{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,15]],"date-time":"2026-10-15T00:33:56Z","timestamp":1792024436211,"version":"4.4.0"},"reference-count":57,"publisher":"eLife Sciences Publications, Ltd","license":[{"start":{"date-parts":[[2017,9,26]],"date-time":"2017-09-26T00:00:00Z","timestamp":1506384000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2017,9,26]],"date-time":"2017-09-26T00:00:00Z","timestamp":1506384000000},"content-version":"am","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"},{"start":{"date-parts":[[2017,9,26]],"date-time":"2017-09-26T00:00:00Z","timestamp":1506384000000},"content-version":"tdm","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["R01-056322"],"award-info":[{"award-number":["R01-056322"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000928","name":"Welch Foundation","doi-asserted-by":"publisher","award":["I-1544"],"award-info":[{"award-number":["I-1544"]}],"id":[{"id":"10.13039\/100000928","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000011","name":"Howard Hughes Medical Institute","doi-asserted-by":"publisher","id":[{"id":"10.13039\/100000011","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Iowa State University & Roy J. Carver Charitable Trust"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"abstract":"<jats:p>The Rho GTPase Rac1 activates the WAVE regulatory complex (WRC) to drive Arp2\/3 complex-mediated actin polymerization, which underpins diverse cellular processes. Here we report the structure of a WRC-Rac1 complex determined by cryo-electron microscopy. Surprisingly, Rac1 is not located at the binding site on the Sra1 subunit of the WRC previously identified by mutagenesis and biochemical data. Rather, it binds to a distinct, conserved site on the opposite end of Sra1. Biophysical and biochemical data on WRC mutants confirm that Rac1 binds to both sites, with the newly identified site having higher affinity and both sites required for WRC activation. Our data reveal that the WRC is activated by simultaneous engagement of two Rac1 molecules, suggesting a mechanism by which cells may sense the density of active Rac1 at membranes to precisely control actin assembly.<\/jats:p>","DOI":"10.7554\/elife.29795","type":"journal-article","created":{"date-parts":[[2017,9,26]],"date-time":"2017-09-26T07:00:15Z","timestamp":1506409215000},"source":"Crossref","is-referenced-by-count":181,"title":["Rac1 GTPase activates the WAVE regulatory complex through two distinct binding sites"],"prefix":"10.7554","volume":"6","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-6366-159X","authenticated-orcid":true,"given":"Baoyu","family":"Chen","sequence":"first","affiliation":[{"name":"Department of Biophysics, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"},{"name":"Howard Hughes Medical Institute, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hui-Ting","family":"Chou","sequence":"additional","affiliation":[{"name":"Department of Cell Biology, Harvard Medical School, Boston, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chad A","family":"Brautigam","sequence":"additional","affiliation":[{"name":"Department of Biophysics, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"},{"name":"Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wenmin","family":"Xing","sequence":"additional","affiliation":[{"name":"Department of Biophysics, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"},{"name":"Howard Hughes Medical Institute, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sheng","family":"Yang","sequence":"additional","affiliation":[{"name":"Roy J. Carver Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lisa","family":"Henry","sequence":"additional","affiliation":[{"name":"Department of Biophysics, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"},{"name":"Howard Hughes Medical Institute, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lynda K","family":"Doolittle","sequence":"additional","affiliation":[{"name":"Department of Biophysics, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"},{"name":"Howard Hughes Medical Institute, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Thomas","family":"Walz","sequence":"additional","affiliation":[{"name":"Rockefeller University, New York, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0775-7917","authenticated-orcid":true,"given":"Michael