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MLK3 mediates impact of PKG1α on cardiac function and controls blood pressure through separate mechanisms.

ABSTRACT: cGMP-dependent protein kinase 1α (PKG1α) promotes left ventricle (LV) compensation after pressure overload. PKG1-activating drugs improve heart failure (HF) outcomes but are limited by vasodilation-induced hypotension. Signaling molecules that mediate PKG1α cardiac therapeutic effects but do not promote PKG1α-induced hypotension could therefore represent improved therapeutic targets. We investigated roles of mixed lineage kinase 3 (MLK3) in mediating PKG1α effects on LV function after pressure overload and in regulating BP. In a transaortic constriction HF model, PKG activation with sildenafil preserved LV function in MLK3+/+ but not MLK3-/- littermates. MLK3 coimmunoprecipitated with PKG1α. MLK3-PKG1α cointeraction decreased in failing LVs. PKG1α phosphorylated MLK3 on Thr277/Ser281 sites required for kinase activation. MLK3-/- mice displayed hypertension and increased arterial stiffness, though PKG stimulation with sildenafil or the soluble guanylate cyclase (sGC) stimulator BAY41-2272 still reduced BP in MLK3-/- mice. MLK3 kinase inhibition with URMC-099 did not affect BP but induced LV dysfunction in mice. These data reveal MLK3 as a PKG1α substrate mediating PKG1α preservation of LV function but not acute PKG1α BP effects. Mechanistically, MLK3 kinase-dependent effects preserved LV function, whereas MLK3 kinase-independent signaling regulated BP. These findings suggest augmenting MLK3 kinase activity could preserve LV function in HF but avoid hypotension from PKG1α activation.

SUBMITTER: Calamaras TD

PROVIDER: S-EPMC8492323 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

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MLK3 mediates impact of PKG1α on cardiac function and controls blood pressure through separate mechanisms.

Calamaras Timothy D TD Pande Suchita S Baumgartner Robert Au RA Kim Seung Kyum SK McCarthy Joseph C JC Martin Gregory L GL Tam Kelly K McLaughlin Angela L AL Wang Guang-Rong GR Aronovitz Mark J MJ Lin Weiyu W Aguirre Jonathan I JI Baca Paulina P Liu Peiwen P Richards Daniel A DA Davis Roger J RJ Karas Richard H RH Jaffe Iris Z IZ Blanton Robert M RM

JCI insight 20210922 18

cGMP-dependent protein kinase 1α (PKG1α) promotes left ventricle (LV) compensation after pressure overload. PKG1-activating drugs improve heart failure (HF) outcomes but are limited by vasodilation-induced hypotension. Signaling molecules that mediate PKG1α cardiac therapeutic effects but do not promote PKG1α-induced hypotension could therefore represent improved therapeutic targets. We investigated roles of mixed lineage kinase 3 (MLK3) in mediating PKG1α effects on LV function after pressure o ...[more]

PMID: 34324442

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Project description:RationaleStimulated PKG1α (protein kinase G-1α) phosphorylates TSC2 (tuberous sclerosis complex 2) at serine 1365, potently suppressing mTORC1 (mechanistic [mammalian] target of rapamycin complex 1) activation by neurohormonal and hemodynamic stress. This reduces pathological hypertrophy and dysfunction and increases autophagy. PKG1α oxidation at cysteine-42 is also induced by these stressors, which blunts its cardioprotective effects.ObjectiveWe tested the dependence of mTORC1 activation on PKG1α C42 oxidation and its capacity to suppress such activation by soluble GC-1 (guanylyl cyclase 1) activation.Methods and resultsCardiomyocytes expressing wild-type (WT) PKG1α (PKG1αWT) or cysteine-42 to serine mutation redox-dead (PKG1αCS/CS) were exposed to ET-1 (endothelin 1). Cells expressing PKG1αWT exhibited substantial mTORC1 activation (p70 S6K [p70 S6 kinase], 4EBP1 [elF4E binding protein-1], and Ulk1 [Unc-51-like kinase 1] phosphorylation), reduced autophagy/autophagic flux, and abnormal protein aggregation; all were markedly reversed by PKG1αCS/CS expression. Mice with global knock-in of PKG1αCS/CS subjected to pressure overload (PO) also displayed markedly reduced mTORC1 activation, protein aggregation, hypertrophy, and ventricular dysfunction versus PO in PKG1αWT mice. Cardioprotection against PO was equalized between groups by co-treatment with the mTORC1 inhibitor everolimus. TSC2-S1365 phosphorylation increased in PKG1αCS/CS more than PKG1αWT myocardium following PO. TSC2S1365A/S1365A (TSC2 S1365 phospho-null, created by a serine to alanine mutation) knock-in mice lack TSC2 phosphorylation by PKG1α, and when genetically crossed with PKG1αCS/CS mice, protection against PO-induced mTORC1 activation, cardiodepression, and mortality in PKG1αCS/CS mice was lost. Direct stimulation of GC-1 (BAY-602770) offset disparate mTORC1 activation between PKG1αWT and PKG1αCS/CS after PO and blocked ET-1 stimulated mTORC1 in TSC2S1365A-expressing myocytes.ConclusionsOxidation of PKG1α at C42 reduces its phosphorylation of TSC2, resulting in amplified PO-stimulated mTORC1 activity and associated hypertrophy, dysfunction, and depressed autophagy. This is ameliorated by direct GC-1 stimulation.

