A Corrigendum to this article was published on 01 October 2010
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Abstract
Cell-free reconstitution of membrane traffic reactions and the morphological characterization of membrane intermediates that accumulate under these conditions have helped to elucidate the physical and molecular mechanisms involved in membrane transport1,2,3. To gain a better understanding of endocytosis, we have reconstituted vesicle budding and fission from isolated plasma membrane sheets and imaged these events. Electron and fluorescence microscopy, including subdiffraction-limit imaging by stochastic optical reconstruction microscopy (STORM)4,5,6, revealed F-BAR (FBP17) domain coated tubules nucleated by clathrin-coated buds when fission was blocked by GTPγS. Triggering fission by replacing GTPγS with GTP led not only to separation of clathrin-coated buds, but also to vesicle formation by fragmentation of the tubules. These results suggest a functional link between FBP17-dependent membrane tubulation and clathrin-dependent budding. They also show that clathrin spatially directs plasma membrane invaginations that lead to the generation of endocytic vesicles larger than those enclosed by the coat.
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Clathrin mediates membrane fission and budding by constricting membrane pores
Membrane transformations of fusion and budding
Molecular mechanics underlying flat-to-round membrane budding in live secretory cells
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31 August 2010
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Acknowledgements
We thank Aurelien Roux and Yuxin Mao for advice and help; Lijun Liu, Frank Wilson and Christoph Rahner for critical technical assistance. This work was supported in part by the G. Harold and Leila Y. Mathers Charitable Foundation, the W.M. Keck Foundation and the National Institutes of Health grants (NS36251 and DK45735) to P.D.C., and National Institutes of Health grant (GM068518) to X.Z.; P.D.C. and X.Z. are Howard Hughes Medical Institute Investigators.
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Bo Huang
Present address: Current address: Department of Pharmaceutical Chemistry, Department of Biochemistry and Biophysics, University of California, San Francisco, CA 94158, USA.,
Authors and Affiliations
Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, 06520, CT, USA
Min Wu, Andrea Raimondi & Pietro De Camilli
Department of Cell Biology, Yale University School of Medicine, New Haven, 06520, CT, USA
Min Wu, Morven Graham, Andrea Raimondi & Pietro De Camilli
Department of Neurobiology, Yale University School of Medicine, New Haven, 06520, CT, USA
Pietro De Camilli
Program in Cellular Neuroscience, Neurodegeneration and Repair, Yale University School of Medicine, New Haven, 06520, CT, USA
Min Wu, Andrea Raimondi & Pietro De Camilli
Department of Chemistry and Chemical Biology, Harvard University, Cambridge, 02138, MA, USA
Bo Huang & Xiaowei Zhuang
Department of Physics, Harvard University, Cambridge, 02138, MA, USA
Xiaowei Zhuang
Department of Cell Biology and Physiology, Washington University School of Medicine, 660 South Euclid Avenue, St. Louis, 63110, MO, USA
John E. Heuser
Howard Hughes Medical Institute, Harvard University, Cambridge, 02138, MA, USA
Bo Huang & Xiaowei Zhuang
- Min Wu
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- Bo Huang
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Contributions
M.W. and P.D.C. designed the experiments and wrote the manuscript; M.W. performed experiments. Experimental work was also contributed by B.H. (STORM), J.E.H. (electron microscopy), A.R. (electron microscopy) and M.G. (electron microscopy). B.H. and X.Z. also contributed to discussion and manuscript preparation.
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Correspondence to Pietro De Camilli.
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Wu, M., Huang, B., Graham, M. et al. Coupling between clathrin-dependent endocytic budding and F-BAR-dependent tubulation in a cell-free system. Nat Cell Biol 12, 902–908 (2010). https://doi.org/10.1038/ncb2094
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DOI: https://doi.org/10.1038/ncb2094
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