PMID- 32502416 OWN - NLM STAT- MEDLINE DCOM- 20210809 LR - 20210809 IS - 1879-0445 (Electronic) IS - 0960-9822 (Print) IS - 0960-9822 (Linking) VI - 30 IP - 13 DP - 2020 Jul 6 TI - Tyrosine Phosphorylation of the Myosin Regulatory Light Chain Controls Non-muscle Myosin II Assembly and Function in Migrating Cells. PG - 2446-2458.e6 LID - S0960-9822(20)30573-X [pii] LID - 10.1016/j.cub.2020.04.057 [doi] AB - Active non-muscle myosin II (NMII) enables migratory cell polarization and controls dynamic cellular processes, such as focal adhesion formation and turnover and cell division. Filament assembly and force generation depend on NMII activation through the phosphorylation of Ser19 of the regulatory light chain (RLC). Here, we identify amino acid Tyr (Y) 155 of the RLC as a novel regulatory site that spatially controls NMII function. We show that Y155 is phosphorylated in vitro by the Tyr kinase domain of epidermal growth factor (EGF) receptor. In cells, phosphorylation of Y155, or its phospho-mimetic mutation (Glu), prevents the interaction of RLC with the myosin heavy chain (MHCII) to form functional NMII units. Conversely, Y155 mutation to a structurally similar but non-phosphorylatable amino acid (Phe) restores the more dynamic cellular functions of NMII, such as myosin filament formation and nascent adhesion assembly, but not those requiring stable actomyosin bundles, e.g., focal adhesion elongation or migratory front-back polarization. In live cells, phospho-Y155 RLC is prominently featured in protrusions, where it prevents NMII assembly. Our data indicate that Y155 phosphorylation constitutes a novel regulatory mechanism that contributes to the compartmentalization of NMII assembly and function in live cells. CI - Copyright (c) 2020 Elsevier Inc. All rights reserved. FAU - Aguilar-Cuenca, Rocio AU - Aguilar-Cuenca R AD - Instituto de Investigacion Sanitaria-Hospital Universitario de la Princesa, 28006 Madrid, Spain; Universidad Autonoma de Madrid School of Medicine, 28006 Madrid, Spain. FAU - Llorente-Gonzalez, Clara AU - Llorente-Gonzalez C AD - Molecular Mechanisms Program, Centro de Investigacion del Cancer and Instituto de Biologia Molecular y Celular del Cancer, Consejo Superior de Investigaciones Cientificas (CSIC)-University of Salamanca, 37007 Salamanca, Spain. FAU - Chapman, Jessica R AU - Chapman JR AD - Department of Chemistry, University of Virginia, Charlottesville, VA 22903, USA. FAU - Talayero, Vanessa C AU - Talayero VC AD - Molecular Mechanisms Program, Centro de Investigacion del Cancer and Instituto de Biologia Molecular y Celular del Cancer, Consejo Superior de Investigaciones Cientificas (CSIC)-University of Salamanca, 37007 Salamanca, Spain. FAU - Garrido-Casado, Marina AU - Garrido-Casado M AD - Molecular Mechanisms Program, Centro de Investigacion del Cancer and Instituto de Biologia Molecular y Celular del Cancer, Consejo Superior de Investigaciones Cientificas (CSIC)-University of Salamanca, 37007 Salamanca, Spain. FAU - Delgado-Arevalo, Cristina AU - Delgado-Arevalo C AD - Instituto de Investigacion Sanitaria-Hospital Universitario de la Princesa, 28006 Madrid, Spain; Universidad Autonoma de Madrid School of Medicine, 28006 Madrid, Spain. FAU - Millan-Salanova, Maria AU - Millan-Salanova M AD - Molecular Mechanisms Program, Centro de Investigacion del Cancer and Instituto de Biologia Molecular y Celular del Cancer, Consejo Superior de Investigaciones Cientificas (CSIC)-University of Salamanca, 37007 Salamanca, Spain. FAU - Shabanowitz, Jeffrey AU - Shabanowitz J AD - Department of Chemistry, University of Virginia, Charlottesville, VA 22903, USA. FAU - Hunt, Donald F AU - Hunt DF AD - Department of Pathology, University of Virginia, Charlottesville, VA 22903, USA; Department of Chemistry, University of Virginia, Charlottesville, VA 22903, USA. FAU - Sellers, James R AU - Sellers JR AD - Cell Biology and Developmental Biology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA. FAU - Heissler, Sarah M AU - Heissler SM AD - Department of Physiology and Cell Biology, The Ohio State University Wexner Medical Center, Columbus, OH 43210, USA. FAU - Vicente-Manzanares, Miguel AU - Vicente-Manzanares M AD - Molecular Mechanisms Program, Centro de Investigacion del Cancer and Instituto de Biologia Molecular y Celular del Cancer, Consejo Superior de Investigaciones Cientificas (CSIC)-University of Salamanca, 37007 Salamanca, Spain. Electronic address: miguel.vicente@csic.es. LA - eng GR - K22 HL131869/HL/NHLBI NIH HHS/United States GR - R01 GM037537/GM/NIGMS NIH HHS/United States GR - Z01 HL001786/ImNIH/Intramural NIH HHS/United States GR - ZIA HL001786/ImNIH/Intramural NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural PT - Research Support, N.I.H., Intramural PT - Research Support, Non-U.S. Gov't DEP - 20200604 PL - England TA - Curr Biol JT - Current biology : CB JID - 9107782 RN - 0 (Myosin Light Chains) RN - 42HK56048U (Tyrosine) RN - EC 3.6.1.- (Myosin Type II) SB - IM MH - A549 Cells MH - Animals MH - CHO Cells MH - Cell Movement/*physiology MH - Cricetulus MH - HEK293 Cells MH - Humans MH - Myosin Light Chains/*metabolism MH - Myosin Type II/*metabolism MH - Phosphorylation MH - Sf9 Cells MH - Spodoptera/physiology MH - Tyrosine/*metabolism PMC - PMC7343590 MID - NIHMS1589970 OTO - NOTNLM OT - actin OT - cytoskeleton OT - migration OT - myosin OT - phosphorylation OT - tyrosine COIS- Declaration of Interests The authors declare no competing interests. EDAT- 2020/06/06 06:00 MHDA- 2021/08/10 06:00 PMCR- 2021/07/06 CRDT- 2020/06/06 06:00 PHST- 2019/04/02 00:00 [received] PHST- 2020/03/04 00:00 [revised] PHST- 2020/04/22 00:00 [accepted] PHST- 2020/06/06 06:00 [pubmed] PHST- 2021/08/10 06:00 [medline] PHST- 2020/06/06 06:00 [entrez] PHST- 2021/07/06 00:00 [pmc-release] AID - S0960-9822(20)30573-X [pii] AID - 10.1016/j.cub.2020.04.057 [doi] PST - ppublish SO - Curr Biol. 2020 Jul 6;30(13):2446-2458.e6. doi: 10.1016/j.cub.2020.04.057. Epub 2020 Jun 4.