Project description:BackgroundAlthough nitroglycerin has remained in clinical use since 1879, the mechanism by which it relaxes blood vessels to lower blood pressure remains incompletely understood. Nitroglycerin undergoes metabolism that generates several reaction products, including oxidants, and this bioactivation process is essential for vasodilation. Protein kinase G (PKG) mediates classic nitric oxide-dependent vasorelaxation, but the 1α isoform is also independently activated by oxidation that involves interprotein disulfide formation within this homodimeric protein complex. We hypothesized that nitroglycerin-induced vasodilation is mediated by disulfide activation of PKG1α.Methods and resultsTreating smooth muscle cells or isolated blood vessels with nitroglycerin caused PKG1α disulfide dimerization. PKG1α disulfide formation was increased in wild-type mouse aortas by in vivo nitroglycerin treatment, but this oxidation was lost as tolerance developed. To establish whether kinase oxidation underlies nitroglycerin-induced vasodilation in vivo, we used a Cys42Ser PKG1α knock-in mouse that cannot transduce oxidant signals because it does not contain the vital redox-sensing thiol. This redox-dead knock-in mouse was substantively deficient in hypotensive response to nitroglycerin compared with wild-type littermates as measured in vivo by radiotelemetry. Resistance blood vessels from knock-ins were markedly less sensitive to nitroglycerin-induced vasodilation (EC(50)=39.2 ± 10.7 μmol/L) than wild-types (EC(50)=12.1 ± 2.9 μmol/L). Furthermore, after ≈24 hours of treatment, wild-type controls stopped vasodilating to nitroglycerin, and the vascular sensitivity to nitroglycerin was decreased, whereas this tolerance phenomenon, which routinely hampers the management of hypertensive patients, was absent in knock-ins.ConclusionsPKG1α disulfide formation is a significant mediator of nitroglycerin-induced vasodilation, and tolerance to nitroglycerin is associated with loss of kinase oxidation.

Project description:BackgroundIt is unclear how cardiac autonomic function, as indicated by heart rate (HR), heart rate variability (HRV), HR increase during exercise, and HR recovery after exercise, is related to blood pressure (BP). We aimed to examine the observational and genetic evidence for a potential causal effect of these HR(V) traits on BP.MethodsWe performed multivariable adjusted linear regression using Lifelines and UK Biobank cohorts to investigate the relationship between HR(V) traits and BP. Linkage disequilibrium score regression was conducted to examine genetic correlations. We used two-sample Mendelian randomization (2SMR) to examine potential causal relations between HR(V) traits and BP.ResultsObservational analyses showed negative associations of all HR(V) traits with BP, except for HR, which was positively associated. Genetic correlations were directionally consistent with the observational associations, but most significant genetic correlations between HR(V) traits and BP were limited to diastolic blood pressure (DBP). 2SMR analyses suggested a potentially causal relationship between HR(V) traits and DBP but not systolic blood pressure (SBP). No reverse effect of BP on HR(V) traits was found. One standard deviation (SD) unit increase in HR was associated with a 1.82 mmHg elevation of DBP. In contrast, one ln(ms) unit increase of the root mean square of the successive differences (RMSSD) and corrected RMSSD (RMSSDc), decreased DBP by 1.79 and 1.83 mmHg, respectively. For HR increase and HR recovery at 50 s, every additional SD increase was associated with a lower DBP by 2.05 and 1.47 mmHg, respectively. Results of secondary analyses with pulse pressure as outcome were inconsistent between observational and 2SMR analyses, as well as between HR(V) traits, and therefore inconclusive.ConclusionBoth observational and genetic evidence show strong associations between indices of cardiac autonomic function and DBP, suggesting that a larger relative contribution of the sympathetic versus the parasympathetic nervous system to cardiac function may cause elevated DBP.

Dataset Information

MLK3 mediates impact of PKG1α on cardiac function and controls blood pressure through separate mechanisms.

Publications

MLK3 mediates impact of PKG1α on cardiac function and controls blood pressure through separate mechanisms.

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