Actin Filament Attachments for Sustained Motility In Vitro Are Maintained by Filament Bundling

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dc.contributor.authorHu, Xiaohuaen
dc.contributor.authorKuhn, Jeffrey R.en
dc.contributor.departmentBiological Sciencesen
dc.date.accessioned2018-10-31T12:46:11Zen
dc.date.available2018-10-31T12:46:11Zen
dc.date.issued2012-02-16en
dc.description.abstractWe reconstructed cellular motility in vitro from individual proteins to investigate how actin filaments are organized at the leading edge. Using total internal reflection fluorescence microscopy of actin filaments, we tested how profilin, Arp2/3, and capping protein (CP) function together to propel thin glass nanofibers or beads coated with N-WASP WCA domains. Thin nanofibers produced wide comet tails that showed more structural variation in actin filament organization than did bead substrates. During sustained motility, physiological concentrations of Mg2+ generated actin filament bundles that processively attached to the nanofiber. Reduction of total Mg2+ abolished particle motility and actin attachment to the particle surface without affecting actin polymerization, Arp2/3 nucleation, or filament capping. Analysis of similar motility of microspheres showed that loss of filament bundling did not affect actin shell formation or symmetry breaking but eliminated sustained attachments between the comet tail and the particle surface. Addition of Mg2+, Lys-Lys2+, or fascin restored both comet tail attachment and sustained particle motility in low Mg2+ buffers. TIRF microscopic analysis of filaments captured by WCA-coated beads in the absence of Arp2/3, profilin, and CP showed that filament bundling by polycation or fascin addition increased barbed end capture by WCA domains. We propose a model in which CP directs barbed ends toward the leading edge and polycation-induced filament bundling sustains processive barbed end attachment to the leading edge.en
dc.description.versionPublished versionen
dc.format.mimetypeapplication/pdfen
dc.identifier.doihttps://doi.org/10.1371/journal.pone.0031385en
dc.identifier.eissn1932-6203en
dc.identifier.issue2en
dc.identifier.othere31385en
dc.identifier.pmid22359589en
dc.identifier.urihttp://hdl.handle.net/10919/85591en
dc.identifier.volume7en
dc.language.isoenen
dc.publisherPLOSen
dc.rightsCreative Commons Attribution 4.0 Internationalen
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/en
dc.titleActin Filament Attachments for Sustained Motility In Vitro Are Maintained by Filament Bundlingen
dc.title.serialPLOS ONEen
dc.typeArticle - Refereeden
dc.type.dcmitypeTexten

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