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Imaging actin and dynamin recruitment during invagination of single clathrin-coated pits

Nature Cell Biology volume 4, pages 691–698 (2002)Cite this article

Abstract

As a final step in endocytosis, clathrin-coated pits must separate from the plasma membrane and move into the cytosol as a coated vesicle. Because these events involve minute movements that conventional light microscopy cannot resolve, they have not been observed directly and their dynamics remain unexplored. Here, we used evanescent field (EF) microscopy to observe single clathrin-coated pits or vesicles as they draw inwards from the plasma membrane and finally lose their coats. This inward movement occurred immediately after a brief burst of dynamin recruitment and was accompanied by transient actin assembly. Therefore, dynamin may provide the trigger and actin may provide the force for movement into the cytosol.

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Figure 1: DsRed-labelled clathrin-coated structures mediate endocytosis.
Figure 2: Three ways to image live 3T3 cells.
Figure 3: Internalization of a clathrin-coated pit is accompanied by recruitment of actin.
Figure 4: Actin recruitment and other events during internalization of coated pits.
Figure 5: Recruitment of dynamin.
Figure 6: Dynamin and actin are recruited sequentially.
Figure 7: Fluorescence changes and endocytic structures.

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Acknowledgements

We thank P. de Camilli and S. Schmid for helpful discussions, and D. Zenisek, J. Taraska and T. Blackmer for their helpful comments on the manuscript. This work was supported by National Institutes of Health grants MH60600 and DK44239.

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  1. Vollum Institute L-474, Oregon Health and Sciences University, 3181 SW Sam Jackson Park Rd., Portland, 97201, OR, USA

    Christien J. Merrifield, Morris E. Feldman, Lei Wan & Wolfhard Almers

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Correspondence to Wolfhard Almers.

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Merrifield, C., Feldman, M., Wan, L. et al. Imaging actin and dynamin recruitment during invagination of single clathrin-coated pits. Nat Cell Biol 4, 691–698 (2002). https://doi.org/10.1038/ncb837

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