K","family":"Rosen","sequence":"additional","affiliation":[{"name":"Department of Biophysics, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"},{"name":"Howard Hughes Medical Institute, University of Texas Southwestern Medical Center at Dallas, Dallas, United States"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"4374","published-online":{"date-parts":[[2017,9,26]]},"reference":[{"key":"bib1","doi-asserted-by":"publisher","first-page":"W529","DOI":"10.1093\/nar\/gkq399","article-title":"ConSurf 2010: calculating evolutionary conservation in sequence and structure of proteins and nucleic acids","volume":"38","author":"Ashkenazy","year":"2010","journal-title":"Nucleic Acids Research"},{"key":"bib2","doi-asserted-by":"publisher","first-page":"81","DOI":"10.1073\/pnas.0408399102","article-title":"Studying multiprotein complexes by multisignal sedimentation velocity analytical ultracentrifugation","volume":"102","author":"Balbo","year":"2005","journal-title":"PNAS"},{"key":"bib3","doi-asserted-by":"publisher","first-page":"e04123","DOI":"10.7554\/eLife.04123","article-title":"Phase transitions of multivalent proteins can promote clustering of membrane receptors","volume":"3","author":"Banjade","year":"2014","journal-title":"eLife"},{"key":"bib4","doi-asserted-by":"publisher","first-page":"565","DOI":"10.1016\/j.ceb.2013.04.001","article-title":"Membrane and actin dynamics interplay at lamellipodia leading edge","volume":"25","author":"Bisi","year":"2013","journal-title":"Current Opinion in Cell Biology"},{"key":"bib5","doi-asserted-by":"publisher","first-page":"237","DOI":"10.1038\/nrm2867","article-title":"A nucleator arms race: cellular control of actin assembly","volume":"11","author":"Campellone","year":"2010","journal-title":"Nature Reviews Microbiology"},{"key":"bib6","doi-asserted-by":"publisher","first-page":"2745","DOI":"10.15252\/embj.201488837","article-title":"Nanoscale segregation of actin nucleation and elongation factors determines dendritic spine protrusion","volume":"33","author":"Chazeau","year":"2014","journal-title":"The EMBO Journal"},{"key":"bib7","doi-asserted-by":"publisher","first-page":"533","DOI":"10.1038\/nature09623","article-title":"Structure and control of the actin regulatory WAVE complex","volume":"468","author":"Chen","year":"2010","journal-title":"Nature"},{"key":"bib8","doi-asserted-by":"publisher","first-page":"195","DOI":"10.1016\/j.cell.2013.11.048","article-title":"The WAVE regulatory complex links diverse receptors to the actin cytoskeleton","volume":"156","author":"Chen","year":"2014","journal-title":"Cell"},{"key":"bib9","doi-asserted-by":"publisher","first-page":"55","DOI":"10.1016\/B978-0-12-397924-7.00004-2","article-title":"Biochemical reconstitution of the WAVE regulatory complex","volume":"540","author":"Chen","year":"2014","journal-title":"Methods in Enzymology"},{"key":"bib10","doi-asserted-by":"publisher","first-page":"569","DOI":"10.1016\/j.devcel.2014.08.001","article-title":"Ena\/VASP proteins cooperate with the WAVE complex to regulate the actin cytoskeleton","volume":"30","author":"Chen","year":"2014","journal-title":"Developmental Cell"},{"key":"bib11","doi-asserted-by":"publisher","first-page":"557","DOI":"10.1038\/nmeth.3891","article-title":"Quantitative assessment of fluorescent proteins","volume":"13","author":"Cranfill","year":"2016","journal-title":"Nature Methods"},{"key":"bib12","doi-asserted-by":"publisher","first-page":"2569","DOI":"10.1101\/gad.9.21.2569","article-title":"Abi-2, a novel SH3-containing protein interacts with the c-Abl tyrosine kinase and modulates c-Abl transforming activity","volume":"9","author":"Dai","year":"1995","journal-title":"Genes & Development"},{"key":"bib13","doi-asserted-by":"publisher","first-page":"167","DOI":"10.1016\/0092-8674(86)90495-2","article-title":"Biological and biochemical properties of human rasH genes mutated at codon 61","volume":"44","author":"Der","year":"1986","journal-title":"Cell"},{"key":"bib14","doi-asserted-by":"publisher","first-page":"W522","DOI":"10.1093\/nar\/gkm276","article-title":"PDB2PQR: expanding and upgrading automated preparation of biomolecular structures for molecular simulations","volume":"35","author":"Dolinsky","year":"2007","journal-title":"Nucleic Acids