Vimentin promotes actin assembly by stabilizing ATP-actin subunits at the barbed end

Vimentin intermediate filaments play essential roles in maintaining cell integrity and regulating numerous cellular functions. In particular, vimentin cooperates with the actin cytoskeleton in key cellular processes that rely on actin dynamics, such as such as migration, division, and mechanosensing. While there is evidence that these two cytoskeletal components interact in cells, the underlying molecular mechanisms are only partially understood. Actin and vimentin can interact through biochemical signaling pathways or via cross-linkers, but whether they engage in a direct protein-protein interaction has remained controversial, in part because such interactions are difficult to isolate and characterize in cells. Using in vitro reconstitution and total internal reflection fluorescence microscopy to monitor the elongation of single actin filaments, we show that vimentin promotes actin elongation at the barbed end in a dose-dependent manner. Neither the vimentin tail nor head domains are required for this effect, and both filamentous and non-filamentous vimentin enhance actin elongation. We find that this acceleration results from a stabilization of actin subunits at the barbed end. Strikingly, this effect depends on the nucleotide state of actin, as the acceleration is only observed for the elongation from ATP-actin, and not ADP-actin monomers. This stabilization also enables vimentin to promote the nucleation of actin filaments. Consistently, magnetic pull-down assays demonstrate a direct, albeit weak, interaction between vimentin and actin monomers. Altogether, these findings identify vimentin as an unexpected new actor in the regulation of actin dynamics at the barbed end and bring new insights into the functional role of vimentin through cytoskeletal crosstalk.