Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

The GM130 and GRASP65 Golgi proteins cycle through and define a subdomain of the intermediate compartment

Nature Cell Biology volume 3, pages 1101–1113 (2001)Cite this article

Abstract

Integrating the pleomorphic membranes of the intermediate compartment (IC) into the array of Golgi cisternae is a crucial step in membrane transport, but it is poorly understood. To gain insight into this step, we investigated the dynamics by which cis-Golgi matrix proteins such as GM130 and GRASP65 associate with, and incorporate, incoming IC elements. We found that GM130 and GRASP65 cycle via membranous tubules between the Golgi complex and a constellation of mobile structures that we call late IC stations. These stations are intermediate between the IC and the cis-Golgi in terms of composition, and they receive cargo from earlier IC elements and deliver it to the Golgi complex. Late IC elements are transient in nature and sensitive to fixatives; they are seen in only a fraction of fixed cells, whereas they are always visible in living cells. Finally, late IC stations undergo homotypic fusion and establish tubular connections between themselves and the Golgi. Overall, these features indicate that late IC stations mediate the transition between IC elements and the cis-Golgi face.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: GM130 cycles between the Golgi complex and peripheral compartments via tubules.
Figure 2: GM130-positive peripheral structures have compositional features of an IC subdomain.
Figure 3: Ultrastructural analysis of GM130-positive tubules and puncta with correlative light-electron microscopy (CLEM) and cryo-immunoelectromicroscopy (cryo-EM).
Figure 4: GM130-positive L-IC elements contain and transport secretory cargo.
Figure 5: GM130-positive L-IC elements transport secretory cargo.
Figure 6: GM130 distribution after synchronization at 15 °C.
Figure 7: p115 and GRASP65 control the dynamics of GM130-positive L-IC stations and tubules.
Figure 8: Interfering with GM130-tubule dynamics with G95 inhibits the transport of VSV-G protein to the Golgi complex.

Similar content being viewed by others

References

  1. Godi, A. et al. ADP ribosylation factor regulates spectrin binding to the Golgi complex. Proc. Natl Acad. Sci. USA 95, 8607–8612 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Godi, A. et al. ARF mediates recruitment of PtdIns-4-OH kinase-beta and stimulates synthesis of PtdIns(4,5)P2 on the Golgi complex. Nature Cell Biol. 1, 280–287 (1999).

    Article  CAS  PubMed  Google Scholar 

  3. De Matteis, M. A. & Morrow, J. S. Spectrin tethers and mesh in the biosynthetic pathway. J. Cell Sci. 113, 2331–2343 (2000).

    CAS  PubMed  Google Scholar 

  4. Nakamura, N. et al. Characterization of a cis-Golgi matrix protein, GM130. J. Cell Biol. 131, 1715–1726 (1995).

    Article  CAS  PubMed  Google Scholar 

  5. Slusarewicz, P., Nilsson, T., Hui, N., Watson, R. & Warren, G. Isolation of a matrix that binds medial Golgi enzymes. J. Cell Biol. 124, 405–413 (1994).

    Article  CAS  PubMed  Google Scholar 

  6. Barr, F. A., Puype, M., Vandekerckhove, J. & Warren, G. GRASP65, a protein involved in the stacking of Golgi cisternae. Cell 91, 253–262 (1997).

    Article  CAS  PubMed  Google Scholar 

  7. Shorter, J. & Warren, G. A role for the vesicle tethering protein, p115, in the post-mitotic stacking of reassembling Golgi cisternae in a cell-free system. J. Cell Biol. 146, 57–70 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Sonnichsen, B. et al. A role for giantin in docking COPI vesicles to Golgi membranes. J. Cell Biol. 140, 1013–1021 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Seemann, J., Jokitalo, E., Pypaert, M. & Warren, G. Matrix proteins can generate the higher order architecture of the Golgi apparatus. Nature 407, 1022–1026 (2000).

    Article  CAS  PubMed  Google Scholar 

  10. Klumperman, J. Transport between ER and Golgi. Curr. Opin. Cell Biol. 12, 445–449 (2000).

    Article  CAS  PubMed  Google Scholar 

  11. Krijnse-Locker, J., Ericsson, M., Rottier, P. J. & Griffiths, G. Characterization of the budding compartment of mouse hepatitis virus: evidence that transport from the RER to the Golgi complex requires only one vesicular transport step. J. Cell Biol. 124, 55–70 (1994).

