Structure of the F-tractin–F-actin complex

AbstractF-tractin is a short peptide widely used to visualize the actin cytoskeleton in live eukaryotic cells. Similar to other actin-binding probes, F-tractin alters actin organization and impairs cell migration when expressed at high levels. In addition, the probe has been reported to directly induce actin bundling. To elucidate the mechanism behind these effects, we determined the structure of the F-tractin–F-actin complex using electron cryo-microscopy. Our analysis revealed that the F-tractin peptide consists of a flexible N-terminal region and an amphipathic C-terminal helix. The N-terminal part is completely dispensable for F-actin binding but is responsible for the actin bundling effect. The C-terminal helical region interacts with a hydrophobic pocket formed by two neighboring actin subunits, a region identified as an interface for many other actin-binding polypeptides, including Lifeact, the most widely used actin-binding probe. Thus, rather than contrasting F-tractin and Lifeact, our data indicate that these peptides have analogous modes of interaction with F-actin. Our study dissects the structural elements of F-tractin and provides a mechanistic basis for the selection and future development of actin probes.