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EEA1 links PI(3)K function to Rab5 regulation of endosome fusion

Nature volume 394, pages 494–498 (1998)Cite this article

Abstract

GTPases and lipid kinases regulate membrane traffic along the endocytic pathway by mechanisms that are not completely understood1,2,3,4. Fusion between early endosomes requires phosphatidyl-inositol-3-OH kinase (PI(3)K) activity5,6,7 as well as the small GTPase Rab5 (ref. 8). Excess Rab5–GTP complex restores endosome fusion when PI(3)K is inhibited5,9. Here we identify the early-endosomal autoantigen EEA1 (10–12) which binds the PI(3)K product phosphatidylinositol-3-phosphate, as a new Rab5 effector that is required for endosome fusion. The association of EEA1 with the endosomal membrane requires Rab5–GTP and PI(3)K activity, and excess Rab5–GTP stabilizes the membrane association of EEA1 even when PI(3)K is inhibited. The identification of EEA1 as a direct Rab5 effector provides a molecular link between PI(3)K and Rab5, and its restricted distribution to early endosomes10 indicates that EEA1 may confer directionality to Rab5-dependent endocytic transport.

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Figure 1: Biochemical interaction between EEA1 and Rab5.
Figure 2: The C-terminal part of EEA1 inhibits basal and Rab5-stimulated early-endosome fusion in vivo.
Figure 3: EEA1 is required for early endosome fusion in vitro.
Figure 4: The membrane association of EEA1 is regulated by Rab5 and PI(3)K.

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References

  1. Novick, P. & Zerial, M. The diversity of Rab proteins in vesicle transport. Curr. Opin. Cell Biol. 9, 496–504 (1997).

    Article  CAS  Google Scholar 

  2. Olkkonen, V. M. & Stenmark, H. Role of rab GTPases in membrane traffic. Int. Rev. Cytol. 176, 1–85 (1997).

    Article  CAS  Google Scholar 

  3. Roth, M. G. & Sternweis, P. C. The role of lipid signaling in constitutive membrane traffic. Curr. Opin. Cell Biol. 9, 519–526 (1997).

    Article  CAS  Google Scholar 

  4. De Camilli, P., Emr, S. D., McPherson, P. S. & Novick, P. Phosphoinositides as regulators in membrane traffic. Science 271, 1533–1539 (1996).

    Article  ADS  CAS  Google Scholar 

  5. Li, G.et al. Evidence for phosphatidylinositol 3-kinase as a regulator of endocytosis via activation of Rab5. Proc. Natl Acad. Sci. USA 92, 10207–10211 (1995).

    Article  ADS  CAS  Google Scholar 

  6. Jones, A. T. & Clague, M. J. Phosphatidylinositol 3-kinase activity is required for early endosome fusion. Biochem. J. 311, 31–34 (1995).

    Article  CAS  Google Scholar 

  7. Spiro, D. J., Boll, W., Kirchhausen, T. & Wessling-Resnick, M. Wortmannin alters the transferrin receptor endocytic pathway in vivo and in vitro. Mol. Biol. Cell 7, 355–367 (1996).

    Article  CAS  Google Scholar 

  8. Gorvel, J. P., Chavrier, P., Zerial, M. & Gruenberg, J. rab5 controls early endosome fusion in vitro. Cell 64, 915–925 (1991).

    Article  CAS  Google Scholar 

  9. Jones, A. T., Mills, I. G., Scheidig, A. J., Alexandrov, K. & Clague, M. J. Inhibition of endosome fusion by wortmannin persists in the presence of activated rab5. Mol. Biol. Cell 9, 323–332 (1998).

    Article  CAS  Google Scholar 

  10. Mu, F. T.et al. EEA1, an early endosome-associated protein. EEA1 is a conserved alpha-helical peripheral membrane protein flanked by cysteine “fingers” and contains a calmodulin-binding IQ motif. J. Biol. Chem. 270, 13503–13511 (1995).

    Article  CAS  Google Scholar 

  11. Patki, V., Virbasius, J., Lane, W. S., Toh, B. H., Shpetner, H. S. & Corvera, S. Identification of an early endosomal protein regulated by phosphatidylinositol 3-kinase. Proc. Natl Acad. Sci. USA 94, 7326–7330 (1997).

    Article  ADS  CAS  Google Scholar 

  12. Gaullier, J.-M. et al. Afunctional PtdIns(3)P-binding motif. Nature 394, 433–434 (1998).

    Article  ADS  Google Scholar 

  13. Stenmark, H., Aasland, R., Toh, B. H. & D'Arrigo, A. Endosomal localization of the autoantigen EEA1 is mediated by a zinc-binding FYVE finger. J. Biol. Chem. 271, 24048–24054 (1996).

    Article  CAS  Google Scholar 

  14. Vitale, G.et al. Distinct Rab-binding domains mediate the interaction of rabaptin-5 with GTP-bound rab4 and rab5. EMBO J. 17, 1941–1951 (1998).