    Article  CAS  PubMed  Google Scholar 

  12. Sitia, R. & Meldolesi, J. Endoplasmic reticulum: a dynamic patchwork of specialized subregions. Mol. Biol. Cell 3, 1067–1072 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Mellman, I. & Simons, K. The Golgi complex: in vitro veritas? Cell 68, 829–840 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Bannykh, S. I. & Balch, W. E. Membrane dynamics at the endoplasmic reticulum-Golgi interface. J. Cell Biol. 138, 1–4 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Lotti, L. V., Torrisi, M. R., Pascale, M. C. & Bonatti, S. Immunocytochemical analysis of the transfer of vesicular stomatitis virus G glycoprotein from the intermediate compartment to the Golgi complex. J. Cell Biol. 118, 43–50 (1992).

    Article  CAS  PubMed  Google Scholar 

  16. Pelham, H. R. Control of protein exit from the endoplasmic reticulum. Annu. Rev. Cell Biol. 5, 1–23 (1989).

    Article  CAS  PubMed  Google Scholar 

  17. Saraste, J. & Svensson, K. Distribution of the intermediate elements operating in ER to Golgi transport. J. Cell Sci. 100, 415–430 (1991).

    CAS  PubMed  Google Scholar 

  18. Ying, M., Flatmark, T. & Saraste, J. The p58-positive pre-golgi intermediates consist of distinct subpopulations of particles that show differential binding of COPI and COPII coats and contain vacuolar H(+)-ATPase. J. Cell Sci. 113, 3623–3638 (2000).

    CAS  PubMed  Google Scholar 

  19. Presley, J. F. et al. ER-to-Golgi transport visualized in living cells. Nature 389, 81–85 (1997).

    Article  CAS  PubMed  Google Scholar 

  20. Scales, S. J., Pepperkok, R. & Kreis, T. E. Visualization of ER-to-Golgi transport in living cells reveals a sequential mode of action for COPII and COPI. Cell 90, 1137–1148 (1997).

    Article  CAS  PubMed  Google Scholar 

  21. Hauri, H. P., Kappeler, F., Andersson, H. & Appenzeller, C. ERGIC-53 and traffic in the secretory pathway. J. Cell Sci. 113, 587–596 (2000).

    CAS  PubMed  Google Scholar 

  22. Hammond, A. T. & Glick, B. S. Dynamics of transitional endoplasmic reticulum sites in vertebrate cells. Mol. Biol. Cell 11, 3013–3030 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Shima, D. T., Scales, S. J., Kreis, T. E. & Pepperkok, R. Segregation of COPI-rich and anterograde-cargo-rich domains in endoplasmic-reticulum-to-Golgi transport complexes. Curr. Biol. 9, 821–824 (1999).

    Article  CAS  PubMed  Google Scholar 

  24. Stephens, D. J., Lin-Marq, N., Pagano, A., Pepperkok, R. & Paccaud, J. P. COPI-coated ER-to-Golgi transport complexes segregate from COPII in close proximity to ER exit sites. J. Cell Sci. 113, 2177–2185 (2000).

    CAS  PubMed  Google Scholar 

  25. Perez-Vilar, J., Hidalgo, J. & Velasco, A. Presence of terminal N-acetylgalactosamine residues in subregions of the endoplasmic reticulum is influenced by cell differentiation in culture. J. Biol. Chem. 266, 23967–23976 (1991).

    CAS  PubMed  Google Scholar 

  26. Klumperman, J. et al. The recycling pathway of protein ERGIC-53 and dynamics of the ER-Golgi intermediate compartment. J. Cell Sci. 111, 3411–3425 (1998).

    CAS  PubMed  Google Scholar 

  27. Tang, B. L., Low, S. H., Hauri, H. P. & Hong, W. Segregation of ERGIC53 and the mammalian KDEL receptor upon exit from the 15 degrees C compartment. Eur. J. Cell Biol. 68, 398–410 (1995).

