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External push and internal pull forces recruit curvature-sensing N-BAR domain proteins to the plasma membrane

Nature Cell Biology volume 14, pages 874–881 (2012)Cite this article

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Abstract

Many of the more than 20 mammalian proteins with N-BAR domains1,2 control cell architecture3 and endocytosis4,5 by associating with curved sections of the plasma membrane6. It is not well understood whether N-BAR proteins are recruited directly by processes that mechanically curve the plasma membrane or indirectly by plasma-membrane-associated adaptor proteins that recruit proteins with N-BAR domains that then induce membrane curvature. Here, we show that externally induced inward deformation of the plasma membrane by cone-shaped nanostructures (nanocones) and internally induced inward deformation by contracting actin cables both trigger recruitment of isolated N-BAR domains to the curved plasma membrane. Markedly, live-cell imaging in adherent cells showed selective recruitment of full-length N-BAR proteins and isolated N-BAR domains to plasma membrane sub-regions above nanocone stripes. Electron microscopy confirmed that N-BAR domains are recruited to local membrane sites curved by nanocones. We further showed that N-BAR domains are periodically recruited to curved plasma membrane sites during local lamellipodia retraction in the front of migrating cells. Recruitment required myosin-II-generated force applied to plasma-membrane-connected actin cables. Together, our results show that N-BAR domains can be directly recruited to the plasma membrane by external push or internal pull forces that locally curve the plasma membrane.

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Figure 1: Nanocones induce inward curved plasma membrane deformations at the basal plasma membrane of adherent cells.
Figure 2: Nanocone-induced membrane deformation triggers N-BAR domain recruitment to the plasma membrane.
Figure 3: N-BAR domains are dynamically recruited to local membrane sites at the leading edge of migrating 3T3 cells.
Figure 4: Myosin-II-dependent contraction of actin cables induces N-BAR domain recruitment to the lamellipodia plasma membrane.
Figure 5: External push and internal pull forces applied to local plasma membrane sites recruit N-BAR domains to inward curved membrane.

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Acknowledgements

The authors thank the members of the Meyer laboratory for comments and discussion. M.G. was supported by the Swiss National Science Foundation (No. PBBSP3-123159), and a Novartis Jubilaeumsstiftung and Stanford Deans Postdoctoral Fellowship. S.J. was supported by a Korea Foundation for Advanced Studies graduate fellowship. Y.C. acknowledges the partial support from a DOE-EFRC at Stanford: Center on Nanostructuring for Efficient Energy Conversion (No. DE-SC0001060). T.M. acknowledges financial support from the National Institutes of Health, MH064801, MH095087 and GM063702.

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Author notes

  1. Yi I. Wu

    Present address: Present address: Center for Cell Analysis and Modeling, University of Conneticut Health Center, 400 Farmington Avenue, Cell and Genome Sciences Building, Farmington, Connecticut 06032, USA,

Authors and Affiliations

  1. Department of Chemical and Systems Biology, Stanford University, 318 Campus Drive, Stanford, California 94305, Clark Building W200, USA

    Milos Galic, Feng-Chiao Tsai & Tobias Meyer

  2. Department of Electrical Engineering, Stanford University, 476 Lomita Mall, Stanford, California 94305, McCullough Building 217, USA

    Sangmoo Jeong

  3. Cell Sciences Imaging Facility, Stanford University School of Medicine, Stanford, California 94305, USA

    Lydia-Marie Joubert

  4. Department of Pharmacology, Lineberger Comprehensive Cancer Center, University of North Carolina, 120 Mason Farm Road, Genetic Medicine Building, Chapel Hill, North Carolina 27599, USA

    Yi I. Wu & Klaus M. Hahn

  5. Department of Materials Science and Engineering, Stanford University, 476 Lomita Mall, Stanford, California 94305, McCullough Building 343, USA

    Yi Cui

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Contributions

M.G. performed all experiments and analysed the data. F-C.T. developed the temporal cross-correlation analysis. S.J. and Y.C. designed the nanocones. L-M.J. helped with the SEM. Y.I.W. and K.M.H. developed the photoactivatable RAC construct. M.G. and T.M. designed the experiments, interpreted the results and wrote the manuscript. All authors discussed the results and the manuscript. T.M. supervised the study.

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Correspondence to Milos Galic or Tobias Meyer.

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Galic, M., Jeong, S., Tsai, FC. et al. External push and internal pull forces recruit curvature-sensing N-BAR domain proteins to the plasma membrane. Nat Cell Biol 14, 874–881 (2012). https://doi.org/10.1038/ncb2533

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