This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Hameau, L.
Right arrow Articles by Mirambeau, G.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Hameau, L.
Right arrow Articles by Mirambeau, G.

 Previous Article  |  Next Article 

Journal of Virology, April 2001, p. 3301-3313, Vol. 75, No. 7
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.7.3301-3313.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Human Immunodeficiency Virus Type 1 Central DNA Flap: Dynamic Terminal Product of Plus-Strand Displacement DNA Synthesis Catalyzed by Reverse Transcriptase Assisted by Nucleocapsid Protein

Laurence Hameau,1 Josette Jeusset,1 Sophie Lafosse,1 Dominique Coulaud,1 Etienne Delain,1 Torsten Unge,2 Tobias Restle,3 Eric Le Cam,1 and Gilles Mirambeau1,*

Laboratoire de Microscopie Moléculaire et Cellulaire, CNRS UMR 8532, Institut Gustave Roussy, 94805 Villejuif Cedex, France1; Department of Molecular Biology, University of Uppsala, S-111 11 Uppsala, Sweden2; and Abteilung Physikalische Biochemie, Max-Planck-Institut für Molekulare Physiologie, 44227 Dortmund, Germany3

Received 5 October 2000/Accepted 20 December 2000

To terminate the reverse transcription of the human immunodeficiency virus type 1 (HIV-1) genome, a final step occurs within the center of the proviral DNA generating a 99-nucleotide DNA flap (6). This step, catalyzed by reverse transcriptase (RT), is defined as a discrete strand displacement (SD) synthesis between the first nucleotide after the central priming (cPPT) site and the final position of the central termination sequence (CTS) site. Using recombinant HIV-1 RT and a circular single-stranded DNA template harboring the cPPT-CTS sequence, we have developed an SD synthesis-directed in vitro termination assay. Elongation, strand displacement, and complete central flap behavior were analyzed using electrophoresis and electron microscopy approaches. Optimal conditions to obtain complete central flap, which ended at the CTS site, have been defined in using nucleocapsid protein (NCp), the main accessory protein of the reverse transcription complex. A full-length HIV-1 central DNA flap was then carried out in vitro. Its synthesis appears faster in the presence of the HIV-1 NCp or the T4-encoded SSB protein (gp32). Finally, a high frequency of strand transfer was shown during the SD synthesis along the cPPT-CTS site with RT alone. This reveals a local and efficient 3'-5' branch migration which emphasizes some important structural fluctuations within the flap. These fluctuations may be stabilized by the NCp chaperone activity. The biological implications of the RT-directed NCp-assisted flap synthesis are discussed within the context of reverse transcription complexes, assembly of the preintegration complexes, and nuclear import of the HIV-1 proviral DNA to the nucleus toward their chromatin targets.

* Corresponding author. Mailing address: Laboratoire de Microscopie Moléculaire et Cellulaire, CNRS UMR 8532, Institut Gustave Roussy, 94805 Villejuif Cedex, France. Phone: 331-42-11-48-80. Fax: 331-42-11-52-76. E-mail: mirambe{at}igr.fr.

Journal of Virology, April 2001, p. 3301-3313, Vol. 75, No. 7
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.7.3301-3313.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Jang, S. I., Kim, Y. H., Paik, S. Y., You, J. C. (2007). Development of a Cell-Based Assay Probing the Specific Interaction between the Human Immunodeficiency Virus Type 1 Nucleocapsid and Psi RNA In Vivo. J. Virol. 81: 6151-6155 [Abstract] [Full Text]  
  • Hamon, L., Pastre, D., Dupaigne, P., Breton, C. L., Cam, E. L., Pietrement, O. (2007). High-resolution AFM imaging of single-stranded DNA-binding (SSB) protein--DNA complexes. Nucleic Acids Res 35: e58-e58 [Abstract] [Full Text]  
  • Wurtzer, S., Goubard, A., Mammano, F., Saragosti, S., Lecossier, D., Hance, A. J., Clavel, F. (2006). Functional Central Polypurine Tract Provides Downstream Protection of the Human Immunodeficiency Virus Type 1 Genome from Editing by APOBEC3G and APOBEC3B.. J. Virol. 80: 3679-3683 [Abstract] [Full Text]  
  • Kankia, B. I., Barany, G., Musier-Forsyth, K. (2005). Unfolding of DNA quadruplexes induced by HIV-1 nucleocapsid protein. Nucleic Acids Res 33: 4395-4403 [Abstract] [Full Text]  
  • Lanciault, C., Champoux, J. J. (2004). Single Unpaired Nucleotides Facilitate HIV-1 Reverse Transcriptase Displacement Synthesis through Duplex RNA. J. Biol. Chem. 279: 32252-32261 [Abstract] [Full Text]  
  • Lyonnais, S., Gorelick, R. J., Mergny, J.-L., Le Cam, E., Mirambeau, G. (2003). G-quartets direct assembly of HIV-1 nucleocapsid protein along single-stranded DNA. Nucleic Acids Res 31: 5754-5763 [Abstract] [Full Text]  
  • Lyonnais, S., Hounsou, C., Teulade-Fichou, M.-P., Jeusset, J., Cam, E. L., Mirambeau, G. (2002). G-quartets assembly within a G-rich DNA flap. A possible event at the center of the HIV-1 genome. Nucleic Acids Res 30: 5276-5283 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»