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A pathway for phagosome maturation during engulfment of apoptotic cells

Nature Cell Biology volume 10, pages 556–566 (2008)Cite this article

Abstract

Removal of apoptotic cells is critical for the physiological well-being of the organism1 and defects in corpse removal have been linked to disease states2,3. Genes regulating corpse recognition and internalization have been identified4, but few molecules involved in the processing of internalized corpses are known. Through a combination of targeted and unbiased reverse genetic screens in Caenorhabditis elegans, and studies in mammalian cells, we have identified genes required for maturation of apoptotic-cell-containing phagosomes. We have further ordered these candidates, which include the GTPases RAB-5 and RAB-7 and the HOPS complex, into a coherent linear pathway for the maturation of apoptotic cells within phagosomes. In depth analysis of two additional candidate genes, the phosphatidylinositol 3 kinase (PI(3)K) vps-34 (A001762) and dyn-1/dynamin, showed an accumulation of internalized, but undegraded, corpses within abnormal Rab5-negative phagosomes. We ordered these candidates in our pathway, with DYN-1 functioning upstream of VPS-34 in the recruitment and/or retention of RAB-5 to the phagosome. Finally, we have also identified a previously undescribed biochemical complex containing Vps34, dynamin and Rab5GDP, thus providing a mechanism for Rab5 recruitment to the nascent phagosome.

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Figure 1: RAB-5, RAB-7 and the HOPS complex are required for maturation of apoptotic-cell-containing phagosomes.
Figure 2: A reverse genetic screen identifies DYN-1 and VPS-34, which are required for efficient corpse removal.
Figure 3: DYN-1 colocalizes with RAB-5 and is required for RAB-5 recruitment to phagosomes containing apoptotic cells.
Figure 4: DYN-1 is required for efficient recruitment of RAB-5 and 2 × FYVE domain to phagosomes containing engulfed apoptotic cells in the C.elegans gonad.
Figure 5: Role of DYN-1 in phagosome maturation is evolutionarily conserved.

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References

  1. Ravichandran, K. S. & Lorenz, U. Engulfment of apoptotic cells: signals for a good meal. Nature Rev. Immunol. 7, 964–974 (2007).

    Article  CAS  Google Scholar 

  2. Kawane, K. et al. Chronic polyarthritis caused by mammalian DNA that escapes from degradation in macrophages. Nature 443, 998–1002 (2006).

    Article  CAS  Google Scholar 

  3. Schrijvers, D. M., De Meyer, G. R., Kockx, M. M., Herman, A. G. & Martinet, W. Phagocytosis of apoptotic cells by macrophages is impaired in atherosclerosis. Arterioscler. Thromb. Vasc. Biol. 25, 1256–1261 (2005).

    Article  CAS  Google Scholar 

  4. Kinchen, J. M. & Hengartner, M. O. Tales of cannibalism, suicide, and murder: programmed cell death in C. elegans. Curr. Top. Dev. Biol. 65, 1–45 (2005).

    CAS  PubMed  Google Scholar 

  5. Henson, P. M. & Hume, D. A. Apoptotic cell removal in development and tissue homeostasis. Trends Immunol. 27, 244–250 (2006).

    Article  CAS  Google Scholar 

  6. Franz, S. et al. Apoptosis and autoimmunity: when apoptotic cells break their silence. Curr. Rheumatol. Rep. 8, 245–247 (2006).

    Article  CAS  Google Scholar 

  7. Gaipl, U. S. et al. Inefficient clearance of dying cells and autoreactivity. Curr. Top. Microbiol. Immunol. 305, 161–176 (2006).

    CAS  PubMed  Google Scholar 

  8. Lettre, G. & Hengartner, M. O. Developmental apoptosis in C. elegans: a complex CEDnario. Nature Rev. Mol. Cell Biol. 7, 97–108 (2006).

    Article  CAS  Google Scholar 

  9. Gumienny, T. L., Lambie, E., Hartwieg, E., Horvitz, H. R. & Hengartner, M. O. Genetic control of programmed cell death in the Caenorhabditis elegans hermaphrodite germline. Development 126, 1011–1022 (1999).

    CAS  PubMed  Google Scholar 

  10. Kinchen, J. M. & Ravichandran, K. S. Journey to the grave: signaling events regulating removal of apoptotic cells. J. Cell Sci. 120, 2143–2149 (2007).

    Article  CAS  Google Scholar 

  11. Mangahas, P. M. & Zhou, Z. Clearance of apoptotic cells in Caenorhabditis elegans. Semin. Cell Dev. Biol. 16, 295–306 (2005).

