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Autophagy machinery mediates macroendocytic processing and entotic cell death by targeting single membranes

Nature Cell Biology volume 13, pages 1335–1343 (2011)Cite this article

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Abstract

Autophagy normally involves the formation of double-membrane autophagosomes that mediate bulk cytoplasmic and organelle degradation. Here we report the modification of single-membrane vacuoles in cells by autophagy proteins. LC3 (Light chain 3) a component of autophagosomes, is recruited to single-membrane entotic vacuoles, macropinosomes and phagosomes harbouring apoptotic cells, in a manner dependent on the lipidation machinery including ATG5 and ATG7, and the class III phosphatidylinositol-3-kinase VPS34. These downstream components of the autophagy machinery, but not the upstream mammalian Tor (mTor)-regulated ULK–ATG13–FIP200 complex, facilitate lysosome fusion to single membranes and the degradation of internalized cargo. For entosis, a live-cell-engulfment program, the autophagy-protein-dependent fusion of lysosomes to vacuolar membranes leads to the death of internalized cells. As pathogen-containing phagosomes can be targeted in a similar manner, the death of epithelial cells by this mechanism mimics pathogen destruction. These data demonstrate that proteins of the autophagy pathway can target single-membrane vacuoles in cells in the absence of pathogenic organisms.

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Figure 1: Entotic cell death involves recruitment of LC3 to a single-membrane vacuole.
Figure 2: Autophagy machinery in host cells controls the fate of starving internalized cells.
Figure 3: Inhibition of apoptosis and entotic cell death promotes anchorage-independent growth.
Figure 4: LC3 recruits to single-membrane vacuoles containing apoptotic cells to facilitate corpse degradation.
Figure 5: LC3 is recruited to macropinosomes independently of macroautophagy.
Figure 6: LGG-1 recruitment to apoptotic phagosomes during C. elegans embryonic development.

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Acknowledgements

We thank J. Durgan, I. Ganley, X. Jiang, G. Mouneimne, E. Yao, A. Spencer and members of the Overholtzer laboratory for helpful discussions, reagents and for reading the manuscript. We also thank N. Lampen of the Memorial Sloan Kettering Cancer Center Electron Microscopy Facility for processing of electron microscopy samples. This work was financially supported by a grant from the National Cancer Institute (CA154649; M.O.), The Geoffrey Beene Cancer Research Center at MSKCC (M.O.), the Louis V. Gerstner, Jr. Young Investigators Fund (M.O. and C.M.H.) and the Alfred W. Bressler Scholar Fund (C.M.H.).

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Authors and Affiliations

  1. Cell Biology Program, Memorial Sloan Kettering Cancer Center, New York, New York 10065, USA

    Oliver Florey, Sung Eun Kim, Cole M. Haynes & Michael Overholtzer

  2. BCMB Allied Program, Weill Cornell Medical College, 1300 York Avenue, New York, New York 10065, USA

    Sung Eun Kim, Cole M. Haynes & Michael Overholtzer

  3. Department of Physiology, University of Arizona, Tucson, Arizona 85721, USA

    Cynthia P. Sandoval

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Contributions

O.F. and M.O. designed, carried out experiments and wrote the paper. S.E.K. and C.P.S. contributed experimental assistance and data. C.M.H. provided worm strains and carried out RNAi feeding of C. elegans.

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Correspondence to Michael Overholtzer.

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Florey, O., Kim, S., Sandoval, C. et al. Autophagy machinery mediates macroendocytic processing and entotic cell death by targeting single membranes. Nat Cell Biol 13, 1335–1343 (2011). https://doi.org/10.1038/ncb2363

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