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:

Visualization of a missing link in retrovirus capsid assembly

Nature volume 457, pages 694–698 (2009)Cite this article

Abstract

For a retrovirus such as HIV to be infectious, a properly formed capsid is needed; however, unusually among viruses, retrovirus capsids are highly variable in structure. According to the fullerene conjecture, they are composed of hexamers and pentamers of capsid protein (CA), with the shape of a capsid varying according to how the twelve pentamers are distributed and its size depending on the number of hexamers. Hexamers have been studied in planar and tubular arrays, but the predicted pentamers have not been observed. Here we report cryo-electron microscopic analyses of two in-vitro-assembled capsids of Rous sarcoma virus. Both are icosahedrally symmetric: one is composed of 12 pentamers, and the other of 12 pentamers and 20 hexamers. Fitting of atomic models of the two CA domains into the reconstructions shows three distinct inter-subunit interactions. These observations substantiate the fullerene conjecture, show how pentamers are accommodated at vertices, support the inference that nucleation is a crucial morphologic determinant, and imply that electrostatic interactions govern the differential assembly of pentamers and hexamers.

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: RSV CA protein assembles in vitro in 0.5 M phosphate buffer into small isometric particles.
Figure 3: The 30-nm RSV CA double-layer capsid.
Figure 2: Three-dimensional reconstruction of the RSV CA T = 1 capsid.
Figure 4: Pseudo-atomic models of the RSV-CA subunit, pentamer and hexamer.
Figure 5: Inter-subunit interactions in the T = 1 capsid, represented in stereo views.

Similar content being viewed by others

References

  1. Vogt, V. M. in Retroviruses (eds Coffin, J. M., Hughes, S. H. & Varmus, H.) 27–70 (Cold Spring Harbor Laboratory, 1997)

    Google Scholar 

  2. Benjamin, J., Ganser-Pornillos, B. K., Tivol, W. F., Sundquist, W. I. & Jensen, G. J. Three-dimensional structure of HIV-1 virus-like particles by electron cryotomography. J. Mol. Biol. 346, 577–588 (2005)

    Article  CAS  Google Scholar 

  3. Kingston, R. L., Olson, N. H. & Vogt, V. M. The organization of mature Rous sarcoma virus as studied by cryoelectron microscopy. J. Struct. Biol. 136, 67–80 (2001)

    Article  CAS  Google Scholar 

  4. Butan, C., Winkler, D. C., Heymann, J. B., Craven, R. C. & Steven, A. C. RSV capsid polymorphism correlates with polymerization efficiency and envelope glycoprotein content: implications that nucleation controls morphogenesis. J. Mol. Biol. 376, 1168–1181 (2008)

    Article  CAS  Google Scholar 

  5. Yeager, M., Wilson-Kubalek, E. M., Weiner, S. G., Brown, P. O. & Rein, A. Supramolecular organization of immature and mature murine leukemia virus revealed by electron cryo-microscopy: implications for retroviral assembly mechanisms. Proc. Natl Acad. Sci. USA 95, 7299–7304 (1998)

    Article  ADS  CAS  Google Scholar 

  6. Briggs, J. A., Wilk, T., Welker, R., Kräusslich, H. G. & Fuller, S. D. Structural organization of authentic, mature HIV-1 virions and cores. EMBO J. 22, 1707–1715 (2003)

    Article  CAS  Google Scholar 

  7. Briggs, J. A. et al. The mechanism of HIV-1 core assembly: insights from three-dimensional reconstructions of authentic virions. Structure 14, 15–20 (2006)

    Article  CAS  Google Scholar 

  8. Gamble, T. R. et al. Crystal structure of human cyclophilin A bound to the amino-terminal domain of HIV-1 capsid. Cell 87, 1285–1294 (1996)

    Article  CAS  Google Scholar 

  9. Momany, C. et al. Crystal structure of dimeric HIV-1 capsid protein. Nature Struct. Biol. 3, 763–770 (1996)

    Article  CAS  Google Scholar 

  10. Gitti, R. K. et al. Structure of the amino-terminal core domain of the HIV-1 capsid protein. Science 273, 231–235 (1996)

    Article  ADS  CAS  Google Scholar 

  11. Gamble, T. R. et al. Structure of the carboxy-terminal dimerization domain of the HIV-1 capsid protein. Science 278, 849–853 (1997)

    Article  ADS  CAS  Google Scholar 

  12. Khorasanizadeh, S., Campos-Olivas, R. & Summers, M. F. Solution structure of the capsid protein from the human T-cell leukemia virus type-I. J. Mol. Biol. 291, 491–505 (1999)