    Article  CAS  Google Scholar 

  12. Gruenberg, J. & van der Goot, F. G. Mechanisms of pathogen entry through the endosomal compartments. Nature Rev. Mol. Cell Biol. 7, 495–504 (2006).

    Article  CAS  Google Scholar 

  13. Grosshans, B. L., Ortiz, D. & Novick, P. Rabs and their effectors: achieving specificity in membrane traffic. Proc. Natl Acad. Sci. USA 103, 11821–11827 (2006).

    Article  CAS  Google Scholar 

  14. Horazdovsky, B. F. & Emr, S. D. The VPS16 gene product associates with a sedimentable protein complex and is essential for vacuolar protein sorting in yeast. J. Biol. Chem. 268, 4953–4962 (1993).

    CAS  PubMed  Google Scholar 

  15. Sato, T. K., Rehling, P., Peterson, M. R. & Emr, S. D. Class C Vps protein complex regulates vacuolar SNARE pairing and is required for vesicle docking/fusion. Mol. Cell 6, 661–671 (2000).

    Article  CAS  Google Scholar 

  16. Vieira, O. V. et al. Distinct roles of class I and class III phosphatidylinositol 3-kinases in phagosome formation and maturation. J. Cell Biol. 155, 19–25 (2001).

    Article  CAS  PubMed Central  Google Scholar 

  17. Gengyo-Ando, K. et al. The SM protein VPS-45 is required for RAB-5-dependent endocytic transport in Caenorhabditis elegans. EMBO Rep. 8, 152–157 (2007).

    Article  CAS  PubMed Central  Google Scholar 

  18. Hayakawa, A. et al. The WD40 and FYVE domain containing protein 2 defines a class of early endosomes necessary for endocytosis. Proc. Natl Acad. Sci. USA 103, 11928–11933 (2006).

    Article  CAS  Google Scholar 

  19. Carney, D. S., Davies, B. A. & Horazdovsky, B. F. Vps9 domain-containing proteins: activators of Rab5 GTPases from yeast to neurons. Trends Cell Biol. 16, 27–35 (2006).

    Article  CAS  Google Scholar 

  20. Sato, M. et al. Caenorhabditis elegans RME-6 is a novel regulator of RAB-5 at the clathrin-coated pit. Nature Cell Biol. 7, 559–569 (2005).

    Article  CAS  Google Scholar 

  21. Lettre, G. et al. Genome-wide RNAi identifies p53-dependent and -independent regulators of germ cell apoptosis in C. elegans. Cell Death Differ. 11, 1198–1203 (2004).

    Article  CAS  Google Scholar 

  22. Kinchen, J. M. et al. Two pathways converge at CED-10 to mediate actin rearrangement and corpse removal in C. elegans. Nature 434, 93–99 (2005).

    Article  CAS  Google Scholar 

  23. Kamath, R. S. et al. Systematic functional analysis of the Caenorhabditis elegans genome using RNAi. Nature 421, 231–237 (2003).

    Article  CAS  Google Scholar 

  24. Thompson, H. M., Skop, A. R., Euteneuer, U., Meyer, B. J. & McNiven, M. A. The large GTPase dynamin associates with the spindle midzone and is required for cytokinesis. Curr. Biol. 12, 2111–2117 (2002).

    Article  CAS  PubMed Central  Google Scholar 

  25. Clark, S. G., Shurland, D. L., Meyerowitz, E. M., Bargmann, C. I. & van der Bliek, A. M. A dynamin GTPase mutation causes a rapid and reversible temperature-inducible locomotion defect in C. elegans. Proc. Natl Acad. Sci. USA 94, 10438–10443 (1997).

    Article  CAS  Google Scholar 

  26. Yu, X., Odera, S., Chuang, C. H., Lu, N. & Zhou, Z. C. elegans Dynamin mediates the signaling of phagocytic receptor CED-1 for the engulfment and degradation of apoptotic cells. Dev. Cell 10, 743–757 (2006).

    Article  CAS  Google Scholar 

  27. Orth, J. D. & McNiven, M. A. Dynamin at the actin-membrane interface. Curr. Opin. Cell Biol. 15, 31–39 (2003).

    Article  CAS  Google Scholar 

  28. Krueger, E. W., Orth, J. D., Cao, H. & McNiven, M. A. A dynamin-cortactin-Arp2/3 complex mediates actin reorganization in growth factor-stimulated cells. Mol. Biol. Cell 14, 1085–1096 (2003).

    Article  CAS  PubMed Central  Google Scholar 

  29. Schafer, D. A. Coupling actin dynamics and membrane dynamics during endocytosis. Curr. Opin. Cell Biol. 14, 76–81 (2002).

