Tyrosine phosphorylation of eNOS regulates myocardial survival after an ischaemic insult

Role of PYK2

Sofia Iris Bibli, Zongmin Zhou, Sven Zukunft, Beate Fisslthaler, Ioanna Andreadou, Csaba Szabo, Peter Brouckaert, Ingrid Fleming, Andreas Papapetropoulos

    Research output: Contribution to journalArticle

    10 Citations (Scopus)

    Abstract

    Aims Endothelial nitric oxide (NO) synthase (eNOS) is known to play a cardioprotective protective.However, the molecular mechanisms regulating eNOS activity during ischaemia/reperfusion (I/R) injury are incompletely understood.eNOS is a substrate for several kinases that positively or negatively affect its enzymatic activity.Herein, we sought to correlate eNOS phosphorylation status with cardiomyocyte survival and we investigated the contribution of the proline-rich tyrosine kinase 2 (PYK2)/eNOS axis to the regulation of myocardial infarct size in vivo.Methods and results Exposure of H9c2 cardiomyocytes to H2O2 lead to PYK2 phosphorylation on its activator site (Y402) and eNOS phosphorylation on the inhibitor site Y656 and the activator site S1176.Both H2O2induced eNOS phosphorylation events were abolished by PYK2 pharmacological inhibition or gene knockdown.Activity assays demonstrated that phosphorylation of the tyrosine inhibitory site exerts a dominant effect over S1176.In cardiomyocytes subjected to oxidative stress or oxygen-glucose deprivation, inhibition of PYK2 limited cell injury; this effect was prevented by inhibition of NO production.In vivo, ischaemia-reperfusion induced an early activation of PYK2, leading to eNOS phosphorylation on Y656, which, in turn, reduced NO output, as judged by the low tissue levels of its downstream effector cGMP.Moreover, pharmacological blockade of PYK2 alleviated eNOS inhibition and prevented cardiac damage following I/R injury in wild-type, but not in eNOS KO mice.Conclusion The current studies demonstrate that PYK2 is a pivotal regulator of eNOS function in myocardial infarction and identify PYK2 as a novel therapeutic target for cardioprotection.All rights reserved.

    Original languageEnglish (US)
    Pages (from-to)926-937
    Number of pages12
    JournalCardiovascular Research
    Volume113
    Issue number8
    DOIs
    StatePublished - Jul 1 2017

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    Focal Adhesion Kinase 2
    Tyrosine
    Phosphorylation
    Cardiac Myocytes
    Reperfusion Injury
    Nitric Oxide
    Myocardial Infarction
    Pharmacology
    Gene Knockdown Techniques
    Nitric Oxide Synthase Type III
    Reperfusion
    Oxidative Stress
    Phosphotransferases
    Ischemia
    Oxygen
    Glucose

    Keywords

    • Cardioprotection
    • Cyclic nucleotide
    • eNOS
    • Myocardial infarction
    • Reperfusion injury

    ASJC Scopus subject areas

    • Physiology
    • Cardiology and Cardiovascular Medicine
    • Physiology (medical)

    Cite this

    Tyrosine phosphorylation of eNOS regulates myocardial survival after an ischaemic insult : Role of PYK2. / Bibli, Sofia Iris; Zhou, Zongmin; Zukunft, Sven; Fisslthaler, Beate; Andreadou, Ioanna; Szabo, Csaba; Brouckaert, Peter; Fleming, Ingrid; Papapetropoulos, Andreas.

    In: Cardiovascular Research, Vol. 113, No. 8, 01.07.2017, p. 926-937.

    Research output: Contribution to journalArticle

    Bibli, SI, Zhou, Z, Zukunft, S, Fisslthaler, B, Andreadou, I, Szabo, C, Brouckaert, P, Fleming, I & Papapetropoulos, A 2017, 'Tyrosine phosphorylation of eNOS regulates myocardial survival after an ischaemic insult: Role of PYK2', Cardiovascular Research, vol. 113, no. 8, pp. 926-937. https://doi.org/10.1093/cvr/cvx058
    Bibli, Sofia Iris ; Zhou, Zongmin ; Zukunft, Sven ; Fisslthaler, Beate ; Andreadou, Ioanna ; Szabo, Csaba ; Brouckaert, Peter ; Fleming, Ingrid ; Papapetropoulos, Andreas. / Tyrosine phosphorylation of eNOS regulates myocardial survival after an ischaemic insult : Role of PYK2. In: Cardiovascular Research. 2017 ; Vol. 113, No. 8. pp. 926-937.
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    abstract = "Aims Endothelial nitric oxide (NO) synthase (eNOS) is known to play a cardioprotective protective.However, the molecular mechanisms regulating eNOS activity during ischaemia/reperfusion (I/R) injury are incompletely understood.eNOS is a substrate for several kinases that positively or negatively affect its enzymatic activity.Herein, we sought to correlate eNOS phosphorylation status with cardiomyocyte survival and we investigated the contribution of the proline-rich tyrosine kinase 2 (PYK2)/eNOS axis to the regulation of myocardial infarct size in vivo.Methods and results Exposure of H9c2 cardiomyocytes to H2O2 lead to PYK2 phosphorylation on its activator site (Y402) and eNOS phosphorylation on the inhibitor site Y656 and the activator site S1176.Both H2O2induced eNOS phosphorylation events were abolished by PYK2 pharmacological inhibition or gene knockdown.Activity assays demonstrated that phosphorylation of the tyrosine inhibitory site exerts a dominant effect over S1176.In cardiomyocytes subjected to oxidative stress or oxygen-glucose deprivation, inhibition of PYK2 limited cell injury; this effect was prevented by inhibition of NO production.In vivo, ischaemia-reperfusion induced an early activation of PYK2, leading to eNOS phosphorylation on Y656, which, in turn, reduced NO output, as judged by the low tissue levels of its downstream effector cGMP.Moreover, pharmacological blockade of PYK2 alleviated eNOS inhibition and prevented cardiac damage following I/R injury in wild-type, but not in eNOS KO mice.Conclusion The current studies demonstrate that PYK2 is a pivotal regulator of eNOS function in myocardial infarction and identify PYK2 as a novel therapeutic target for cardioprotection.All rights reserved.",
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    T1 - Tyrosine phosphorylation of eNOS regulates myocardial survival after an ischaemic insult

