Interplay of myosin phosphatase and protein phosphatase-2A in the regulation of endothelial nitric-oxide synthase phosphorylation and nitric oxide production

Róbert Bátori, Bálint Bécsi, Dénes Nagy, Zoltán Kónya, Csaba Hegedũs, Zsuzsanna Bordán, Alexander Dmitriyevich Verin, Beáta Lontay, Ferenc Erdõdi

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Abstract

The inhibitory phosphorylation of endothelial nitric oxide (NO) synthase (eNOS) at Thr497 (eNOS pThr497) by protein kinase C or RhoA-activated kinase is a major regulatory determinant of eNOS activity. The signalling mechanisms involved in the dephosphorylation of eNOSpThr497 have not yet been clarified. This study identifies myosin phosphatase (MP) holoenzyme consisting of protein phosphatase-1 catalytic subunit (PP1c) and MP target subunit-1 (MYPT1) as an eNOSpThr497 phosphatase. In support of this finding are: (i) eNOS and MYPT1 interacts in various endothelial cells (ECs) and in in vitro binding assays (ii) MYPT1 targets and stimulates PP1c toward eNOSpThr497 substrate (iii) phosphorylation of MYPT1 at Thr696 (MYPT1pThr696) controls the activity of MP on eNOSpThr497. Phosphatase inhibition suppresses both NO production and transendothelial resistance (TER) of ECs. In contrast, epigallocatechin-3-gallate (EGCG) signals ECs via the 67 kDa laminin-receptor (67LR) resulting in protein kinase A dependent activation of protein phosphatase-2A (PP2A). PP2A dephosphorylates MYPT1 pThr696 and thereby stimulates MP activity inducing dephosphorylation of eNOSpThr497 and the 20 kDa myosin II light chains. Thus an interplay of MP and PP2A is involved in the physiological regulation of EC functions implying that an EGCG dependent activation of these phosphatases leads to enhanced NO production and EC barrier improvement.

Original languageEnglish (US)
Article number44698
JournalScientific Reports
Volume7
DOIs
StatePublished - Mar 16 2017

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Myosin-Light-Chain Phosphatase
Protein Phosphatase 2
Nitric Oxide Synthase Type III
Nitric Oxide
Endothelial Cells
Phosphorylation
Phosphoric Monoester Hydrolases
Protein Phosphatase 1
Laminin Receptors
Myosin Type II
Myosin Light Chains
Holoenzymes
Cyclic AMP-Dependent Protein Kinases
Protein Kinase C
Phosphotransferases

ASJC Scopus subject areas

  • General

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Interplay of myosin phosphatase and protein phosphatase-2A in the regulation of endothelial nitric-oxide synthase phosphorylation and nitric oxide production. / Bátori, Róbert; Bécsi, Bálint; Nagy, Dénes; Kónya, Zoltán; Hegedũs, Csaba; Bordán, Zsuzsanna; Verin, Alexander Dmitriyevich; Lontay, Beáta; Erdõdi, Ferenc.

In: Scientific Reports, Vol. 7, 44698, 16.03.2017.

Research output: Contribution to journalArticle

Bátori, Róbert ; Bécsi, Bálint ; Nagy, Dénes ; Kónya, Zoltán ; Hegedũs, Csaba ; Bordán, Zsuzsanna ; Verin, Alexander Dmitriyevich ; Lontay, Beáta ; Erdõdi, Ferenc. / Interplay of myosin phosphatase and protein phosphatase-2A in the regulation of endothelial nitric-oxide synthase phosphorylation and nitric oxide production. In: Scientific Reports. 2017 ; Vol. 7.
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AU - Nagy, Dénes

AU - Kónya, Zoltán

AU - Hegedũs, Csaba

AU - Bordán, Zsuzsanna

AU - Verin, Alexander Dmitriyevich

AU - Lontay, Beáta

AU - Erdõdi, Ferenc

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AB - The inhibitory phosphorylation of endothelial nitric oxide (NO) synthase (eNOS) at Thr497 (eNOS pThr497) by protein kinase C or RhoA-activated kinase is a major regulatory determinant of eNOS activity. The signalling mechanisms involved in the dephosphorylation of eNOSpThr497 have not yet been clarified. This study identifies myosin phosphatase (MP) holoenzyme consisting of protein phosphatase-1 catalytic subunit (PP1c) and MP target subunit-1 (MYPT1) as an eNOSpThr497 phosphatase. In support of this finding are: (i) eNOS and MYPT1 interacts in various endothelial cells (ECs) and in in vitro binding assays (ii) MYPT1 targets and stimulates PP1c toward eNOSpThr497 substrate (iii) phosphorylation of MYPT1 at Thr696 (MYPT1pThr696) controls the activity of MP on eNOSpThr497. Phosphatase inhibition suppresses both NO production and transendothelial resistance (TER) of ECs. In contrast, epigallocatechin-3-gallate (EGCG) signals ECs via the 67 kDa laminin-receptor (67LR) resulting in protein kinase A dependent activation of protein phosphatase-2A (PP2A). PP2A dephosphorylates MYPT1 pThr696 and thereby stimulates MP activity inducing dephosphorylation of eNOSpThr497 and the 20 kDa myosin II light chains. Thus an interplay of MP and PP2A is involved in the physiological regulation of EC functions implying that an EGCG dependent activation of these phosphatases leads to enhanced NO production and EC barrier improvement.

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