    Article  CAS  Google Scholar 

  15. Stenmark, H., Parton, R. G., Steele-Mortimer, O., Lütcke, A., Gruenberg, J. & Zerial, M. Inhibition of rab5 GTPase activity stimulates membrane fusion in endocytosis. EMBO J. 13, 1287–1296 (1994).

    Article  CAS  Google Scholar 

  16. Horiuchi, H.et al. Anovel Rab5 GDP/GTP exchange factor complexed to rabaptin-5 links nucleotide exchange to effector recruitment and function. Cell 90, 1149–1159 (1997).

    Article  CAS  Google Scholar 

  17. Shpetner, H., Joly, M., Hartley, D. & Corvera, S. Potential sites of PI-3 kinase function in the endocytic pathway revealed by the PI-3 kinase inhibitor, wortmannin. J. Cell Biol. 132, 595–605 (1996).

    Article  CAS  Google Scholar 

  18. Rybin, V.et al. GTPase activity of Rab5 acts as a timer for endocytic membrane fusion. Nature 383, 266–269 (1996).

    Article  ADS  CAS  Google Scholar 

  19. D'Arrigo, A., Bucci, C., Toh, B. H. & Stenmark, H. Microtubules are involved in bafilomycin A1-induced tubulation and Rab5-dependent vacuolation of early endosomes. Eur. J. Cell Biol. 72, 95–103 (1997).

    CAS  PubMed  Google Scholar 

  20. Stenmark, H., Vitale, G., Ullrich, O. & Zerial, M. Rabaptin-5 is a direct effector of the small GTPase Rab5 in endocytic membrane fusion. Cell 83, 423–432 (1995).

    Article  CAS  Google Scholar 

  21. Chavrier, P., Parton, R. G., Hauri, H. P., Simons, K. & Zerial, M. Localization of low molecular weight GTP binding proteins to exocytic and endocytic compartments. Cell 62, 317–329 (1990).

    Article  CAS  Google Scholar 

  22. Bucci, C.et al. The small GTPase rab5 functions as a regulatory factor in the early endocytic pathway. Cell 70, 715–728 (1992).

    Article  CAS  Google Scholar 

  23. Vojtek, A. B., Hollenberg, S. M. & Cooper, J. A. Mammalian Ras interacts directly with the serine/threonine kinase Raf. Cell 74, 205–214 (1993).

    Article  CAS  Google Scholar 

  24. Stenmark, H., Bucci, C. & Zerial, M. Expression of Rab GTPases using recombinant vaccinia viruses. Methods Enzymol. 257, 155–164 (1995).

    Article  CAS  Google Scholar 

  25. Evan, G. I., Lewis, G. K., Ramsay, G. & Bishop, J. M. Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product. Mol. Cell. Biol. 5, 3610–3616 (1985).

    Article  CAS  Google Scholar 

  26. Tooze, J. & Hollinshead, M. Tubular early endosome networks in AtT20 and other cells. J. Cell Biol. 115, 635–653 (1991).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank E. Rønning for technical assistance; D. Warren for help with the baculovirus system; N. Salmon and T. Nordeng for help with confocal microscopy; V. Rybin for providing the Rab5D136N–Rep1 complex; E. Kolpakova for anti-MBP antibodies; and H. McBride, S. Olsnes, K.Sandvig and B. Sönnichsen for comments on the manuscript. H.S. was supported by the Top Research Programme, the Research Council of Norway, the Norwegian Cancer Society and the Novo Nordisk Foundation. This work was supported by a European Community TMR grant (to H.S. and M.Z.).

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Author notes

  1. Carol Murphy

    Present address: University of Ioannina Medical School, 45110, Ioannina, Greece

  2. Anne Simonsen, Roger Lippe, Judy Callaghan and Ban-Hock Toh: These authors contributed equally to this work

Authors and Affiliations

  1. Department of Biochemistry, The Norwegian Radium Hospital, Montebello, N-0310, Oslo, Norway

    Anne Simonsen, Jean-Michel Gaullier & Harald Stenmark

  2. EMBL, Meyerhofstrasse, D-69012 1, Heidelberg, Germany

    Roger Lippe, Savvas Christoforidis, Carol Murphy & Marino Zerial

  3. EM-unit, Institute of Biology, PO Box 1050, Blindern, N-0316, Oslo, Norway

    Andreas Brech

  4. Department of Pathology and Immunology, Monash Medical School, Prahran, 3181, Victoria, Australia

    Judy Callaghan & Ban-Hock Toh

  5. Max Planck Institute for Molecular Cell Biology and Genetics, 01307, Dresden, Germany

    Roger Lippe, Savvas Christoforidis, Carol Murphy & Marino Zerial

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Correspondence to Harald Stenmark.

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Simonsen, A., Lippe, R., Christoforidis, S. et al. EEA1 links PI(3)K function to Rab5 regulation of endosome fusion. Nature 394, 494–498 (1998). https://doi.org/10.1038/28879

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