Rapid formation and high diffusibility of actin-cofilin cofilaments at lowpH

Cofilin is a small actin-binding protein that is known to bind both F-actin and G-actin, severing the former. The interaction of cofilin with actin is pH-sensitive, F-actin being preferentially bound at low pH and G-actin at higher pH, within the physiological range. Diffusion coefficients of F-acti n with cofilin were measured by the fluorescence recovery after photobleach ing (FRAP) technique. This has the potential for simultaneous and direct me asurement of average polymer length via the average diffusion coefficient o f the polymers (D-LM) as well as the fraction of polymerized actin, f(LM), present in solution. In the range of cofilin-actin ratios up to 1 : 1 and a t both pH 6.5 and pH 8.0, the diffusion coefficients of the polymers increa sed with the amount of cofilin present in the complex, in a co-operative ma nner to a plateau. We interpret this as indicating co-operative binding/sev ering and that filaments less than a certain length cannot be severed furth er. Under the conditions used here, filaments were found to be more motile at pH 6.5 than at pH 8.0. At pH 8.0, some actin is expected to be sequester ed as ADP-actin-cofilin complexes, with the remaining actin being present a s long slowly diffusing filaments. At pH 6.5, however, cofilin binds to F-a ctin to form short rapidly diffusing cofilaments. These filaments form very rapidly from cofilin-actin monomeric complexes, possibly indicating that t his complex is able to polymerize without dissociation. These findings may be relevant to the nuclear import of actin-cofilin complexes.