title: Okadaic acid induces phosphorylation and translocation of myosin phosphatase target subunit 1 influencing myosin phosphorylation, stress fiber assembly and cell migration in HepG2 cells.
abstract: It was determined that the myosin phosphatase (MP) activity and content of myosin phosphatase target subunit 1 (MYPT1) were correlated in subcellular fractions of human hepatocarcinoma (HepG2) cells. In control cells MYPT1 was localized in the cytoplasm and in the nucleus, as determined by confocal microscopy. Treatment of HepG2 cells with 50 nM okadaic acid (OA), a cell-permeable phosphatase inhibitor, induced several changes: 1) a marked redistribution of MYPT1 to the plasma membrane associated with an increased level of phosphorylation of MYPT1 at Thr695. Both effects showed only a slight influence with the Rho-kinase inhibitor, Y-27632; 2) an increase in phosphorylation of MYPT1 at Thr850 associated with its accumulation in the perinuclear region and nucleus. These effects were markedly reduced by Y-27632; 3) an increased phosphorylation of the 20 kDa myosin II light chain at Ser19 associated with an increased location of myosin II at the cell center. These effects were partially counteracted by Y-27632; 4) an increase in stress fiber formation and a decrease in cell migration, both OA-induced effects were blocked by Y-27632. In HepG2 lysates, OA (5-100 nM) did not affect MP activity but inhibited PP2A activity. These results indicate that OA induces differential phosphorylation and translocation of MYPT1, dependent on PP2A and, to varying extents, on ROK. These changes are associated with an increased level of myosin II phosphorylation and attenuation of hepatic cell migration.
authors.affiliation: Department of Medical Chemistry, Research Center for Molecular Medicine, Medical and Health Science Center, University of Debrecen, H-4026 Debrecen, Bem tér 18/B, Hungary.
authors.forename: Beáta
authors.initials: B
authors.lastname: Lontay
authors.forename: Andrea
authors.initials: A
authors.lastname: Kiss
authors.forename: Pál
authors.initials: P
authors.lastname: Gergely
authors.forename: David J
authors.initials: DJ
authors.lastname: Hartshorne
authors.forename: Ferenc
authors.initials: F
authors.lastname: Erdodi
mesh_terms: D000577:Amides / Q000494:pharmacology; D051017:Apoptosis Regulatory Proteins; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects*; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D064550:Death-Associated Protein Kinases; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008856:Microscopy, Fluorescence; D018994:Myosin Light Chains / Q000378:metabolism; D043263:Myosin-Light-Chain Phosphatase / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D019319:Okadaic Acid / Q000494:pharmacology*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects; D011725:Pyridines / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D022502:Stress Fibers / Q000187:drug effects*; D013347:Subcellular Fractions; D054460:rho-Associated Kinases
chemical_list: D000577:Amides; D051017:Apoptosis Regulatory Proteins; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; C466107:MRLC2 protein, human; D018994:Myosin Light Chains; D011725:Pyridines; C108830:Y 27632; D019319:Okadaic Acid; D064550:Death-Associated Protein Kinases; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases; D043263:Myosin-Light-Chain Phosphatase; C490372:PPP1R12A protein, human; D009218:Myosins