Letter | Published:

Structural basis for retroviral integration into nucleosomes

Nature volume 523, pages 366369 (16 July 2015) | Download Citation

Abstract

Retroviral integration is catalysed by a tetramer of integrase (IN) assembled on viral DNA ends in a stable complex, known as the intasome1,2. How the intasome interfaces with chromosomal DNA, which exists in the form of nucleosomal arrays, is currently unknown. Here we show that the prototype foamy virus (PFV) intasome is proficient at stable capture of nucleosomes as targets for integration. Single-particle cryo-electron microscopy reveals a multivalent intasome–nucleosome interface involving both gyres of nucleosomal DNA and one H2A–H2B heterodimer. While the histone octamer remains intact, the DNA is lifted from the surface of the H2A–H2B heterodimer to allow integration at strongly preferred superhelix location ±3.5 positions. Amino acid substitutions disrupting these contacts impinge on the ability of the intasome to engage nucleosomes in vitro and redistribute viral integration sites on the genomic scale. Our findings elucidate the molecular basis for nucleosome capture by the viral DNA recombination machinery and the underlying nucleosome plasticity that allows integration.

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Accessions

Primary accessions

Electron Microscopy Data Bank

Gene Expression Omnibus

Data deposits

The cryo-EM electron density map has been deposited in the Electron Microscopy Data Bank under accession number EMD-2992. Integration sites have been deposited in the NCBI Gene Expression Omnibus under accession number GSE67730.

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Acknowledgements

This work was supported by the European Union FP7 HIVINNOV consortium grant 305137 (to P.C.), the US National Institute of General Medical Sciences P50 grant GM082251-06 (to P.C.) and the US National Institutes of Health R01 grant AI070042-08 (to A.N.E.). Data collection was in part funded by the Netherlands Centre for Electron Nanoscopy (NeCEN) by grants from the Nederlandse Organisatie voor Wetenschappelijk Onderzoek (project 175.010.2009.001) and by the European Union’s Regional Development Fund through ‘Kansen voor West’ (project 21Z.014). We would like to thank L. Collinson, R. Carzaniga and Kirsty MacLellan-Gibson for EM access, R. Horton-Harpin for provision of HeLa cell pellets and assistance with tissue culture. We also thank F. Santoni, N. Sweeny and all our colleagues for helpful discussions.

Author information

Author notes

    • Stephen Hare

    Present address: Division of Molecular Biosciences, Imperial College London, South Kensington Campus, London SW7 2AZ, UK

Affiliations

  1. Chromatin Structure and Mobile DNA, The Francis Crick Institute, Blanche Lane, South Mimms EN6 3LD, UK

    • Daniel P. Maskell
    • , Paul Lesbats
    •  & Peter Cherepanov
  2. Architecture and Dynamics of Macromolecular Machines, Clare Hall Laboratories, The Francis Crick Institute, Blanche Lane, South Mimms EN6 3LD, UK

    • Ludovic Renault
    •  & Alessandro Costa
  3. National Institute for Biological Standards and Control, Microscopy and Imaging, Blanche Lane, South Mimms EN6 3QG, UK

    • Ludovic Renault
  4. Department of Cancer Immunology and AIDS, Dana-Farber Cancer Institute, 450 Brookline Avenue, Boston, Massachusetts 02215, USA

    • Erik Serrao
    •  & Alan N. Engelman
  5. NeCEN, Gorlaeus Laboratory, Einsteinweg 55, Leiden, 2333, the Netherlands

    • Rishi Matadeen
  6. Division of Medicine, Imperial College London, St-Mary’s Campus, Norfolk Place, London W2 1PG, UK

    • Stephen Hare
    •  & Peter Cherepanov
  7. Institute of Virology, Technische Universität Dresden, Fetscherstr. 74, Dresden 01307, Germany

    • Dirk Lindemann

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Contributions

D.P.M. analysed interactions of the PFV intasome and nucleosomes, discovered conditions to produce the stable intasome–nucleosome complex and prepared it for EM; L.R., D.P.M. and A.C. performed all EM work with the exception of cryo-EM grid preparation and screening, which was performed by L.R.; R.M. collected cryo-EM data; L.R. and A.C. determined the structure. S.H. designed the co-dependent K120E–D273K PFV IN pair; D.L. designed and provided wild-type PFV vector constructs; P.C. cloned PFV vector mutants; P.C. and P.L. carried out PFV infections; E.S. and A.N.E. developed the protocol and carried out sequencing of PFV integration sites; P.C. analysed integration site distributions.

Corresponding authors

Correspondence to Alessandro Costa or Peter Cherepanov.

Extended data

Supplementary information

Videos

  1. 1.

    The PFV intasome - nucleosome complex at 7.8 Å resolution

    This video displays the overall structure, intasome-nucleosome interface and the key components of the complex.

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DOI

https://doi.org/10.1038/nature14495

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