Disulfide-activated protein kinase G Iα regulates cardiac diastolic relaxation and fine-tunes the Frank–Starling response

Scotcher, Jenna; Prysyazhna, Oleksandra; Boguslavskyi, Andrii; Kistamás, Kornél; Hadgraft, Natasha; Martin, Eva Denise; Worthington, Jenny; Rudyk, Olena; Cutillas, Pedro R.; Cuello, Friederike; Shattock, Michael J.; Marber, Michael; Conte, Maria R.; Greenstein, Adam; Greensmith, David J.; Venetucci, Luigi; Timms, John F.; Eaton, Philip

doi:10.1038/ncomms13187

Cited by 38 publications

(28 citation statements)

References 47 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although it failed to reach significance, this is in line with the previously published data for younger mice [8] , [12] . KI mice fed a normal diet had smaller left ventricular diastolic dimension and volume than the WT mice on a normal diet, consistent with the diastolic dysfunction observed in the KI mice previously [8] , [10] . The obesogenic high fat diet caused both systolic and diastolic dysfunction in the WT mice ( Fig.…”

Section: Resultssupporting

confidence: 81%

“…Protein kinase G (PKG) is the end-effector kinase that mediates nitric oxide (NO)-dependent vasorelaxation [7] , with the Iα isoform mediating oxidant-dependent vasorelaxation to maintain normal healthy blood pressure [8] , [9] and regulate diastolic relaxation [10] . Whilst these finding support a role for oxidant-activated PKG Iα in the maintenance of health, oxidation of this kinase have also been observed in the pathogenesis of cardiovascular disease.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Examining a role for PKG Iα oxidation in the pathogenesis of cardiovascular dysfunction during diet-induced obesity

Rudyk¹,

Eaton²

2017

Free Radical Biology and Medicine

Self Cite

View full text Add to dashboard Cite

BackgroundProtein kinase G (PKG) Iα is the end-effector kinase that mediates nitric oxide (NO)-dependent and oxidant-dependent vasorelaxation to maintain blood pressure during health. A hallmark of cardiovascular disease is attenuated NO production, which in part is caused by NO Synthase (NOS) uncoupling, which in turn increases oxidative stress because of superoxide generation. NOS uncoupling promotes PKG Iα oxidation to the interprotein disulfide state, likely mediated by superoxide-derived hydrogen peroxide, and because the NO-cyclic guanosine monophosphate (cGMP) pathway otherwise negatively regulates oxidation of the kinase to its active disulfide dimeric state. Diet-induced obesity is associated with NOS uncoupling, which may in part contribute to the associated cardiovascular dysfunction due to exacerbated PKG Iα disulfide oxidation to the disulfide state. This is a rational hypothesis because PKG Iα oxidation is known to significantly contribute to heart failure that arises from chronic myocardial oxidative stress.Methods and resultsBovine arterial endothelial cells (BAECs) or smooth muscle cells (SMCs) were exposed to drugs that uncouple NOS. These included 1,3-bis(2-chloroethyl)−1-nitrosourea (BCNU) which promotes its S-glutathiolation, 4-diamino-6-hydroxy-pyrimidine (DAHP) which inhibits guanosine-5′-triphosphate-cyclohydrolase 2 to prevent BH4 synthesis or methotrexate (MTX) which inhibits the regeneration of BH4 from BH2 by dihydrofolate reductase. While all the drugs mentioned above induced robust PKG Iα disulfide dimerization in cells, exposure of BAECs to NOS inhibitor L-NMMA did not. Increased PKG Iα disulfide formation occurred in hearts and aortae from mice treated in vivo with DAHP (10 mM in a drinking water for 3 weeks). Redox-dead C42S PKG Iα knock-in (KI) mice developed less pronounced cardiac posterior wall hypertrophy and did not develop cardiac dysfunction, assessed by echocardiography, compared to the wild-type (WT) mice after chronic DAHP treatment. WT or KI mice were then subjected to a diet-induced obesity protocol by feeding them with a high fat Western-type diet (RM 60% AFE) for 27 weeks, which increased body mass, adiposity, plasma leptin, resistin and glucagon levels comparably in each genotype. Obesity-induced hypertension, assessed by radiotelemetry, was mild and transient in the WT, while the basally hypertensive KI mice were resistant to further increases in blood pressure following high fat feeding. Although the obesogenic diet caused mild cardiac dysfunction in the WT but not the KI mice, gross changes in myocardial structure monitored by echocardiography were not apparent in either genotype. The level of cyclic guanosine monophosphate (cGMP) was decreased in the aortae of WT and KI mice following high fat feeding. PKG Iα oxidation was not evident in the hearts of WT mice fed a high fat diet.ConclusionsDespite robust evidence for PKG Iα oxidation during NOS uncoupling in cell models, it is unlikely that PKG Iα oxidation occurs to a significant extent in vivo during diet-ind...