Research"},{"key":"bib15","doi-asserted-by":"publisher","first-page":"273","DOI":"10.1007\/978-1-62703-538-5_16","article-title":"Measurement and analysis of in vitro actin polymerization","volume":"1046","author":"Doolittle","year":"2013","journal-title":"Methods in Molecular Biology"},{"key":"bib16","doi-asserted-by":"publisher","first-page":"790","DOI":"10.1038\/nature00859","article-title":"Mechanism of regulation of WAVE1-induced actin nucleation by Rac1 and Nck","volume":"418","author":"Eden","year":"2002","journal-title":"Nature"},{"key":"bib17","doi-asserted-by":"publisher","first-page":"1051","DOI":"10.1016\/j.cell.2013.01.051","article-title":"Regulation of WASH-dependent actin polymerization and protein trafficking by ubiquitination","volume":"152","author":"Hao","year":"2013","journal-title":"Cell"},{"key":"bib18","doi-asserted-by":"publisher","first-page":"663","DOI":"10.1038\/373663b0","article-title":"Improved green fluorescence","volume":"373","author":"Heim","year":"1995","journal-title":"Nature"},{"key":"bib19","doi-asserted-by":"publisher","first-page":"251","DOI":"10.1016\/j.cell.2012.06.024","article-title":"A landscape of driver mutations in melanoma","volume":"150","author":"Hodis","year":"2012","journal-title":"Cell"},{"key":"bib20","doi-asserted-by":"publisher","first-page":"47","DOI":"10.1016\/j.jsb.2006.07.003","article-title":"SPARX, a new environment for Cryo-EM image processing","volume":"157","author":"Hohn","year":"2007","journal-title":"Journal of Structural Biology"},{"key":"bib21","doi-asserted-by":"publisher","first-page":"561","DOI":"10.1038\/nsmb.1587","article-title":"The wave regulatory complex is inhibited","volume":"16","author":"Ismail","year":"2009","journal-title":"Nature Structural & Molecular Biology"},{"key":"bib22","doi-asserted-by":"publisher","first-page":"151","DOI":"10.1038\/35004513","article-title":"Autoinhibition and activation mechanisms of the Wiskott-Aldrich syndrome protein","volume":"404","author":"Kim","year":"2000","journal-title":"Nature"},{"key":"bib23","doi-asserted-by":"publisher","first-page":"14449","DOI":"10.1073\/pnas.1107666108","article-title":"WAVE regulatory complex activation by cooperating GTPases Arf and Rac1","volume":"108","author":"Koronakis","year":"2011","journal-title":"PNAS"},{"key":"bib24","doi-asserted-by":"publisher","first-page":"1006","DOI":"10.1038\/ng.2359","article-title":"Exome sequencing identifies recurrent somatic RAC1 mutations in melanoma","volume":"44","author":"Krauthammer","year":"2012","journal-title":"Nature Genetics"},{"key":"bib25","doi-asserted-by":"publisher","first-page":"63","DOI":"10.1038\/nmeth.2727","article-title":"Quantifying the local resolution of cryo-EM density maps","volume":"11","author":"Kucukelbir","year":"2014","journal-title":"Nature Methods"},{"key":"bib26","doi-asserted-by":"publisher","first-page":"1508","DOI":"10.1126\/science.1245503","article-title":"C57BL\/6N mutation in cytoplasmic FMRP interacting protein 2 regulates cocaine response","volume":"342","author":"Kumar","year":"2013","journal-title":"Science"},{"key":"bib27","doi-asserted-by":"publisher","first-page":"260","DOI":"10.1006\/abio.1996.0238","article-title":"Program DYNAFIT for the analysis of enzyme kinetic data: application to HIV proteinase","volume":"237","author":"Kuzmic","year":"1996","journal-title":"Analytical Biochemistry"},{"key":"bib28","first-page":"155","article-title":"Structure and role of WASP and WAVE in Rho GTPase signalling in cancer","volume":"11","author":"Lane","year":"2014","journal-title":"Cancer Genomics & Proteomics"},{"key":"bib29","doi-asserted-by":"publisher","first-page":"512","DOI":"10.1016\/j.molcel.2009.10.024","article-title":"Activation of the WAVE complex by coincident signals controls actin assembly","volume":"36","author":"Lebensohn","year":"2009","journal-title":"Molecular Cell"},{"key":"bib30","doi-asserted-by":"publisher","first-page":"789","DOI":"10.1083\/jcb.151.4.789","article-title":"Fission yeast myosin-I, Myo1p, stimulates actin assembly by Arp2\/3 complex and shares functions with WASp","volume":"151","author":"Lee","year":"2000","journal-title":"The Journal of Cell Biology"},{"key":"bib31","doi-asserted-by":"publisher","first-page":"584","DOI":"10.1038\/nmeth.2472","article-title":"Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM","volume":"10","author":"Li","year":"2013","journal-title":"Nature