Agonists Trigger G Protein-mediated Activation of the CPI-17 Inhibitor Phosphoprotein of Myosin Light Chain Phosphatase to Enhance Vascular Smooth Muscle Contractility

Kitazawa, Takio; Eto, Masumi; Woodsome, Terence P.; Brautigan, David L.

doi:10.1074/jbc.275.14.9897

Cited by 307 publications

(312 citation statements)

References 27 publications

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“…Most of the ROCK substrates are cellular proteins associated with the regulation of actin cytoskeleton. Besides MLC and MYPT1, this important subset of ROCK substrates include CPI-17 [63], calponin [58], LIM kinases [80,99,132], ezrin/radixin/moesin (ERM) [83], adducin [37], sodium-hydrogen exchanger (NHE1) [137] and ZIP kinase [43].…”

Section: Substrates Of Rockmentioning

confidence: 99%

Rho kinase in the regulation of cell death and survival

Shi

Wei

2007

Arch. Immunol. Ther. Exp.

219

183

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SummaryRho kinase (ROCK) belongs to a family of serine/threonine kinases that are activated via interaction with Rho GTPases. ROCK is involved in a wide range of fundamental cellular functions such as contraction, adhesion, migration, and proliferation. Recent studies have shown that ROCK plays an important role in the regulation of apoptosis in various cell types and animal disease models. Two ROCK isoforms, ROCK1 and ROCK2, are assumed to be function redundant, based largely on kinase construct overexpression, and chemical inhibitors (Y27632 and fasudil), which inhibit both ROCK1 and ROCK2. Gene targeting and RNA interference approaches allow further dissection of distinct cellular, physiological and patho-physiological functions of the two ROCK isoforms. This review, based on recent molecular, cellular and animal studies, focuses on the current understanding of ROCK signaling in the regulation of apoptosis and highlights the new findings from recently generated ROCK-deficient mice.

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Section: Substrates Of Rockmentioning

confidence: 99%

Rho kinase in the regulation of cell death and survival

Shi

Wei

2007

Arch. Immunol. Ther. Exp.

219

183

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“…PKC acts by phosphorylating an endogenous 17 kDa inhibitory protein, PKC-activated 17 kDa inhibitor protein of type 1 phosphatase (CPI-17). Phosphorylation of CPI-17 greatly augments its ability to bind to the catalytic subunit of MLCP, causing inhibition of MLCP activity [24][25][26]. Although Rho kinase is capable of activating CPI-17 in vitro, albeit to a lesser extent when compared with PKC, its predominant effect in vivo appears to be via phosphorylation of the large, 110-130 kDa regulatory myosin phosphatase targeting subunit (MYPT1), and inhibition of MLCP activity [27].…”

Section: Phosphorylation Of Sermentioning

confidence: 99%

“…Hydrolysis of phosphatidylcholine by PLD yields phosphatidic acid, which is dephosphorylated to diacylglycerol, leading to sustained activation of various protein kinase C (PKC) isoenzymes. Rho kinase and PKC, either singly or co-operatively, are involved in the inhibition of MLCP [12][13][14][15][16][17][23][24][25][26]. PKC acts by phosphorylating an endogenous 17 kDa inhibitory protein, PKC-activated 17 kDa inhibitor protein of type 1 phosphatase (CPI-17).…”

Section: Phosphorylation Of Sermentioning

confidence: 99%

“…Phosphorylation of CPI-17, MYPT1 and MLC 20 was measured by Western-blot analysis using phospho-specific antibodies as described previously [25,37,[41][42][43]. Dispersed muscle cells were treated with ACh for different time periods in the presence or absence of diphenylacetoxy-N-methylpiperidine (4-DAMP), methoctramine, bisindolylmaleimide or Y27632 and solubilized on ice for 1 h in a medium containing 20 mM Tris/HCl (pH 8.0), 1 mM DTT, 100 mM NaCl, 0.5 % SDS, 0.75 % deoxycholate, 1 mM PMSF, 10 µg/ml leupeptin and 100 µg/ml aprotinin.…”

Section: Western-blot Analysis Of Phosphoproteins (Cpi-17 Mypt1 and mentioning

confidence: 99%

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Differential signalling by muscarinic receptors in smooth muscle: m2-mediated inactivation of myosin light chain kinase via Gi3, Cdc42/Rac1 and p21-activated kinase 1 pathway, and m3-mediated MLC20 (20 kDa regulatory light chain of myosin II) phosphorylation via Rho-associated kinase/myosin phosphatase targeting subunit 1 and protein kinase C/CPI-17 pathway

