Phosphorylation-Induced Conformational Switching of CPI-17 Produces a Potent Myosin Phosphatase Inhibitor

Eto, Masumi; Kitazawa, Takio; Matsuzawa, Fumiko; Aikawa, Sei Ichi; Kirkbride, Jason A.; Isozumi, Noriyoshi; Nishimura, Yumi; Brautigan, David L.; Ohki, Shiro

doi:10.1016/j.str.2007.10.014

Cited by 46 publications

(49 citation statements)

References 61 publications

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“…As indicated in a recent paper by Eto et al (17), CPI-17 phosphorylation induced conformational changes in CPI-17, making it a potent inhibitor of MLCP. Thus, phosphorylated CPI-17 caused increased inhibition of MLCP activity (27).…”

Section: Discussionmentioning

confidence: 88%

Deletion of SM-B, the high ATPase isoform of myosin, upregulates the PKC-mediated signal transduction pathway in murine urinary bladder smooth muscle

Hypolite¹,

Chang²,

LaBelle³

et al. 2009

American Journal of Physiology-Renal Physiology

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Hypolite JA, Chang S, LaBelle E, Babu GJ, Periasamy M, Wein AJ, Chacko S. Deletion of SM-B, the high ATPase isoform of myosin, upregulates the PKC-mediated signal transduction pathway in murine urinary bladder smooth muscle. Am J Physiol Renal Physiol 296: F658 -F665, 2009. First published December 3, 2008 doi:10.1152/ajprenal.90221.2008.-Detrusor smooth muscle (DSM) hypertrophy induced by partial bladder outlet obstruction (PBOO) is associated with changes in the NH 2-terminal myosin heavy chain isoform from predominantly SM-B to SM-A, alteration in the Ca 2ϩ sensitization pathway, and the contractile characteristics from phasic to tonic in rabbits. We utilized the SM-B knockout (KO) mouse to determine whether a shift from SM-B to SM-A without PBOO is associated with changes in the signal transduction pathway mediated via PKC and CPI-17, which keeps the myosin phosphorylation (MLC 20) level high by inhibiting the myosin phosphatase. DSM strips from SM-B KO mice generated more force in response to electrical field stimulation, KCl, carbachol, and phorbol 12,13-dibutyrate than that of age-matched wild-type mice. There was no difference in the ED 50 for carbachol but the maximum response was greater for the SM-B KO mice. DSM from SM-B KO mice revealed increased mass and hypertrophy. The KO mice also showed an overexpression of PKC-␣, increased levels of phospho-CPI-17, and an elevated level of IP 3 and DAG upon stimulation with carbachol. Two-dimensional gel electrophoresis revealed an increased level of MLC 20 phosphorylation in response to carbachol. Together, these changes may be responsible for the higher level of force generation and maintenance by the DSM from the SM-B KO bladders. In conclusion, our data show that ablation of SM-B is associated with alteration of PKC-mediated signal transduction and CPI-17-mediated Ca 2ϩ sensitization pathway that regulate smooth muscle contraction. Interestingly, similar changes are also present in PBOO-induced DSM compensatory response in the rabbit model in which SM-B is downregulated. smooth muscle contraction; alternative splicing; CPI-17; light chain phosphorylation MYOSIN II IS THE MAJOR COMPONENT of the thick filament in smooth muscles from all sources. Smooth muscle myosin II is regulated by alternative splicing at both 3Ј and 5Ј end of the myosin heavy chain (MHC) pre-mRNA producing COOH-terminal (SM1 and SM2) and NH 2 -terminal(SM-A and SM-B) MHC isoforms, respectively. Alternative splicing at the 5Ј end inserts a 21-nt insertion that encodes a seven amino acid sequence in the NH 2 -terminal head region of the myosin near the ATP-binding site (3,26). Studies have shown that smooth muscle myosin containing the SM-B isoform (with the 7-amino acid insert) have a higher actin-activated ATPase activity and a higher shortening velocity compared with those smooth muscle which predominantly consist of SM-A, lacking the seven amino acid insert (14,26,29). Studies have also shown that smooth muscles containing primarily this high-velocity isoform (SM-B), such as the urin...