    T2 - Role of PYK2

    AU - Bibli, Sofia Iris

    AU - Zhou, Zongmin

    AU - Zukunft, Sven

    AU - Fisslthaler, Beate

    AU - Andreadou, Ioanna

    AU - Szabo, Csaba

    AU - Brouckaert, Peter

    AU - Fleming, Ingrid

    AU - Papapetropoulos, Andreas

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    N2 - Aims Endothelial nitric oxide (NO) synthase (eNOS) is known to play a cardioprotective protective.However, the molecular mechanisms regulating eNOS activity during ischaemia/reperfusion (I/R) injury are incompletely understood.eNOS is a substrate for several kinases that positively or negatively affect its enzymatic activity.Herein, we sought to correlate eNOS phosphorylation status with cardiomyocyte survival and we investigated the contribution of the proline-rich tyrosine kinase 2 (PYK2)/eNOS axis to the regulation of myocardial infarct size in vivo.Methods and results Exposure of H9c2 cardiomyocytes to H2O2 lead to PYK2 phosphorylation on its activator site (Y402) and eNOS phosphorylation on the inhibitor site Y656 and the activator site S1176.Both H2O2induced eNOS phosphorylation events were abolished by PYK2 pharmacological inhibition or gene knockdown.Activity assays demonstrated that phosphorylation of the tyrosine inhibitory site exerts a dominant effect over S1176.In cardiomyocytes subjected to oxidative stress or oxygen-glucose deprivation, inhibition of PYK2 limited cell injury; this effect was prevented by inhibition of NO production.In vivo, ischaemia-reperfusion induced an early activation of PYK2, leading to eNOS phosphorylation on Y656, which, in turn, reduced NO output, as judged by the low tissue levels of its downstream effector cGMP.Moreover, pharmacological blockade of PYK2 alleviated eNOS inhibition and prevented cardiac damage following I/R injury in wild-type, but not in eNOS KO mice.Conclusion The current studies demonstrate that PYK2 is a pivotal regulator of eNOS function in myocardial infarction and identify PYK2 as a novel therapeutic target for cardioprotection.All rights reserved.

    AB - Aims Endothelial nitric oxide (NO) synthase (eNOS) is known to play a cardioprotective protective.However, the molecular mechanisms regulating eNOS activity during ischaemia/reperfusion (I/R) injury are incompletely understood.eNOS is a substrate for several kinases that positively or negatively affect its enzymatic activity.Herein, we sought to correlate eNOS phosphorylation status with cardiomyocyte survival and we investigated the contribution of the proline-rich tyrosine kinase 2 (PYK2)/eNOS axis to the regulation of myocardial infarct size in vivo.Methods and results Exposure of H9c2 cardiomyocytes to H2O2 lead to PYK2 phosphorylation on its activator site (Y402) and eNOS phosphorylation on the inhibitor site Y656 and the activator site S1176.Both H2O2induced eNOS phosphorylation events were abolished by PYK2 pharmacological inhibition or gene knockdown.Activity assays demonstrated that phosphorylation of the tyrosine inhibitory site exerts a dominant effect over S1176.In cardiomyocytes subjected to oxidative stress or oxygen-glucose deprivation, inhibition of PYK2 limited cell injury; this effect was prevented by inhibition of NO production.In vivo, ischaemia-reperfusion induced an early activation of PYK2, leading to eNOS phosphorylation on Y656, which, in turn, reduced NO output, as judged by the low tissue levels of its downstream effector cGMP.Moreover, pharmacological blockade of PYK2 alleviated eNOS inhibition and prevented cardiac damage following I/R injury in wild-type, but not in eNOS KO mice.Conclusion The current studies demonstrate that PYK2 is a pivotal regulator of eNOS function in myocardial infarction and identify PYK2 as a novel therapeutic target for cardioprotection.All rights reserved.

    KW - Cardioprotection

    KW - Cyclic nucleotide

    KW - eNOS

    KW - Myocardial infarction

    KW - Reperfusion injury

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