title: Direct demonstration of actin filament annealing in vitro.
abstract: Direct electron microscopic examination confirms that short actin filaments rapidly anneal end-to-end in vitro, leading over time to an increase in filament length at steady state. During annealing of mixtures of native unlabeled filaments and glutaraldehyde-fixed filaments labeled with myosin subfragment-1, the structural polarity within heteropolymers is conserved absolutely. Annealing does not appear to require either ATP hydrolysis or the presence of exogenous actin monomers, suggesting that joining occurs through the direct association of filament ends. During recovery from sonication the initial rate of annealing is consistent with a second-order reaction involving the collision of two filament ends with an apparent annealing rate constant of 10(7) M-1s-1. This rapid phase lasts less than 10 s and is followed by a slow phase lasting minutes to hours. Annealing is calculated to contribute minimally to filament elongation during the initial stages of self-assembly. However, the rapid rate of annealing of sonicated fixed filaments observed in vitro suggests that it may be an efficient mechanism for repairing breaks in filaments and that annealing together with polymer-severing mechanisms may contribute significantly to the dynamics and function of actin filaments in vivo.
authors.affiliation: Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
authors.forename: D B
authors.initials: DB
authors.lastname: Murphy
authors.forename: R O
authors.initials: RO
authors.lastname: Gray
authors.forename: W A
authors.initials: WA
authors.lastname: Grasser
authors.forename: T D
authors.initials: TD
authors.lastname: Pollard
mesh_terms: D008841:Actin Cytoskeleton / Q000648:ultrastructure*; D000199:Actins / Q000502:physiology*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D003599:Cytoskeleton / Q000648:ultrastructure*; D066298:In Vitro Techniques; D008854:Microscopy, Electron; D011485:Protein Binding; D011817:Rabbits; D013010:Sonication; D013997:Time Factors
chemical_list: D000199:Actins; D000255:Adenosine Triphosphate