Integrating the actin and vimentin cytoskeletons: Adhesion-dependent formation of fimbrin-vimentin complexes in macrophages

Cells adhere to the substratum through specialized structures that are link ed to the actin cytoskeleton. Recent studies report that adhesion also invo lves the intermediate filament (IF) and microtubule cytoskeletons, although their mechanisms of interaction are unknown. Here we report evidence for a novel adhesion-dependent interaction between components of the actin and I F cytoskeletons. In biochemical fractionation experiments, fimbrin and vime ntin coprecipitate from detergent extracts of macrophages using vimentin- o r fimbrin-specific antisera. Fluorescence microscopy confirms the biochemic al association. Both proteins colocalized to podosomes in the earliest stag es of cell adhesion and spreading. The complex is also found in filopodia a nd retraction fibers. After detergent extraction, fimbrin and vimentin stai ning of podosomes, filopodia, and retraction fibers are lost, confirming th at the complex is localized to these structures. A 1:4 stoichiometry of fim brin binding to vimentin and a low percentage (1%) of the extracted vimenti n suggest that fimbrin interacts with a vimentin subunit. A fimbrin-binding site was identified in the NH2-terminal domain of vimentin and the vimenti n binding site at residues 143-188 in the CH1 domain of fimbrin. Based on t hese observations, we propose that a fimbrin-vimentin complex may be involv ed in directing the assembly of the vimentin cytoskeleton at cell adhesion sites.