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Distinct AAA-ATPase p97 complexes function in discrete steps of nuclear assembly

Nature Cell Biology volume 3, pages 1086–1091 (2001)Cite this article

Abstract

Although nuclear envelope (NE) assembly is known to require the GTPase Ran, the membrane fusion machinery involved is uncharacterized. NE assembly involves formation of a reticular network on chromatin, fusion of this network into a closed NE and subsequent expansion. Here we show that p97, an AAA-ATPase previously implicated in fusion of Golgi and transitional endoplasmic reticulum (ER) membranes together with the adaptor p47, has two discrete functions in NE assembly. Formation of a closed NE requires the p97–Ufd1–Npl4 complex, not previously implicated in membrane fusion. Subsequent NE growth involves a p97–p47 complex. This study provides the first insights into the molecular mechanisms and specificity of fusion events involved in NE formation.

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Figure 1: Formation of NE and ER in vitro.
Figure 2: p97 is involved in NE and ER formation.
Figure 3: Depletion of p97 adaptors.
Figure 4: Different functions for Ufd1–Npl4 and p47.
Figure 5: Schematic step-wise model of NE formation.

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References

  1. Gerace, L. & Burke, B. Functional organization of the nuclear envelope. Annu. Rev. Cell Biol. 4, 335–374 (1988).

    Article  CAS  Google Scholar 

  2. Gant, T. M. & Wilson K. L. Nuclear assembly. Annu. Rev. Cell Dev. Biol. 13, 669–695 (1997).

    Article  CAS  Google Scholar 

  3. Franke, W. W., Scheer, U., Krohne, G. & Jarasch, E. D. The nuclear envelope and the architecture of the nuclear periphery. J. Cell Biol. 91, 39s–50s (1981).

    Article  CAS  Google Scholar 

  4. Ellenberg, J. et al. Nuclear membrane dynamics and reassembly in living cells: targeting of an inner nuclear membrane protein in interphase and mitosis. J. Cell Biol. 138, 1193–1206 (1997).

    Article  CAS  Google Scholar 

  5. Powell, L. & Burke, B. Internuclear exchange of an inner nuclear membrane protein (p55) in heterokaryons: in vivo evidence for the interaction of p55 with the nuclear lamina. J. Cell Biol. 111, 2225–2234 (1990).

    Article  CAS  Google Scholar 

  6. Soullam, B. & Worman, H. J. Signals and structural features involved in integral membrane protein targeting to the inner nuclear membrane. J. Cell Biol. 130, 15–27 (1995).

    Article  CAS  Google Scholar 

  7. Lohka, M. J. & Masui, Y. Formation in vitro of sperm pronuclei and mitotic chromosomes induced by amphibian ooplasmic components. Science 220, 719–721 (1983).

    Article  CAS  Google Scholar 

  8. Burke, B. & Gerace, L. A cell free system to study reassembly of the nuclear envelope at the end of mitosis. Cell 28, 639–652 (1986).

    Article  Google Scholar 

  9. Dreier, L. & Rapoport, T. A. In vitro formation of the endoplasmic reticulum occurs independently of microtubules by a controlled fusion reaction. J. Cell Biol. 148, 883–898 (2000).

    Article  CAS  Google Scholar 

  10. Macaulay, C. & Forbes, D. J. Assembly of the nuclear pore: biochemically distinct steps revealed with NEM, GTPγS, and BAPTA. J. Cell Biol. 132, 5–20 (1996).

    Article  CAS  Google Scholar 

  11. Vigers, G. P. & Lohka, M. J. A distinct vesicle population targets membranes and pore complexes to the nuclear envelope in Xenopus eggs. J. Cell Biol. 112, 545–556 (1991).

    Article  CAS  Google Scholar 

  12. Newport, J. & Dunphy, W. Characterization of the membrane binding and fusion events during nuclear envelope assembly using purified components. J. Cell Biol. 116, 295–306 (1992).

    Article  CAS  Google Scholar 

  13. Marshall, I. C. B. & Wilson, K. L. Nuclear envelope assembly after mitosis. Trends Cell Biol. 7, 69–74 (1997).

    Article  CAS  Google Scholar 

  14. Boman, A. L., Delannoy, M. R., & Wilson, K. L. GTP hydrolysis is required for vesicle fusion during nuclear envelope assembly in vitro. J. Cell Biol. 116, 281–294 (1992).

    Article  CAS  Google Scholar 

  15. Hetzer, M., Bilbao-Cortes, D., Walther, T. C., Gruss, O. J. & Mattaj, I. W. GTP hydrolysis by Ran is required for nuclear envelope assembly. Mol. Cell 5, 1013–1024 (2000).

    Article  CAS  Google Scholar 

  16. Zhang, C. & Clarke, P. R. Chromatin-independent nuclear envelope assembly induced by Ran GTPase in Xenopus egg extracts. Science 26, 1429–1432 (2000).

    Article  Google Scholar 

  17. Zhang, C. & Clarke, P. R. Roles of Ran-GTP and Ran-GDP in precursor vesicle recruitment and fusion during nuclear envelope assembly in a human cell-free system. Curr. Biol. 6, 208–212 (2001).

    Article  Google Scholar 

  18. Benavente, R., Dabauvalle, M. C., Scheer, U. & Chaly, N. Functional role of newly formed pore complexes in postmitotic nuclear reorganization. Chromosoma 98, 233–241 (1989).

    Article  CAS  Google Scholar 

  19. Wilson, K. L. & Newport, J. A trypsin-sensitive receptor on membrane vesicles is required for nuclear envelope formation in vitro. J. Cell Biol. 107, 57–68 (1988).