Methods"},{"key":"bib32","doi-asserted-by":"publisher","first-page":"732","DOI":"10.1038\/35047107","article-title":"IRSp53 is an essential intermediate between Rac and WAVE in the regulation of membrane ruffling","volume":"408","author":"Miki","year":"2000","journal-title":"Nature"},{"key":"bib33","doi-asserted-by":"publisher","first-page":"334","DOI":"10.1016\/S1047-8477(03)00069-8","article-title":"Accurate determination of local defocus and specimen tilt in electron microscopy","volume":"142","author":"Mindell","year":"2003","journal-title":"Journal of Structural Biology"},{"key":"bib34","doi-asserted-by":"publisher","first-page":"8326","DOI":"10.1523\/JNEUROSCI.1482-08.2008","article-title":"Cdk5 phosphorylation of WAVE2 regulates oligodendrocyte precursor cell migration through nonreceptor tyrosine kinase Fyn","volume":"28","author":"Miyamoto","year":"2008","journal-title":"Journal of Neuroscience"},{"key":"bib35","doi-asserted-by":"publisher","first-page":"85","DOI":"10.3109\/10409238.2014.999191","article-title":"Structures of Ras superfamily effector complexes: What have we learnt in two decades?","volume":"50","author":"Mott","year":"2015","journal-title":"Critical Reviews in Biochemistry and Molecular Biology"},{"key":"bib36","doi-asserted-by":"publisher","first-page":"89","DOI":"10.1016\/j.ab.2010.07.017","article-title":"Determination of protein complex stoichiometry through multisignal sedimentation velocity experiments","volume":"407","author":"Padrick","year":"2010","journal-title":"Analytical Biochemistry"},{"key":"bib37","doi-asserted-by":"publisher","first-page":"707","DOI":"10.1146\/annurev.biochem.77.060407.135452","article-title":"Physical mechanisms of signal integration by WASP family proteins","volume":"79","author":"Padrick","year":"2010","journal-title":"Annual Review of Biochemistry"},{"key":"bib38","doi-asserted-by":"publisher","first-page":"E472","DOI":"10.1073\/pnas.1100236108","article-title":"Arp2\/3 complex is bound and activated by two WASP proteins","volume":"108","author":"Padrick","year":"2011","journal-title":"PNAS"},{"key":"bib39","doi-asserted-by":"publisher","first-page":"1605","DOI":"10.1002\/jcc.20084","article-title":"UCSF Chimera--a visualization system for exploratory research and analysis","volume":"25","author":"Pettersen","year":"2004","journal-title":"Journal of Computational Chemistry"},{"key":"bib40","doi-asserted-by":"publisher","first-page":"4061","DOI":"10.1091\/mbc.E10-08-0683","article-title":"A guide to simple and informative binding assays","volume":"21","author":"Pollard","year":"2010","journal-title":"Molecular Biology of the Cell"},{"key":"bib41","doi-asserted-by":"publisher","first-page":"801","DOI":"10.1126\/science.290.5492.801","article-title":"Integration of multiple signals through cooperative regulation of the N-WASP-Arp2\/3 complex","volume":"290","author":"Prehoda","year":"2000","journal-title":"Science"},{"key":"bib42","doi-asserted-by":"publisher","first-page":"7","DOI":"10.1038\/nrm3492","article-title":"New insights into the regulation and cellular functions of the ARP2\/3 complex","volume":"14","author":"Rotty","year":"2013","journal-title":"Nature Reviews Molecular Cell Biology"},{"key":"bib43","doi-asserted-by":"publisher","first-page":"519","DOI":"10.1016\/j.jsb.2012.09.006","article-title":"RELION: implementation of a Bayesian approach to cryo-EM structure determination","volume":"180","author":"Scheres","year":"2012","journal-title":"Journal of Structural Biology"},{"key":"bib44","volume-title":"The PyMOL Molecular Graphics System","author":"Schrodinger","year":"2015","unstructured":"Schrodinger LLC. 2015. The PyMOL Molecular Graphics System. 1.8."},{"key":"bib45","doi-asserted-by":"publisher","first-page":"104","DOI":"10.1016\/S0003-2697(03)00289-6","article-title":"On the analysis of protein self-association by sedimentation velocity analytical ultracentrifugation","volume":"320","author":"Schuck","year":"2003","journal-title":"Analytical Biochemistry"},{"key":"bib46","doi-asserted-by":"publisher","first-page":"285","DOI":"10.1146\/annurev-biophys-060414-034308","article-title":"Specification of Architecture and Function of Actin Structures by Actin Nucleation Factors","volume":"44","author":"Skau","year":"2015","journal-title":"Annual Review of