in male wild type (WT) or disulfide-resistant C42S PKG I␣ knock-in (KI) mice. Echocardiography showed that doxorubicin treatment caused loss of myocardial tissue and depressed left ventricular function in WT mice. Doxorubicin also reduced pro-survival signaling and increased apoptosis in WT hearts. In contrast, KI mice were markedly resistant to the dysfunction induced by doxorubicin in WTs. In follow-on experiments the influence of the PDE5 inhibitor tadalafil on the development of doxorubicin-induced cardiomyopathy in WT and KI mice was investigated. In WT mice, co-administration of tadalafil with doxorubicin reduced PKG I␣ oxidation caused by doxorubicin and also protected against cardiac injury and loss of function. KI mice were again innately resistant to doxorubicin-induced cardiotoxicity, and therefore tadalafil afforded no additional protection. Doxorubicin decreased phosphorylation of RhoA (Ser-188), stimulating its GTPase activity to activate Rho-associated protein kinase (ROCK) in WTs. These pro-apoptotic events were absent in KI mice and were attenuated in WTs coadministered tadalafil. PKG I␣ disulfide formation triggers cardiac injury, and this initiation of maladaptive signaling can be blocked by pharmacological therapies that elevate cGMP, which binds kinase to limit its oxidation.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Phosphodiesterase 5 Inhibition Limits Doxorubicin-induced Heart Failure by Attenuating Protein Kinase G Iα Oxidation

Prysyazhna¹,

Burgoyne²,

Scotcher³

et al. 2016

Journal of Biological Chemistry

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“…Indeed, oxidation has been shown to increase the in vitro interaction between PKGI␣ and two of its interacting proteins, RhoA and MYPT1 (12). In addition, a recent paper by Nakamura et al (15) demonstrated that the C43S mutation appeared to alter PKGI␣ subcellular localization in cardiac myocytes, which suggests a change in association of with interacting/anchoring proteins in cells, but the structural basis for oxidation-induced changes in these interactions was not examined. We are currently pursuing these studies.…”

Section: Pkgi␣ Is Not Activated By Cysmentioning

confidence: 99%

The activity of cGMP-dependent protein kinase Iα is not directly regulated by oxidation-induced disulfide formation at cysteine 43

Kalyanaraman

Zhuang

Pilz

et al. 2017

Journal of Biological Chemistry

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The type I cGMP-dependent protein kinases (PKGs) are key regulators of smooth muscle tone, cardiac hypertrophy, and other physiological processes. The two isoforms PKGI␣ and PKGI␤ are thought to have unique functions because of their tissue-specific expression, different cGMP affinities, and isoform-specific protein-protein interactions. Recently, a non-canonical pathway of PKGI␣ activation has been proposed, in which PKGI␣ is activated in a cGMP-independent fashion via oxidation of Cys 43 , resulting in disulfide formation within the PKGI␣ N-terminal dimerization domain. A "redox-dead" knock-in mouse containing a C43S mutation exhibits phenotypes consistent with decreased PKGI␣ signaling, but the detailed mechanism of oxidation-induced PKGI␣ activation is unknown. Therefore, we examined oxidation-induced activation of PKGI␣, and in contrast to previous findings, we observed that disulfide formation at Cys 43 does not directly activate PKGI␣ in vitro or in intact cells. In transfected cells, phosphorylation of Ras homolog gene family member A (RhoA) and vasodilator-stimulated phosphoprotein was increased in response to 8-CPT-cGMP treatment, but not when disulfide formation in PKGI␣ was induced by H 2 O 2 . Using purified enzymes, we found that the Cys 43 oxidation had no effect on basal kinase activity or K m and V max values; however, PKGI␣ containing the C43S mutation was less responsive to cGMP-induced activation. This reduction in cGMP affinity may in part explain the PKGI␣ lossof-function phenotype of the C43S knock-in mouse. In conclusion, disulfide formation at Cys 43 does not directly activate PKGI␣, and the C43S-mutant PKGI␣ has a higher K a for cGMP. Our results highlight that mutant enzymes should be carefully biochemically characterized before making in vivo inferences.The type I cGMP-dependent protein kinases play key roles in regulating vascular tone, intestinal motility, memory formation, and nociception in the spinal cord (1). The kinases are activated by cGMP, and cellular cGMP levels are increased by the activity of soluble and particulate guanylate cyclases, which are activated by nitric oxide or small peptides, respectively. Conversely, cellular cGMP levels are decreased by phosphodiesterases. Pharmacological cGMP-elevating agents include nitric-oxide donors (nitroglycerin, nitroprusside), direct guanylate cyclase activators (riociguat), and phosphodiesterase inhibitors (sildenafil, tadalafil). These agents are used clinically to treat cardiac ischemia, systemic and pulmonary hypertension, and erectile dysfunction (2).The type I PKG gene produces splice variants (PKGI␣ and PKGI␤)2 that differ in their first ϳ100 amino acids (3). The kinases have a similar domain structure, which can be roughly divided into N-terminal regulatory and C-terminal catalytic domains. The regulatory domain contains a number of functional subdomains. At the extreme N termini, leucine/isoleucine zippers (LZs) mediate homodimerization and facilitate binding to specific interacting proteins (4 -9). After the LZs, each i...

