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Electron tomography reveals unbranched networks of actin filaments in lamellipodia

Nature Cell Biology volume 12, pages 429–435 (2010)Cite this article

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Abstract

Eukaryotic cells can initiate movement using the forces exerted by polymerizing actin filaments to extend lamellipodial and filopodial protrusions. In the current model, actin filaments in lamellipodia are organized in a branched, dendritic network. We applied electron tomography to vitreously frozen 'live' cells, fixed cells and cytoskeletons, embedded in vitreous ice or in deep-negative stain. In lamellipodia from four cell types, including rapidly migrating fish keratocytes, we found that actin filaments are almost exclusively unbranched. The vast majority of apparent filament junctions proved to be overlapping filaments, rather than branched end-to-side junctions. Analysis of the tomograms revealed that actin filaments terminate at the membrane interface within a zone several hundred nanometres wide at the lamellipodium front, and yielded the first direct measurements of filament densities. Actin filament pairs were also identified as lamellipodium components and bundle precursors. These data provide a new structural basis for understanding actin-driven protrusion during cell migration.

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Figure 1: Cryo-electron tomography of lamellipodia.
Figure 2: Correlated live-cell imaging and electron tomography of protruding goldfish fibroblast lamellipodium.
Figure 3: Actin filaments overlap and do not form a dendritic array in keratocyte lamellipodia.
Figure 4: The keratocyte lamellipodium tapers towards the tip with the filament ends distributed over a wide zone.
Figure 5: Schemes of actin filament organization.
Figure 6: Hypothetical scheme of actin network generation in lamellipodia.

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Acknowledgements

We thank T. Kulcsar for help with graphics, M. Brandstaetter for support in electron microscopy, T. Stradal and K. Rottner for the p16 ArpC5 construct, and R. Tsien for mCherry. J.V.S. was supported by the Austrian Science Research Council and the Vienna Science Research and Technology Fund (WWTF). G.P.R and J.V.S. acknowledge the support by the City of Vienna/Zentrum für Innovation und Technologie through the spot of excellence grant “Center of Molecular and Cellular Nanostructure”.

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  1. Edit Urban and Sonja Jacob: These two authors contributed equally to the work.

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  1. Institute of Molecular Biotechnology, Dr Bohr Gasse 3–7, 1030, Vienna, Austria

    Edit Urban, Sonja Jacob, Maria Nemethova, Guenter P. Resch & J. Victor Small

  2. Research Institute of Molecular Pathology, Dr Bohr Gasse 3–7, 1030, Vienna, Austria

    Guenter P. Resch

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Contributions

E.U. and S.J. prepared experimental material and performed electron tomography, image processing and image analysis. M.N. prepared experimental material and performed correlated live-cell imaging and image analysis. E.U., S.J. and M.N. prepared the figures. G.P.R. set up the tomography facility, provided advice and assistance with tomography, cryofixation and image processing. J.V.S. conceived the project, contributed to image analysis and wrote the manuscript with feedback from co-authors.

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Correspondence to J. Victor Small.

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Urban, E., Jacob, S., Nemethova, M. et al. Electron tomography reveals unbranched networks of actin filaments in lamellipodia. Nat Cell Biol 12, 429–435 (2010). https://doi.org/10.1038/ncb2044

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