    Article  CAS  Google Scholar 

  30. Gold, E. S. et al. Dynamin 2 is required for phagocytosis in macrophages. J. Exp. Med. 190, 1849–1856 (1999).

    Article  CAS  PubMed Central  Google Scholar 

  31. Christoforidis, S. et al. Phosphatidylinositol-3-OH kinases are Rab5 effectors. Nature Cell Biol. 1, 249–252 (1999).

    Article  CAS  Google Scholar 

  32. Roggo, L. et al. Membrane transport in Caenorhabditis elegans: an essential role for VPS34 at the nuclear membrane. EMBO J. 21, 1673–1683 (2002).

    Article  CAS  PubMed Central  Google Scholar 

  33. Chang, H. C., Hull, M. & Mellman, I. The J-domain protein Rme-8 interacts with Hsc70 to control clathrin-dependent endocytosis in Drosophila. J. Cell Biol. 164, 1055–64 (2004).

    Article  CAS  PubMed Central  Google Scholar 

  34. Bieri, T. et al. WormBase: new content and better access. Nucleic Acids Res. 35, D506–D510 (2007).

    Article  CAS  Google Scholar 

  35. Grimsley, C. M., Lu, M., Haney, L. B., Kinchen, J. M. & Ravichandran, K. S. Characterization of a novel interaction between ELMO1 and ERM proteins. J. Biol. Chem. 281, 5928–5937 (2006).

    Article  CAS  Google Scholar 

  36. Tosello-Trampont, A. C. et al. Identification of two signaling submodules within the CrkII/ELMO/Dock180 pathway regulating engulfment of apoptotic cells. Cell Death Differ. 14, 963–972 (2007).

    Article  CAS  Google Scholar 

  37. Lu, M. et al. PH domain of ELMO functions in trans to regulate Rac activation via Dock180. Nature Struct. Mol. Biol. 11, 756–762 (2004).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors would like to thank members of the Ravichandran and Hengartner laboratories for helpful input and suggestions. We thank Dorothy Schafer for dynamin plasmids and purified Dyn2 protein, David Castle, Jim Casanova and Heidi McBride for Rab5 expression constructs, Jae Jung for Vps34 expression constructs, Lukas Neukomm for pLN022 and pLN019 plasmids and Jan Redick and Christie Davis from the Advanced Microscopy Facility for help with confocal microscopy. Some strains used in this work were obtained from the Caenorhabditis Genetics Center (CGC). This work was supported by grants from the NIGMS/NIH to K. S. R. and from the EU Project Apoclear, the Ernst Hadorn Foundation, the University of Zurich and the Swiss National Science Foundation to M. O. H. J. M. K. is an Arthritis Foundation Postdoctoral Fellow.

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Author notes

  1. Lilli Stergiou

    Present address: Present address: Institute of Molecular Systems Biology, ETH Zürich 8093 Zürich, Switzerland.,

  2. Jason M. Kinchen and Kimon Doukoumetzidis: These authors contributed equally to this work

  3. Michael O. Hengartner and Kodi S. Ravichandran: These authors contributed equally to this work

Authors and Affiliations

  1. Beirne Carter Center for Immunology Research, University of Virginia, Charlottesville, 22908, VA, USA

    Jason M. Kinchen, Annie Tosello-Trampont & Kodi S. Ravichandran

  2. Institute of Molecular Biology, University of Zurich, Zurich, CH-8057, Switzerland

    Kimon Doukoumetzidis, Johann Almendinger, Lilli Stergiou & Michael O. Hengartner

  3. Department of Medicine, University of Virginia, Charlottesville, 22908, VA, USA

    Costi D. Sifri

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Contributions

J. M. K., K. D., J. A. and L. S. contributed to the generation of nematode transgenics and fluorescence microscopy studies; K. D. and J. A. conducted the unbiased screen; J. M. K. conducted the targeted screen; J. M. K. performed the mammalian cell culture experiments and wrote the manuscript; all authors contributed to editing the manuscript; A. C. T., C. D. S., M. O. H. and K. S. R. contributed to the data analysis, project planning and writing of the manuscript.

Corresponding authors

Correspondence to Michael O. Hengartner or Kodi S. Ravichandran.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Figures S1, S2, S3, S4, S5, S6, Supplementary Tables S1, S2, S3, S4, S5 and Supplementary Methods (PDF 1741 kb)

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Kinchen, J., Doukoumetzidis, K., Almendinger, J. et al. A pathway for phagosome maturation during engulfment of apoptotic cells. Nat Cell Biol 10, 556–566 (2008). https://doi.org/10.1038/ncb1718

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