    CAS  PubMed  Google Scholar 

  28. Mironov, A. A., Polishchuk, R. S. & Luini, A. Visualizing membrane traffic in vivo by combined video fluorescence and 3D electron microscopy. Trends Cell Biol. 10, 349–353 (2000).

    Article  CAS  PubMed  Google Scholar 

  29. Powell, R. D., Halsey, C. M. & Hainfeld, J. F. Combined fluorescent and gold immunoprobes: reagents and methods for correlative light and electron microscopy. Microsc. Res. Technol. 42, 2–12 (1998).

    Article  CAS  Google Scholar 

  30. Kuismanen, E. & Saraste, J. Low temperature-induced transport blocks as tools to manipulate membrane traffic. Methods Cell Biol. 32, 257–274 (1989).

    Article  CAS  PubMed  Google Scholar 

  31. Barroso, M., Nelson, D. S. & Sztul, E. Transcytosis-associated protein (TAP)/p115 is a general fusion factor required for binding of vesicles to acceptor membranes. Proc. Natl Acad. Sci. USA 92, 527–531 (1995).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Waters, M. G., Clary, D. O. & Rothman, J. E. A novel 115-kD peripheral membrane protein is required for intercisternal transport in the Golgi stack. J. Cell Biol. 118, 1015–1026 (1992).

    Article  CAS  PubMed  Google Scholar 

  33. Alvarez, C., Garcia-Mata, R., Hauri, H. P. & Sztul, E. The p-115 inteactive proteins, GM130 and giantin participate in ER-Golgi traffic. J. Biol. Chem. 276, 2693–2700 (2001).

    Article  CAS  PubMed  Google Scholar 

  34. Seemann, J., Jokitalo, E. J. & Warren, G. The role of the tethering proteins p115 and GM130 in transport through the Golgi apparatus in vivo. Mol. Biol. Cell 11, 635–645 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Barr, F. A., Nakamura, N. & Warren, G. Mapping the interaction between GRASP65 and GM130, components of a protein complex involved in the stacking of Golgi cisternae. EMBO J. 17, 3258–3268 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Nakamura, N., Lowe, M., Levine, T. P., Rabouille, C. & Warren, G. The vesicle docking protein p115 binds GM130, a cis-Golgi matrix protein, in a mitotically regulated manner. Cell 89, 445–455 (1997).

    Article  CAS  PubMed  Google Scholar 

  37. Alvarez, C., Fujita, H., Hubbard, A. & Sztul, E. ER to Golgi transport: Requirement for p115 at a pre-Golgi VTC stage. J. Cell Biol. 147, 1205–1222 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Garcia-Mata, R., Gao, Y., Alvarez, C. & Sztul, E. S. The membrane transport factor p115 recycles only between homologous compartments in intact heterokaryons. Eur. J. Cell Biol. 79, 229–239 (2000).

    Article  CAS  PubMed  Google Scholar 

  39. Hopkins, C. R., Gibson, A., Shipman, M. & Miller, K. Movement of internalized ligand-receptor complexes along a continuous endosomal reticulum. Nature 346, 335–339 (1990).

    Article  CAS  PubMed  Google Scholar 

  40. Blum, R., Stephens, D. J. & Schulz, I. Lumenal targeted GFP, used as a marker of soluble cargo, visualises rapid ERGIC to golgi traffic by a tubulo-vesicular network. J. Cell Sci. 113, 3151–3159 (2000).

    CAS  PubMed  Google Scholar 

  41. Moyer, B. D., Allan, B. B. & Balch, W. E. Rab1 Interaction with a GM130 Effector Complex Regulates COPII Vesicle cis-Golgi Tethering. Traffic 2, 268–276 (2001).

    Article  CAS  PubMed  Google Scholar 

  42. Chao, D. S. et al. SNARE membrane trafficking dynamics in vivo. J. Cell Biol. 144, 869–881 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Hay, J. C. et al. Localization, dynamics, and protein interactions reveal distinct roles for ER and Golgi SNAREs. J. Cell Biol. 141, 1489–1502 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Weide, T. et al. The Golgi matrix protein GM130: a specific interacting partner of the small GTPase rab1b. EMBO Rep. 2, 336–341 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Allan, B. B., Moyer, B. D. & Balch, W. E. Rab1 recruitment of p115 into a cis-SNARE complex: programming budding COPII vesicles for fusion. Science 289, 444–448 (2000).

