Reconstitution and regulation of actin gel-sol transformation with purified filamin and villin.
Gel-sol transformation of actin filaments, a process essential for cell motility, can be reconstituted in vitro and regulated in a predictable fashion by the combined action of villin and filamin. Measurements made in a low shear falling ball viscometer show that mixtures of actin, villin, and filamin exist either as a gel (yield point greater than or equal to 140 dynes/cm2) or as a low viscosity liquid depending on the relative ration of villin:actin. Filamin induces gelation of F-actin by forming stable cross-links between actin filaments. Villin inhibits filamin-induced F-actin gelation, but the effect can be overcome by increasing the amount of filamin. Sedimentation assays show that villin does not inhibit gelation of actin by preventing filamin from binding to F-actin. Results from viscosity measurements and filament length determinations show that villin increases actin filament number by reducing the average filament length without altering the total amount of polymer. Because the gel point of a fixed amount of polymer is sharply dependent on the ratio of cross-links to number of polymers, the solation effect of villin might be explained by its effect on filament number. Based on the network theory of gel formation, calculations of the amount of additional cross-linker required to overcome the effect of a known increase in the number of actin filaments agree reasonably well with experimental findings. These results document the existence of cellular proteins which could regulate gel-sol transformation in vivo by their effect on actin polymer length and, therefore, on actin filament number.
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002645:Chickens; D003285:Contractile Proteins / Q000378:metabolism*; D064448:Filamins; D005782:Gels; D007413:Intestinal Mucosa; D007700:Kinetics; D046911:Macromolecular Substances; D008840:Microfilament Proteins; D008854:Microscopy, Electron; D008871:Microvilli; D008970:Molecular Weight; D009130:Muscle, Smooth; D009132:Muscles / Q000378:metabolism; D011817:Rabbits; D012995:Solubility; D014783:Viscosity
D000199:Actins; D002352:Carrier Proteins; D003285:Contractile Proteins; D064448:Filamins; D005782:Gels; D046911:Macromolecular Substances; D008840:Microfilament Proteins; C026615:villin