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

• DOI: 10.1038/srep44698
• Publication Date: 2017-03-16T11:16:39Z
• Authors: Róbert Bátori; Bálint Bécsi; Dénes Nagy; Zoltán Kónya; Csaba Hegedűs; Zsuzsanna Bordán; Alexander Verin; Beáta Lontay; Ferenc Erdődi
• Keywords: Nitric Oxide Synthase Type III; Pulmonary Artery; Nitric Oxide; Models, Biological; Article; Catechin; Cell Line; Myosin-Light-Chain Phosphatase; 03 medical and health sciences; Electric Impedance; Human Umbilical Vein Endothelial Cells; Animals; Humans; Elméleti orvostudományok; Gene Silencing; Protein Phosphatase 2; Phosphorylation; Oxazoles; Protein Kinase C; 0303 health sciences; Orvostudományok; Enzyme Activation; Phosphothreonine; Cattle; Marine Toxins

AbstractThe inhibitory phosphorylation of endothelial nitric oxide (NO) synthase (eNOS) at Thr497 (eNOSpThr497) 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 MYPT1pThr696 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.