ROLE OF NUCLEOTIDE EXCHANGE AND HYDROLYSIS IN THE FUNCTION OF PROFILIN IN ACTIN ASSEMBLY

Profilin, an essential G-actin-binding protein, has two opposite regul atory functions in actin filament assembly. It facilitates assembly at the barbed ends by lowering the critical concentration (Pantaloni, D. , and Carlier, M.-F. (1993) Cell 75, 1007-1014); in contrast it contri butes to the pool of unassembled actin when barbed ends are capped. We proposed that the first of these functions required an input of energ y. How profilin uses the ATP hydrolysis that accompanies actin polymer ization and whether the acceleration of nucleotide exchange on G-actin by profilin participates in its function in filament assembly are the issues addressed here. We show that 1) profilin increases the treadmi lling rate of actin filaments in the presence of Mg2+ ions; 2) when fi laments are assembled from CaATP-actin, which polymerizes in a quasire versible fashion, profilin does not promote assembly at the barbed end s and has only a G-actin-sequestering function; 3) plant profilins do not accelerate nucleotide exchange on G-actin, yet they promote assemb ly at the barbed end. The enhancement of nucleotide exchange by profil in is therefore not involved in its promotion of actin assembly, and t he productive growth of filaments from profilin-actin complex requires the coupling of ATP hydrolysis to profilin-actin assembly, a conditio n fulfilled by Mg-actin, and not by Ca-actin.