    Article  CAS  Google Scholar 

  13. Campos-Olivas, R., Newman, J. L. & Summers, M. F. Solution structure and dynamics of the Rous sarcoma virus capsid protein and comparison with capsid proteins of other retroviruses. J. Mol. Biol. 296, 633–649 (2000)

    Article  CAS  Google Scholar 

  14. Kingston, R. L. et al. Structure and self-association of the Rous sarcoma virus capsid protein. Structure 8, 617–628 (2000)

    Article  CAS  Google Scholar 

  15. Cornilescu, C. C., Bouamr, F., Yao, X., Carter, C. & Tjandra, N. Structural analysis of the N-terminal domain of the human T-cell leukemia virus capsid protein. J. Mol. Biol. 306, 783–797 (2001)

    Article  CAS  Google Scholar 

  16. Mortuza, G. B. et al. High-resolution structure of a retroviral capsid hexameric amino-terminal domain. Nature 431, 481–485 (2004)

    Article  ADS  CAS  Google Scholar 

  17. Li, S., Hill, C. P., Sundquist, W. I. & Finch, J. T. Image reconstructions of helical assemblies of the HIV-1 CA protein. Nature 407, 409–413 (2000)

    Article  ADS  CAS  Google Scholar 

  18. Mayo, K. et al. Analysis of Rous sarcoma virus capsid protein variants assembled on lipid monolayers. J. Mol. Biol. 316, 667–678 (2002)

    Article  CAS  Google Scholar 

  19. Ganser, B. K., Cheng, A., Sundquist, W. I. & Yeager, M. Three-dimensional structure of the M-MuLV CA protein on a lipid monolayer: a general model for retroviral capsid assembly. EMBO J. 22, 2886–2892 (2003)

    Article  CAS  Google Scholar 

  20. Mayo, K. et al. Retrovirus capsid protein assembly arrangements. J. Mol. Biol. 325, 225–237 (2003)

    Article  CAS  Google Scholar 

  21. Ganser-Pornillos, B. K., Cheng, A. & Yeager, M. Structure of full-length HIV-1 CA: a model for the mature capsid lattice. Cell 131, 70–79 (2007)

    Article  CAS  Google Scholar 

  22. Ganser, B. K., Li, S., Klishko, V. Y., Finch, J. T. & Sundquist, W. I. Assembly and analysis of conical models for the HIV-1 core. Science 283, 80–83 (1999)

    Article  ADS  CAS  Google Scholar 

  23. Caspar, D. L. & Klug, A. Physical principles in the construction of regular viruses. Cold Spring Harb. Symp. Quant. Biol. 27, 1–24 (1962)

    Article  CAS  Google Scholar 

  24. Ganser-Pornillos, B. K., von Schwedler, U. K., Stray, K. M., Aiken, C. & Sundquist, W. I. Assembly properties of the human immunodeficiency virus type 1 CA protein. J. Virol. 78, 2545–2552 (2004)

    Article  CAS  Google Scholar 

  25. Heymann, J. B., Butan, C., Winkler, D. C., Craven, R. C. & Steven, A. C. Irregular and semi-regular polyhedral models for Rous sarcoma virus cores. Comput. Math. Meth. Medicine 9, 197–210 (2008)

    Article  MathSciNet  Google Scholar 

  26. Purdy, J. G., Flanagan, J. M., Ropson, I. J., Rennoll-Bankert, K. E. & Craven, R. C. Critical role of conserved hydrophobic residues within the major homology region in mature retroviral capsid assembly. J. Virol. 82, 5951–5961 (2008)

    Article  CAS  Google Scholar 

  27. von Schwedler, U. K. et al. Proteolytic refolding of the HIV-1 capsid protein amino-terminus facilitates viral core assembly. EMBO J. 17, 1555–1568 (1998)

    Article  CAS  Google Scholar 

  28. Lanman, J. et al. Identification of novel interactions in HIV-1 capsid protein assembly by high-resolution mass spectrometry. J. Mol. Biol. 325, 759–772 (2003)

    Article  CAS  Google Scholar 

  29. Briggs, J. A. et al. The stoichiometry of Gag protein in HIV-1. Nature Struct. Mol. Biol. 11, 672–675 (2004)

    Article  CAS  Google Scholar 

  30. Bowzard, J. B., Wills, J. W. & Craven, R. C. Second-site suppressors of Rous sarcoma virus CA mutations: evidence for interdomain interactions. J. Virol. 75, 6850–6856 (2001)