Biophysics"},{"key":"bib47","doi-asserted-by":"publisher","first-page":"970","DOI":"10.1038\/ncb886","article-title":"The WRP component of the WAVE-1 complex attenuates Rac-mediated signalling","volume":"4","author":"Soderling","year":"2002","journal-title":"Nature Cell Biology"},{"key":"bib48","doi-asserted-by":"publisher","first-page":"11","DOI":"10.1186\/1471-2121-6-11","article-title":"Inclusion of Scar\/WAVE3 in a similar complex to Scar\/WAVE1 and 2","volume":"6","author":"Stovold","year":"2005","journal-title":"BMC Cell Biology"},{"key":"bib49","doi-asserted-by":"publisher","first-page":"595","DOI":"10.1126\/science.aad9964","article-title":"Phase separation of signaling molecules promotes T cell receptor signal transduction","volume":"352","author":"Su","year":"2016","journal-title":"Science"},{"key":"bib50","doi-asserted-by":"publisher","first-page":"37","DOI":"10.1038\/nrm2069","article-title":"The WASP-WAVE protein network: connecting the membrane to the cytoskeleton","volume":"8","author":"Takenawa","year":"2007","journal-title":"Nature Reviews Molecular Cell Biology"},{"key":"bib51","doi-asserted-by":"publisher","first-page":"38","DOI":"10.1016\/j.jsb.2006.05.009","article-title":"EMAN2: an extensible image processing suite for electron microscopy","volume":"157","author":"Tang","year":"2007","journal-title":"Journal of Structural Biology"},{"key":"bib52","doi-asserted-by":"publisher","first-page":"1215","DOI":"10.1016\/S1097-2765(03)00139-4","article-title":"Contingent phosphorylation\/dephosphorylation provides a mechanism of molecular memory in WASP","volume":"11","author":"Torres","year":"2003","journal-title":"Molecular Cell"},{"key":"bib53","doi-asserted-by":"publisher","first-page":"3513","DOI":"10.1074\/jbc.M509416200","article-title":"Protein-tyrosine kinase and GTPase signals cooperate to phosphorylate and activate Wiskott-Aldrich syndrome protein (WASP)\/neuronal WASP","volume":"281","author":"Torres","year":"2006","journal-title":"Journal of Biological Chemistry"},{"key":"bib54","doi-asserted-by":"publisher","first-page":"1845","DOI":"10.1373\/clinchem.2005.052381","article-title":"Multiplex human papillomavirus serology based on in situ-purified glutathione s-transferase fusion proteins","volume":"51","author":"Waterboer","year":"2005","journal-title":"Clinical Chemistry"},{"key":"bib55","doi-asserted-by":"publisher","first-page":"166","DOI":"10.3389\/fpls.2013.00166","article-title":"ARP2\/3-dependent growth in the plant kingdom: SCARs for life","volume":"4","author":"Yanagisawa","year":"2013","journal-title":"Frontiers in Plant Science"},{"key":"bib56","doi-asserted-by":"publisher","first-page":"237","DOI":"10.1016\/j.str.2011.12.007","article-title":"Iterative stable alignment and clustering of 2D transmission electron microscope images","volume":"20","author":"Yang","year":"2012","journal-title":"Structure"},{"key":"bib57","doi-asserted-by":"publisher","first-page":"2309","DOI":"10.1016\/j.bpj.2011.03.004","article-title":"On the distribution of protein refractive index increments","volume":"100","author":"Zhao","year":"2011","journal-title":"Biophysical Journal"}],"container-title":["eLife"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/cdn.elifesciences.org\/articles\/29795\/elife-29795-v1.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/cdn.elifesciences.org\/articles\/29795\/elife-29795-v1.xml","content-type":"application\/xml","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/elifesciences.org\/articles\/29795","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,10,11]],"date-time":"2023-10-11T20:59:37Z","timestamp":1697057977000},"score":1,"resource":{"primary":{"URL":"https:\/\/elifesciences.org\/articles\/29795"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2017,9,26]]},"references-count":57,"alternative-id":["10.7554\/eLife.29795"],"URL":"https:\/\/doi.org\/10.7554\/elife.29795","archive":["CLOCKSS"],"relation":{"has-review":[{"id-type":"doi","id":"10.3410\/f.731312248.793538305","asserted-by":"object"},{"id-type":"doi","id":"10.3410\/f.731312248.793567193","asserted-by":"object"},{"id-type":"doi","id":"10.7554\/eLife.29795.022","asserted-by":"object"},{"id-type":"doi","id":"10.7554\/eLife.29795.023","asserted-by":"object"}]},"ISSN":["2050-084X"],"issn-type":[{"value":"2050-084X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2017,9,26]]},"article-number":"e29795"}}