Murthy

Zhou

Grider

et al. 2003

Biochemical Journal

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138

250

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Signalling via m3 and m2 receptors in smooth muscles involved activation of two G-protein-dependent pathways by each receptor. m2 receptors were coupled via Gβγ i3 with activation of phospholipase C-β3, phosphoinositide 3-kinase and Cdc42/Rac1 (where Cdc stands for cell division cycle) and p21-activated kinase 1 (PAK1), resulting in phosphorylation and inactivation of myosin light chain kinase (MLCK). Each step was inhibited by methoctramine and pertussis toxin. PAK1 activity was abolished in cells expressing both Cdc42-DN (where DN stands for dominant negative) and Rac1-DN. MLCK phosphorylation was inhibited by PAK1 antibody, and in cells expressing Cdc42-DN and Rac1-DN. m3 receptors were coupled via Gα q/11 with activation of phospholipase C-β1 and via RhoA with activation of Rho-associated kinase (Rho kinase), phospholipase D and protein kinase C (PKC). Rho kinase and phospholipase D activities were inhibited by C3 exoenzyme and in cells expressing RhoA-DN. PKC activity was inhibited by bisindolylmaleimide, and in cells expressing RhoA-DN; PKC activity was also inhibited partly by Y27632 (44 + − 5 %). PKC-induced phosphorylation of PKC-activated 17 kDa inhibitor protein of type 1 phosphatase (CPI-17) at Thr 38 was abolished by bisindolylmaleimide and inhibited partly by Y27632 (28 + − 3 %). Rho-kinase-induced phosphorylation of myosin phosphatase targeting subunit (MYPT1) and was abolished by Y27632. Sustained phosphorylation of 20 kDa regulatory light chain of myosin II (MLC 20 ) and contraction were abolished by bisindolylmaleimide Y27632 and C3 exoenzyme and in cells expressing RhoA-DN. The results suggest that Rho-kinase-dependent phosphorylation of MYPT1 and PKCdependent phosphorylation and enhancement of CPI-17 binding to the catalytic subunit of MLC phosphatase (MLCP) act co-operatively to inhibit MLCP activity, leading to sustained stimulation of MLC 20 phosphorylation and contraction. Because Y27632 inhibited both Rho kinase and PKC activities, it could not be used to ascertain the contribution of MYPT1 to inhibition of MLCP activity. m2-dependent phosphorylation and inactivation of MLCK precluded its involvement in sustained MLC 20 phosphorylation and contraction.

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“…Phosphorylation of Thr38 of CPI-17 increases over 10 3 -fold in inhibitory potency for MP [2]. CPI-17 is exclusively expressed in smooth muscle and brain [2], and suggested to function controlling vascular tone [3] and maintaining cerebellar memory [4]. A series of biochemical experiments suggested that this inhibition is achieved by the direct binding of phospho-Thr38 on CPI-17 to the catalytic site of MP [5 -7].…”

Section: Introductionmentioning

confidence: 99%

Phosphorylation-induced conformational change responsible for the function of a myosin phosphatase inhibitor, CPI-17

Ohki

Eto

Takada

et al. 2004

Science and Technology of Advanced Materials

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The structures of CPI-17 (Protein kinase-C dependent protein phosphatase-1 (PP1) inhibitor of 17 kDa) in an inactive and an active form have been determined by multidimensional NMR spectroscopy. Comparison of the two structures revealed how the molecular switch turns on at atomic resolution. Using the NMR structure of CPI-17 in the active form, the binding with catalytic domain of PP1 (PP1c) was simulated and the binding model is proposed in this report. When the phospho-Thr38 docks to the catalytic site of PP1, possible interactions for the tight binding are found; one is electrostatic interaction between a negatively charged cluster on phospho-CPI-17 and an acidic groove of PP1c, and the other is hydrophobic interaction between a hydrophobic surface area of phospho-CPI-17 and a hydrophobic groove of PP1c. q

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Agonists Trigger G Protein-mediated Activation of the CPI-17 Inhibitor Phosphoprotein of Myosin Light Chain Phosphatase to Enhance Vascular Smooth Muscle Contractility

Cited by 307 publications

References 27 publications

Rho kinase in the regulation of cell death and survival

Rho kinase in the regulation of cell death and survival

Phosphorylation-induced conformational change responsible for the function of a myosin phosphatase inhibitor, CPI-17

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