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Actin dynamics counteract membrane tension during clathrin-mediated endocytosis

Nature Cell Biology volume 13, pages 1124–1131 (2011)Cite this article

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Abstract

Clathrin-mediated endocytosis is independent of actin dynamics in many circumstances but requires actin polymerization in others. We show that membrane tension determines the actin dependence of clathrin-coat assembly. As found previously, clathrin assembly supports formation of mature coated pits in the absence of actin polymerization on both dorsal and ventral surfaces of non-polarized mammalian cells, and also on basolateral surfaces of polarized cells. Actin engagement is necessary, however, to complete membrane deformation into a coated pit on apical surfaces of polarized cells and, more generally, on the surface of any cell in which the plasma membrane is under tension from osmotic swelling or mechanical stretching. We use these observations to alter actin dependence experimentally and show that resistance of the membrane to propagation of the clathrin lattice determines the distinction between ‘actin dependent and ‘actin independent’. We also find that light-chain-bound Hip1R mediates actin engagement. These data thus provide a unifying explanation for the role of actin dynamics in coated-pit budding.

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Figure 1: Formation of endocytic coated pits and vesicles at the apical and basolateral surfaces of polarized MDCK cells.
Figure 2: Disruption of apical coat formation by pharmacological interference with the small GTPases Rac1 and Arf6 and interference with the function of clathrin light chains.
Figure 3: Actin dependence for endocytic coat formation in cells swelled by hypo-osmotic treatment.
Figure 4: Actin dependence of endocytic coat formation in mechanically stretched cells.
Figure 5: Model depicting the role of actin polymerization during the formation of endocytic clathrin-coated pits.

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Acknowledgements

This work was supported by NIH grant National Institutes of Health GM 075252 (T.K.) and a Harvard Digestive Disease Consortium Feasibility Award (S.B.). We express gratitude to E. Marino (supported by NIH grant U54 AI057159, New England Regional Center of Excellence in Biodefense and Emerging Infectious Diseases) for maintaining the imaging resource used in this study and to S.C. Harrison for help with describing the force contributed by membrane tension and for editorial assistance. We gratefully acknowledge M. Ericsson and I. Kim for preparation of samples for electron microscopic analysis and members of the Kirchhausen laboratory for thought-provoking discussions. We thank M. Arpin (Centre National de la Recherche Scientifique (CNRS)/Morphogenèse et Signalisation cellulaires, Paris, France) and C. Revenu (Centre National de la Recherche Scientifique/Institut Curie, Paris, France) for the antibody specific for villin-1. We also acknowledge S. Robine for important suggestions involving the villin-1/villin-2 depletion experiments.

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  1. Steeve Boulant and Comert Kural: These authors contributed equally to this work

Authors and Affiliations

  1. Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, USA

    Steeve Boulant, Comert Kural, Jean-Christophe Zeeh & Tomas Kirchhausen

  2. Immune Disease Institute and Program in Cellular and Molecular Medicine at Children’s Hospital, Boston, Massachusetts 02115, USA

    Steeve Boulant, Comert Kural, Jean-Christophe Zeeh & Tomas Kirchhausen

  3. Laboratoire de Morphogénèse et Signalisation Cellulaires, Institut Curie Paris, Paris, Cedex 05, France

    Florent Ubelmann

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Contributions

S.B., C.K. and T.K. designed experiments; S.B. and C.K. carried out experiments; S.B. and C.K. analysed data; C.K. developed the analytical tools to analyse the 3D data. F.U. provided expression vectors specific for the experiments with villin-1/villin-2, contributed to the experimental design of the experiments involved in villin-1/villin-2 depletion and created the LLCPK1 cell line. S.B. and T.K. wrote the manuscript. All authors discussed the results and contributed to the final manuscript.

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Correspondence to Tomas Kirchhausen.

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Boulant, S., Kural, C., Zeeh, JC. et al. Actin dynamics counteract membrane tension during clathrin-mediated endocytosis. Nat Cell Biol 13, 1124–1131 (2011). https://doi.org/10.1038/ncb2307

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