    Article  CAS  Google Scholar 

  31. Lokhandwala, P. M., Nguyen, T. L., Bowzard, J. B. & Craven, R. C. Cooperative role of the MHR and the CA dimerization helix in the maturation of the functional retrovirus capsid. Virology 376, 191–198 (2008)

    Article  CAS  Google Scholar 

  32. Edeling, M. A., Smith, C. & Owen, D. Life of a clathrin coat: insights from clathrin and AP structures. Nature Rev. Mol. Cell Biol. 7, 32–44 (2006)

    Article  CAS  Google Scholar 

  33. Heymann, J. B. & Belnap, D. M. Bsoft: Image processing and molecular modeling for electron microscopy. J. Struct. Biol. 157, 3–18 (2007)

    Article  CAS  Google Scholar 

  34. Ludtke, S. J., Baldwin, P. R. & Chiu, W. EMAN: Semiautomated software for high-resolution single-particle reconstructions. J. Struct. Biol. 128, 82–97 (1999)

    Article  CAS  Google Scholar 

  35. Cantele, F., Lanzavecchia, S. & Bellon, P. L. The variance of icosahedral virus models is a key indicator in the structure determination: a model-free reconstruction of viruses, suitable for refractory particles. J. Struct. Biol. 141, 84–92 (2003)

    Article  Google Scholar 

  36. Bubeck, D. et al. Structure of the poliovirus 135S cell-entry intermediate at 10Å resolution reveals the location of an externalized polypeptide that binds to membranes. J. Virol. 79, 7745–7755 (2005)

    Article  CAS  Google Scholar 

  37. Pettersen, E. F. et al. UCSF Chimera—a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004)

    Article  CAS  Google Scholar 

  38. Chacon, P. & Wriggers, W. Multi-resolution contour-based fitting of macromolecular structures. J. Mol. Biol. 317, 375–384 (2002)

    Article  CAS  Google Scholar 

  39. Cheng, N. et al. Handedness of the herpes simplex virus capsid and procapsid. J. Virol. 76, 7855–7859 (2002)

    Article  CAS  Google Scholar 

  40. Chen, D. H., Song, J. L., Chuang, D. T., Chiu, W. & Ludtke, S. J. An expanded conformation of single-ring GroEL–GroES complex encapsulates an 86 kDa substrate. Structure 14, 1711–1722 (2006)

    Article  CAS  Google Scholar 

  41. Saxton, W. O. & Baumeister, W. The correlation averaging of a regularly arranged bacterial cell envelope protein. J. Microsc. 127, 127–138 (1982)

    Article  CAS  Google Scholar 

  42. Tang, C., Ndassa, Y. & Summers, M. F. Structure of the N-terminal 283-residue fragment of the immature HIV-1 Gag polyprotein. Nature Struct. Biol. 9, 537–543 (2002)

    CAS  Google Scholar 

  43. Worthylake, D. K., Wang, H., Yoo, S., Sundquist, W. I. & Hill, C. P. Structures of the HIV-1 capsid protein dimerization domain at 2.6 Å resolution. Acta Crystallogr. D 55, 85–92 (1999)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank J. Flanagan for advice on protein purification and analytic methods and access to equipment, R. Meyers for assistance in electron microscopy at the Penn State College of Medicine and B. Heymann for advice on data analysis. This work was supported by the Intramural Research Program of NIAMS and the IATAP Program (A.C.S.), and funding from NIH grant CA100322, the Pennsylvania Department of Health and the Penn State Cancer Institute (R.C.C.).

Author Contributions A.C.S. and R.C.C. designed the project; J.G.P. prepared the capsids with guidance from R.C.C.; N.C. performed the cryo-electron microscopy; G.C. performed the image reconstruction and modelling; and A.C.S. and G.C. wrote the paper with input from the other authors.

Author information

Authors and Affiliations

  1. Laboratory of Structural Biology, National Institute for Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA ,

    Giovanni Cardone, Naiqian Cheng & Alasdair C. Steven

  2. Department of Microbiology and Immunology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA,

    John G. Purdy & Rebecca C. Craven

Authors
  1. Giovanni Cardone
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. John G. Purdy
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Naiqian Cheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Rebecca C. Craven
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Alasdair C. Steven
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Rebecca C. Craven or Alasdair C. Steven.

Supplementary information

Supplementary Figures

This file contains Supplementary Figures 1-5 with Legends (PDF 3488 kb)

PowerPoint slides

Rights and permissions

Reprints and permissions

About this article

Cite this article

Cardone, G., Purdy, J., Cheng, N. et al. Visualization of a missing link in retrovirus capsid assembly. Nature 457, 694–698 (2009). https://doi.org/10.1038/nature07724

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature07724

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

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