show abstract

Section: Resultssupporting

confidence: 81%

Section: Introductionmentioning

confidence: 99%

Examining a role for PKG Iα oxidation in the pathogenesis of cardiovascular dysfunction during diet-induced obesity

Rudyk¹,

Eaton²

2017

Free Radical Biology and Medicine

Self Cite

View full text Add to dashboard Cite

show abstract

“…However, oxidation of Cys42 may not induce substantive catalytic activation compared to cGMP-dependent activation of the kinase 25 . Given that Cys42 is localised in the N-terminal leucine zipper domain that is important for substrate targeting 26 , the redox state of this thiol may be more important for modulating such interactions, as suggested previously 27 – 29 . Consistent with this regulatory role, Cys42Ser PKGIα knock-in (KI) mice, which cannot form the interprotein disulfide, are hypertensive 30 , illustrating a physiological role for oxidants in blood pressure regulation.…”

Section: Introductionmentioning

confidence: 77%

Oxidant sensor in the cGMP-binding pocket of PKGIα regulates nitroxyl-mediated kinase activity

Donzelli

Goetz

Schmidt

et al. 2017

Sci Rep

Self Cite

View full text Add to dashboard Cite

Despite the mechanisms for endogenous nitroxyl (HNO) production and action being incompletely understood, pharmacological donors show broad therapeutic promise and are in clinical trials. Mass spectrometry and site-directed mutagenesis showed that chemically distinct HNO donors 1-nitrosocyclohexyl acetate or Angeli’s salt induced disulfides within cGMP-dependent protein kinase I-alpha (PKGIα), an interdisulfide between Cys42 of the two identical subunits of the kinase and a previously unobserved intradisulfide between Cys117 and Cys195 in the high affinity cGMP-binding site. Kinase activity was monitored in cells transfected with wildtype (WT), Cys42Ser or Cys117/195Ser PKGIα that cannot form the inter- or intradisulfide, respectively. HNO enhanced WT kinase activity, an effect significantly attenuated in inter- or intradisulfide-deficient PKGIα. To investigate whether the intradisulfide modulates cGMP binding, real-time imaging was performed in vascular smooth muscle cells expressing a FRET-biosensor comprising the cGMP-binding sites of PKGIα. HNO induced FRET changes similar to those elicited by an increase of cGMP, suggesting that intradisulfide formation is associated with activation of PKGIα. Intradisulfide formation in PKGIα correlated with enhanced HNO-mediated vasorelaxation in mesenteric arteries in vitro and arteriolar dilation in vivo in mice. HNO induces intradisulfide formation in PKGIα, inducing the same effect as cGMP binding, namely kinase activation and thus vasorelaxation.

show abstract

“…Ion channels regulate cardiac excitability and contractility with millisecond timescales, raising the possibility that thiol-reactive NO species modulate heart function via protein S-nitrosation on a beat-to beat basis [74]. Recently we reported that disulfide formation in myocardial protein kinase G I rapidly regulates diastolic relaxation [75], consistent with this oxidative modification also being capable of regulation with a millisecond timescale. Whilst both these are possibilities are interesting, the half-life of these modifications has yet to be determined.…”

Section: Further Considerations Of the Stability Of Protein S-nitrosomentioning

confidence: 89%

How widespread is stable protein S-nitrosylation as an end-effector of protein regulation?

Wolhuter

Eaton

2017

Free Radical Biology and Medicine

Self Cite

View full text Add to dashboard Cite

Over the last 25 years protein S-nitrosylation, also known more correctly as S-nitrosation, has been progressively implicated in virtually every nitric oxide-regulated process within the cardiovascular system. The current, widely-held paradigm is that S-nitrosylation plays an equivalent role as phosphorylation, providing a stable and controllable post-translational modification that directly regulates end-effector target proteins to elicit biological responses. However, this concept largely ignores the intrinsic instability of the nitrosothiol bond, which rapidly reacts with typically abundant thiol-containing molecules to generate more stable disulfide bonds. These protein disulfides, formed via a nitrosothiol intermediate redox state, are rationally anticipated to be the predominant endeffector modification that mediates functional alterations when cells encounter nitrosative stimuli. In this review we present evidence and explain our reasoning for arriving at this conclusion that may be controversial to some researchers in the field.

show abstract

Disulfide-activated protein kinase G Iα regulates cardiac diastolic relaxation and fine-tunes the Frank–Starling response

Cited by 38 publications

References 47 publications

Examining a role for PKG Iα oxidation in the pathogenesis of cardiovascular dysfunction during diet-induced obesity

Examining a role for PKG Iα oxidation in the pathogenesis of cardiovascular dysfunction during diet-induced obesity

Oxidant sensor in the cGMP-binding pocket of PKGIα regulates nitroxyl-mediated kinase activity

How widespread is stable protein S-nitrosylation as an end-effector of protein regulation?

Contact Info

Product

Resources

About