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Section: Discussionmentioning

confidence: 88%

Deletion of SM-B, the high ATPase isoform of myosin, upregulates the PKC-mediated signal transduction pathway in murine urinary bladder smooth muscle

Hypolite¹,

Chang²,

LaBelle³

et al. 2009

American Journal of Physiology-Renal Physiology

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“…Therefore, the autoinhibitory domain of MYPT1 likely recognizes the active site modulated by the allosteric regulation with the N-terminal domain of MYPT1. Interestingly, the N-terminal 19 residues of MYPT1, located near the PP1 active site, are directly associated with phosphorylated CPI-17 (59). Therefore, it is also possible that the autoinhibitory domain recognizes the MYPT1 N-terminal segment as a marker of the active site.…”

Section: Discussionmentioning

confidence: 99%

Phosphorylation-dependent Autoinhibition of Myosin Light Chain Phosphatase Accounts for Ca2+ Sensitization Force of Smooth Muscle Contraction

Khromov

Choudhury

Stevenson

et al. 2009

Journal of Biological Chemistry

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140

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“…58,59 Computer modeling predicts direct binding between MYPT1 and phospho-CPI-17, suggesting that MYPT1 may dock CPI-17 to the MLCP holoenzyme, accounting for the specific inhibition of MLCP by CPI-17. 60 Furthermore, when bound to MYPT1, the PPP1Cd catalytic subunit of MLCP is unable to hydrolyze phospho-Thr38 of CPI-17, so phospho-CPI-17 is able to bind at the active site of PPP1Cδ and suppress its activity.…”

Section: Inhibition Of Myosin Light Chain Phosphatase Activity By Cpimentioning

confidence: 99%

Calcium Sensitization Mechanisms in Gastrointestinal Smooth Muscles

Perrino

2016

J Neurogastroenterol Motil

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An increase in intracellular Ca2+ is the primary trigger of contraction of gastrointestinal (GI) smooth muscles. However, increasing the Ca 2+ sensitivity of the myofilaments by elevating myosin light chain phosphorylation also plays an essential role. Inhibiting myosin light chain phosphatase activity with protein kinase C-potentiated phosphatase inhibitor protein-17 kDa (CPI-17) and myosin phosphatase targeting subunit 1 (MYPT1) phosphorylation is considered to be the primary mechanism underlying myofilament Ca 2+ sensitization. The relative importance of Ca 2+ sensitization mechanisms to the diverse patterns of GI motility is likely related to the varied functional roles of GI smooth muscles. Increases in CPI-17 and MYPT1 phosphorylation in response to agonist stimulation regulate myosin light chain phosphatase activity in phasic, tonic, and sphincteric GI smooth muscles. Recent evidence suggests that MYPT1 phosphorylation may also contribute to force generation by reorganization of the actin cytoskeleton. The mechanisms responsible for maintaining constitutive CPI-17 and MYPT1 phosphorylation in GI smooth muscles are still largely unknown. The characteristics of the cell-types comprising the neuroeffector junction lead to fundamental differences between the effects of exogenous agonists and endogenous neurotransmitters on Ca 2+ sensitization mechanisms. The contribution of various cell-types within the tunica muscularis to the motor responses of GI organs to neurotransmission must be considered when determining the mechanisms by which Ca 2+ sensitization pathways are activated. The signaling pathways regulating Ca 2+ sensitization may provide novel therapeutic strategies for controlling GI motility. This article will provide an overview of the current understanding of the biochemical basis for the regulation of Ca

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Phosphorylation-Induced Conformational Switching of CPI-17 Produces a Potent Myosin Phosphatase Inhibitor

Cited by 46 publications

References 61 publications

Deletion of SM-B, the high ATPase isoform of myosin, upregulates the PKC-mediated signal transduction pathway in murine urinary bladder smooth muscle

Deletion of SM-B, the high ATPase isoform of myosin, upregulates the PKC-mediated signal transduction pathway in murine urinary bladder smooth muscle

Phosphorylation-dependent Autoinhibition of Myosin Light Chain Phosphatase Accounts for Ca2+ Sensitization Force of Smooth Muscle Contraction

Calcium Sensitization Mechanisms in Gastrointestinal Smooth Muscles

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