    Article  CAS  Google Scholar 

  20. Drummond, S. et al. Temporal differences in the appearance of NEP-B78 and a LBR-like protein during Xenopus nuclear envelope reassembly reflect the order recruitment of functionally discrete vesicle types. J. Cell Biol. 144, 225–240 (1999).

    Article  CAS  Google Scholar 

  21. Sasagawa, S., Yamamoto, A., Ichimura, T., Omata, S. & Horigome, T. In vitro nuclear assembly with affinity-purified nuclear precursor vesicle fractions, PV1 and PV2. Euro. J. Cell Biol. 78, 593–600 (1999).

    Article  CAS  Google Scholar 

  22. Patel, S. & Latterich, M. The AAA team: related ATPases with diverse functions. Trends Cell Biol. 8, 65–71 (1998).

    Article  CAS  Google Scholar 

  23. Kondo, H et al. p47 is a cofactor for p97-mediated membrane fusion. Nature 388, 75–78 (1997).

    Article  CAS  Google Scholar 

  24. Roy, L. et al. Role of p97 and syntaxin 5 in the assembly of transitional endoplasmic reticulum. Mol. Biol. Cell 11, 2529–2542 (2000).

    Article  CAS  Google Scholar 

  25. Rabouille, C. et al. Syntaxin 5 is a common component of the NSF- and p97-mediated reassembly pathways of Golgi cisternae from mitotic Golgi fragments in vitro. Cell 92, 603–610 (1998).

    Article  CAS  Google Scholar 

  26. Meyer, H. H., Shorter, J. G., Seemann, J., Pappin, D. & Warren, G. A complex of mammalian Ufd1 and Npl4 links the AAA-ATPase, p97, to ubiquitin and nuclear transport pathways. EMBO J. 19, 2181–2192 (2000).

    Article  CAS  Google Scholar 

  27. Latterich, M., Frohlich, K. U. & Schekman, R. Membrane fusion and the cell cycle: Cdc48p participates in the fusion of ER membranes. Cell 82, 885–893 (1995).

    Article  CAS  Google Scholar 

  28. Johnson, E. S., Ma, P. C., Ota, I. M. & Varshavsky, A. A proteolytic pathway that recognizes ubiquitin as a degradation signal. J. Biol. Chem. 270, 17442–17456 (1995).

    Article  CAS  Google Scholar 

  29. Ghislain, M., Dohmen, R. J., Levy, F. & Varshavsky, A. Cdc48p interacts with Ufd3p, a WD repeat protein required for ubiquitin-mediated proteolysis in Saccharomyces cerevisiae. EMBO J. 15, 4884–4899 (1996).

    Article  CAS  Google Scholar 

  30. Hoppe, T. et al. Activation of a membrane-bound transcription factor by regulated ubiquitin/proteasome-dependent processing. Cell 102, 577–586 (2000).

    Article  CAS  Google Scholar 

  31. DeHoratius, C. & Silver, P. A. Nuclear transport defects and nuclear envelope alterations are associated with mutation of the Saccharomyces cerevisiae NPL4 gene. Mol. Biol. Cell 7, 1835–1855 (1996).

    Article  CAS  Google Scholar 

  32. Whaley, W. G. The Golgi apparatus. Cell Biology Monographs, Continuation of Protoplasmatologia, 2 (Springer, Vienna, 1975).

    Google Scholar 

  33. Rossanese, O. W. et al. Golgi structure correlates with transitional endoplasmic reticulum organization in Pichia pastoris and Saccharomyces cerevisiae. J. Cell Biol. 145, 68–81 (1999).

    Article  Google Scholar 

  34. Lavoie, C., Lanoix, J., Kan, F. W. & Paiement, J. Cell-free assembly of rough and smooth endoplasmic reticulum. J. Cell Sci. 109, 1415–1425 (1996).

    CAS  PubMed  Google Scholar 

  35. Davis, L. I. & Blobel, G. Nuclear pore complex contains a family of glycoproteins that includes p62: glycosylation through a previously unidentified cellular pathway. Proc. Natl Acad. Sci. USA 84, 7552–7556 (1987).

    Article  CAS  Google Scholar 

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Acknowledgements

M.H. was supported by EMBO and H.H.M. by the Human Frontiers Science Programme Organisation. We thank Jan Ellenberg and the ELMF for help with microscopy. Thanks to W. Antonin, P. Askjaer, K. Czaplinski, L. Englmeier, J. Ellenberg, V. Hachet, B. Huelsmann, M. Ohno and C. Schatz for critical comments on the manuscript.

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  1. Martin Hetzer and Hemmo H. Meyer: These two authors contributed equally to this work.

Authors and Affiliations

  1. European Molecular Biology Laboratory, Meyerhofstrasse 1, Heidelberg, D-69117, Germany

    Martin Hetzer, Tobias C. Walther, Daniel Bilbao-Cortes & Iain W. Mattaj

  2. Department of Cell Biology, Yale University School of Medicine, 333 Cedar Street, SHM, C441, PO Box 208002, New Haven, 06520-8002, Connecticut, USA

    Hemmo H. Meyer & Graham Warren

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  1. Martin Hetzer
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Correspondence to Iain W. Mattaj.

Supplementary information

Supplementary figures

Figure S1 Nuclei formed in the presence of p97 are functional. (PDF 348 kb)

Figure S2 Characterization of NE proteins in p97-restored nuclei.

Figure S3 Nuclei formed in the absence of p47 have an intact NE.

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Hetzer, M., Meyer, H., Walther, T. et al. Distinct AAA-ATPase p97 complexes function in discrete steps of nuclear assembly. Nat Cell Biol 3, 1086–1091 (2001). https://doi.org/10.1038/ncb1201-1086

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