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“…19,22,23,50 Mouse models of pressure overload-induced hypertrophy/remodeling showed that protein kinase G and GC1 activity is downregulated and that the localization of GCb1 is directly disrupted. 6,51 We observed significant hypertrophy compared with WT in all studied groups ( Figure S8), but no other obvious structural damage was seen (not shown). Conversely, cGMP is a negative regulator of the cardiac hypertrophy-related remodeling process, 52 and it was recently shown that stimulation of GC1 activity protects from pathological remodeling and heart failure after myocardial infarction.…”

Section: Potential Function Of the Cx43-gc1 Associationmentioning

confidence: 80%

“…In humans and animals subjected to a prolonged state of cardiac hypertrophy, the initial compensatory mechanisms and their failure lead to maladaptive structural remodeling associated with decreased Cx43 assembly in GJs at the ID and increased Cx43 membrane lateralization 19, 22, 23, 50. Mouse models of pressure overload–induced hypertrophy/remodeling showed that protein kinase G and GC1 activity is downregulated and that the localization of GCβ1 is directly disrupted 6, 51. We observed significant hypertrophy compared with WT in all studied groups (Figure S8), but no other obvious structural damage was seen (not shown).…”

Section: Discussionmentioning

confidence: 99%

Newly Identified NO‐Sensor Guanylyl Cyclase/Connexin 43 Association Is Involved in Cardiac Electrical Function

Crassous

Shu

Huang

et al. 2017

JAHA

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BackgroundGuanylyl cyclase, a heme‐containing α1β1 heterodimer (GC1), produces cGMP in response to Nitric oxide (NO) stimulation. The NO‐GC1‐cGMP pathway negatively regulates cardiomyocyte contractility and protects against cardiac hypertrophy–related remodeling. We recently reported that the β1 subunit of GC1 is detected at the intercalated disc with connexin 43 (Cx43). Cx43 forms gap junctions (GJs) at the intercalated disc that are responsible for electrical propagation. We sought to determine whether there is a functional association between GC1 and Cx43 and its role in cardiac homeostasis.Methods and Results GC1 and Cx43 immunostaining at the intercalated disc and coimmunoprecipitation from membrane fraction indicate that GC1 and Cx43 are associated. Mice lacking the α subunit of GC1 (GCα1 knockout mice) displayed a significant decrease in GJ function (dye‐spread assay) and Cx43 membrane lateralization. In a cardiac‐hypertrophic model, angiotensin II treatment disrupted the GC1‐Cx43 association and induced significant Cx43 membrane lateralization, which was exacerbated in GCα1 knockout mice. Cx43 lateralization correlated with decreased Cx43‐containing GJs at the intercalated disc, predictors of electrical dysfunction. Accordingly, an ECG revealed that angiotensin II–treated GCα1 knockout mice had impaired ventricular electrical propagation. The phosphorylation level of Cx43 at serine 365, a protein‐kinase A upregulated site involved in trafficking/assembly of GJs, was decreased in these models.Conclusions GC1 modulates ventricular Cx43 location, hence GJ function, and partially protects from electrical dysfunction in an angiotensin II hypertrophy model. Disruption of the NO‐cGMP pathway is associated with cardiac electrical disturbance and abnormal Cx43 phosphorylation. This previously unknown NO/Cx43 signaling could be a protective mechanism against stress‐induced arrhythmia.

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Prevention of PKG1α oxidation augments cardioprotection in the stressed heart

Abstract: B r i e f r e p o r t 2 4 6 8

Cited by 66 publications

References 27 publications

Phosphodiesterase 5 Inhibition Limits Doxorubicin-induced Heart Failure by Attenuating Protein Kinase G Iα Oxidation

Phosphodiesterase 5 Inhibition Limits Doxorubicin-induced Heart Failure by Attenuating Protein Kinase G Iα Oxidation

The activity of cGMP-dependent protein kinase Iα is not directly regulated by oxidation-induced disulfide formation at cysteine 43

Newly Identified NO‐Sensor Guanylyl Cyclase/Connexin 43 Association Is Involved in Cardiac Electrical Function

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