    Article  CAS  PubMed  Google Scholar 

  46. Fritzler, M. J., Hamel, J. C., Ochs, R. L. & Chan, E. K. Molecular characterization of two human autoantigens: unique cDNAs encoding 95- and 160-kD proteins of a putative family in the Golgi complex. J. Exp. Med. 178, 49–62 (1993).

    Article  CAS  PubMed  Google Scholar 

  47. Gallione, C. J. & Rose, J. K. A single amino acid substitution in a hydrophobic domain causes temperature-sensitive cell-surface transport of a mutant viral glycoprotein. J. Virol. 54, 374–382 (1985).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Mironov, A. et al. Role of NAD+ and ADP-ribosylation in the maintenance of the Golgi structure. J. Cell Biol. 139, 1109–1118 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Peters, P. J. & Hunziker, W. Subcellular localization of Rab17 by cryo-immunogold electron microscopy in epithelial cells grown on polycarbonate filters. Methods Enzymol. 329, 210–225 (2001).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank P. Hauri, J. Gruenberg, B. L. Tang, J. Lippincott-Schwartz and F. Barr for antibodies, C. P. Berrie for comments on the manuscript, and Y. Misumi for G95 constructs. Supported in part by grants from Telethon (E.732), the Human Frontier Science Programme (to M.A.D.M.), the European Community (EU grant No. HPRN-CT-2000–00081), the Italian National Research Council (99.00127.PF49 and 99.00133.PF49), and the Italian Association for Cancer Research (AIRC, Milan, Italy). The human GM130 nucleotide sequence data has been submitted to the EMBL/GenBank/DDBJ data libraries with accession number D86427.

Author information

Authors and Affiliations

  1. Department of Cell Biology and Oncology, Istituto di Ricerche Farmacologiche Mario Negri, Consorzio Mario Negri Sud, Santa Maria Imbaro (Chieti), 66030, Italy

    Pierfrancesco Marra, Tania Maffucci, Tiziana Daniele, Giuseppe Di Tullio, Alberto Luini, Gala Beznoussenko, Alexander Mironov & Maria Antonietta De Matteis

  2. Department of Biochemistry, Fukuoka University School of Medicine, 814-0180, Fukuoka, Japan

    Yukio Ikehara

  3. Department of Molecular and Experimental Medicine, The Scripps Research Institute, La Jolla, 92037, California, USA

    Edward K. L. Chan

  4. Maria Antonietta De Matteis

Authors
  1. Pierfrancesco Marra
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Tania Maffucci
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tiziana Daniele
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Giuseppe Di Tullio
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Yukio Ikehara
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Edward K. L. Chan
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Alberto Luini
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Gala Beznoussenko
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Alexander Mironov
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Maria Antonietta De Matteis
    View author publications

    You can also search for this author in PubMed Google Scholar

Supplementary information

Movie 1

Dynamics of GFP-GM130. Time laps images from COS7 cells transfected with GFP-GM130. The numbers in the bottom right corner designate frame numbers and time elapsed in h.mm.ss., respectively. (MOV 6310 kb)

Movie 2

Dynamics of GFP-GM130. Time laps images from COS7 cells transfected with GFP-GM130. The numbers in the bottom left corner designate frame numbers and time elapsed in h.mm.ss., respectively. (MOV 5616 kb)

Movie 3

Dynamics of YFP-GM130 and CFP-VSV-G upon release from the 15 °C block. Time laps images images from COS7 cells transfected with YFP-GM130 (red) and CFP-VSV-G (green) showing the fluorescence recovery after photobleaching of the central Golgi area (white circle) upon release from the 15 °C block. (MOV 6460 kb)

Movie 4

Dynamics of GFP-G95. Time laps images from COS7 cells transfected with GFP-G95. The numbers in the bottom left corner designate frame numbers and time elapsed in h.mm.ss., respectively. (MOV 1082 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Marra, P., Maffucci, T., Daniele, T. et al. The GM130 and GRASP65 Golgi proteins cycle through and define a subdomain of the intermediate compartment. Nat Cell Biol 3, 1101–1113 (2001). https://doi.org/10.1038/ncb1201-1101

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ncb1201-1101

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing