Molecular basis for the dual function of Eps8 on actin dynamics: Bundling and capping

Maud Hertzog, Francesca Milanesi, Larnele Hazelwood, Andrea Disanza, Hongjun Liu, Emilie Perlade, Maria Grazia Malabarba, Sebastiano Pasqualato, Alessio Maiolica, Stefano Confalonieri, Christophe Le Clainche, Nina Offenhauser, Jennifer Block, Klemens Rottner, Pier Paolo Di Fiore, Marie France Carlier, Niels Volkmann, Dorit Hanein, Giorgio Scita

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Abstract

Actin capping and cross-linking proteins regulate the dynamics and architectures of different cellular protrusions. Eps8 is the founding member of a unique family of capping proteins capable of side-binding and bundling actin filaments. However, the structural basis through which Eps8 exerts these functions remains elusive. Here, we combined biochemical, molecular, and genetic approaches with electron microscopy and image analysis to dissect the molecular mechanism responsible for the distinct activities of Eps8. We propose that bundling activity of Eps8 is mainly mediated by a compact four helix bundle, which is contacting three actin subunits along the filament. The capping activity is mainly mediated by a amphipathic helix that binds within the hydrophobic pocket at the barbed ends of actin blocking further addition of actin monomers. Single-point mutagenesis validated these modes of binding, permitting us to dissect Eps8 capping from bundling activity in vitro. We further showed that the capping and bundling activities of Eps8 can be fully dissected in vivo, demonstrating the physiological relevance of the identified Eps8 structural/functional modules. Eps8 controls actin-based motility through its capping activity, while, as a bundler, is essential for proper intestinal morphogenesis of developing Caenorhabditis elegans.

Original languageEnglish
JournalPLoS Biology
Volume8
Issue number6
DOIs
Publication statusPublished - Jun 2010

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ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)
  • Neuroscience(all)

Cite this

Hertzog, M., Milanesi, F., Hazelwood, L., Disanza, A., Liu, H., Perlade, E., Malabarba, M. G., Pasqualato, S., Maiolica, A., Confalonieri, S., Le Clainche, C., Offenhauser, N., Block, J., Rottner, K., Fiore, P. P. D., Carlier, M. F., Volkmann, N., Hanein, D., & Scita, G. (2010). Molecular basis for the dual function of Eps8 on actin dynamics: Bundling and capping. PLoS Biology, 8(6). https://doi.org/10.1371/journal.pbio.1000387