Letter | Published:

Structure and mutagenesis reveal essential capsid protein interactions for KSHV replication

Nature volume 553, pages 521525 (25 January 2018) | Download Citation

Abstract

Kaposi’s sarcoma-associated herpesvirus (KSHV) causes Kaposi’s sarcoma1,2, a cancer that commonly affects patients with AIDS3 and which is endemic in sub-Saharan Africa4. The KSHV capsid is highly pressurized by its double-stranded DNA genome, as are the capsids of the eight other human herpesviruses5. Capsid assembly and genome packaging of herpesviruses are prone to interruption6,7,8,9 and can therefore be targeted for the structure-guided development of antiviral agents. However, herpesvirus capsids—comprising nearly 3,000 proteins and over 1,300 Å in diameter—present a formidable challenge to atomic structure determination10 and functional mapping of molecular interactions. Here we report a 4.2 Å resolution structure of the KSHV capsid, determined by electron-counting cryo-electron microscopy, and its atomic model, which contains 46 unique conformers of the major capsid protein (MCP), the smallest capsid protein (SCP) and the triplex proteins Tri1 and Tri2. Our structure and mutagenesis results reveal a groove in the upper domain of the MCP that contains hydrophobic residues that interact with the SCP, which in turn crosslinks with neighbouring MCPs in the same hexon to stabilize the capsid. Multiple levels of MCP–MCP interaction—including six sets of stacked hairpins lining the hexon channel, disulfide bonds across channel and buttress domains in neighbouring MCPs, and an interaction network forged by the N-lasso domain and secured by the dimerization domain—define a robust capsid that is resistant to the pressure exerted by the enclosed genome. The triplexes, each composed of two Tri2 molecules and a Tri1 molecule, anchor to the capsid floor via a Tri1 N-anchor to plug holes in the MCP network and rivet the capsid floor. These essential roles of the MCP N-lasso and Tri1 N-anchor are verified by serial-truncation mutageneses. Our proof-of-concept demonstration of the use of polypeptides that mimic the smallest capsid protein to inhibit KSHV lytic replication highlights the potential for exploiting the interaction hotspots revealed in our atomic structure to develop antiviral agents.

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Acknowledgements

This project was supported in part by grants from the National Institutes of Health (NIH) (DE025567, GM071940, AI094386, CA091791 and CA177322) and indirectly through a Clinical and Translational Science Institute core voucher award (UL1TR000124) from UCLA’s National Center for Advancing Translational Science. We acknowledge the use of instruments at the Electron Imaging Center for Nanomachines supported by UCLA and by instrumentation grants from the NIH (1S10OD018111, 1U24GM116792) and NSF (DBI-1338135, DMR-1548924).

Author information

Author notes

    • Xinghong Dai
    •  & Danyang Gong

    These authors contributed equally to this work.

Affiliations

  1. Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles (UCLA), Los Angeles, California 90095, USA

    • Xinghong Dai
    • , Hanyoung Lim
    • , Jonathan Jih
    •  & Z. Hong Zhou
  2. The California NanoSystems Institute (CNSI), University of California, Los Angeles (UCLA), Los Angeles, California 90095, USA

    • Xinghong Dai
    • , Ren Sun
    •  & Z. Hong Zhou
  3. Department of Molecular and Medical Pharmacology, University of California, Los Angeles (UCLA), Los Angeles, California 90095, USA

    • Xinghong Dai
    • , Danyang Gong
    • , Ting-Ting Wu
    •  & Ren Sun

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Contributions

Z.H.Z., X.D., D.G. and R.S. designed the project; Z.H.Z., R.S. and T.-T.W. supervised research; D.G. and X.D. prepared the samples; X.D. acquired cryo-EM data and determined the structure; X.D., H.L. and J.J. built atomic models; D.G. performed functional studies; X.D., D.G., Z.H.Z., R.S. and T.-T.W. interpreted the results; Z.H.Z., X.D. and D.G. wrote the paper; R.S. revised the paper; and all authors reviewed the paper.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Ren Sun or Z. Hong Zhou.

Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Extended data

Supplementary information

PDF files

  1. 1.

    Life Sciences Reporting Summary

Videos

  1. 1.

    Overall structure and high resolution features of the KSHV capsid reconstruction

    This video shows the overall structure and high resolution features of the KSHV capsid reconstruction.

  2. 2.

    Structure of a hexon MCP

    The density map of a hexon MCP was segmented out from the 3D reconstruction, displayed as semitransparent grey surface, and superimposed with its atomic model (ribbon).

  3. 3.

    Structure difference between hexon MCP and penton MCP

    This video shows the structural difference between hexon MCP and penton MCP.

  4. 4.

    Densities around a hexon MCP N-lasso region

    Densities around hexon MCP E1 N-lasso (cyan), which lashes the N-arm of MCP C4 (gold) and E-loop of MCP C5 (magenta), were segmented out and superimposed with the corresponding atomic models.

  5. 5.

    MCP network interactions in the capsid floor

    MCP network interactions in the capsid floor.

  6. 6.

    Structure of a triplex

    The density map of triplex Tc was segmented out from the 3D reconstruction, displayed as semitransparent grey surface, and superimposed with its atomic model (ribbon).

  7. 7.

    Structure difference between Tri2A and Tri2B

    This video shows the structural difference between Tri2A and Tri2B.

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DOI

https://doi.org/10.1038/nature25438

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