Summary
Formins are actin polymerization factors that are known to nucleate and elongate actin filaments at the barbed end. In the present study we show that human FHOD1 lacks actin nucleation and elongation capacity, but acts as an actin bundling factor with capping activity toward the filament barbed end. Constitutively active FHOD1 associates with actin filaments in filopodia and lamellipodia at the leading edge, where it moves with the actin retrograde flow. At the base of lamellipodia, FHOD1 is enriched in nascent, bundled actin arcs as well as in more mature stress fibers. This function requires actin-binding domains located N-terminally to the canonical FH1–FH2 element. The bundling phenotype is maintained in the presence of tropomyosin, confirmed by electron microscopy showing assembly of 5 to 10 actin filaments into parallel, closely spaced filament bundles. Taken together, our data suggest a model in which FHOD1 stabilizes actin filaments by protecting barbed ends from depolymerization with its dimeric FH2 domain, whereas the region N-terminal to the FH1 domain mediates F-actin bundling by simultaneously binding to the sides of adjacent F-actin filaments.
Footnotes
↵‡ Present address: University of California San Francisco, Department of Cell and Tissue Biology, San Francisco, CA 94143, USA
↵§ Present address: Institute of Medical Physics and Biophysics, Charité, Universitätsmedizin Berlin, 10117 Berlin, Germany
↵¶ Present address: Department of Cell Pathology, Faculty of Biotechnology, University of Wroclaw, PL-51-148 Wroclaw, Poland
Author contributions
A.S. and M.G. designed the study. A.S. produced proteins and, with the help of S.K., performed and analyzed in vitro actin assembly experiments. H.G.M. and A.J.M. performed cell transfection experiments. H.G.M., U.S. and C.A.S. did in vitro actin polymerization TIRF microscopy. E.B., H.G.M. and S.R. performed electron microscopy measurements. L.D. performed and analyzed live cell imaging. O.T.F. contributed with reagents and discussions. A.S. and M.G. wrote the manuscript with the help of all other authors. All authors discussed the results and commented on the manuscript.
Funding
This work was supported in part by the Deutsche Forschungsgemeinschaft [grant numbers GE-976/4 to M.G., FA-378/6 to O.T.F., MA-807/19 to H.G.M., and RA-1781/1 to S.R.]; and by the Fonds der Chemischen Industrie (stipend to E.B.).
Supplementary material available online at http://jcs.biologists.org/lookup/suppl/doi:10.1242/jcs.126706/-/DC1
- Accepted January 30, 2013.
- © 2013. Published by The Company of Biologists Ltd
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