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 Balakrishnan, M.
Right arrow Articles by Bambara, R. A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Balakrishnan, M.
Right arrow Articles by Bambara, R. A.

 Previous Article  |  Next Article 

Journal of Virology, April 2003, p. 4710-4721, Vol. 77, No. 8
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.8.4710-4721.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Template Dimerization Promotes an Acceptor Invasion-Induced Transfer Mechanism during Human Immunodeficiency Virus Type 1 Minus-Strand Synthesis

Mini Balakrishnan,1 Bernard P. Roques,2 Philip J. Fay,1 and Robert A. Bambara1,3*

Department of Biochemistry and Biophysics,1 the Cancer Center, University of Rochester Medical Center, Rochester, New York 14642,3 Departement de Pharmacochimie Moleculaire et Structurale, U266 INSERM, URA D1500 CNRS, UER des Sciences Pharmaceutiques et Biologiques, 75270 Paris Cedex 06, France2

Received 31 October 2002/ Accepted 20 January 2003

The biochemical mechanism of template switching by human immunodeficiency virus type 1 (HIV-1) reverse transcriptase and the role of template dimerization were examined. Homologous donor-acceptor template pairs derived from the HIV-1 untranslated leader region and containing the wild-type and mutant dimerization initiation sequences (DIS) were used to examine the efficiency and distribution of transfers. Inhibiting donor-acceptor interaction was sufficient to reduce transfers in DIS-containing template pairs, indicating that template dimerization, and not the mere presence of the DIS, promotes efficient transfers. Additionally, we show evidence that the overall transfer process spans an extended region of the template and proceeds through a two-step mechanism. Transfer is initiated through an RNase H-facilitated acceptor invasion step, while synthesis continues on the donor template. The invasion then propagates towards the primer terminus by branch migration. Transfer is completed with the translocation of the primer terminus at a site distant from the invasion point. In our system, most invasions initiated before synthesis reached the DIS. However, transfer of the primer terminus predominantly occurred after synthesis through the DIS. The two steps were separated by 60 to 80 nucleotides. Sequence markers revealed the position of primer terminus switch, whereas DNA oligomers designed to block acceptor-cDNA interactions defined sites of invasion. Within the region of homology, certain positions on the template were inherently more favorable for invasion than others. In templates with DIS, the proximity of the acceptor facilitates invasion, thereby enhancing transfer efficiency. Nucleocapsid protein enhanced the overall efficiency of transfers but did not alter the mechanism.

* Corresponding author. Mailing address: Department of Biochemistry and Biophysics, Box 712, University of Rochester Medical Center, 601 Elmwood Ave., Rochester, NY 14642. Phone: (585) 275-2764. Fax: (585) 271-2683. E-mail: robert_bambara{at}urmc.rochester.edu.

Journal of Virology, April 2003, p. 4710-4721, Vol. 77, No. 8
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.8.4710-4721.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Galli, A., Lai, A., Corvasce, S., Saladini, F., Riva, C., Deho, L., Caramma, I., Franzetti, M., Romano, L., Galli, M., Zazzi, M., Balotta, C. (2008). Recombination analysis and structure prediction show correlation between breakpoint clusters and RNA hairpins in the pol gene of human immunodeficiency virus type 1 unique recombinant forms. J. Gen. Virol. 89: 3119-3125 [Abstract] [Full Text]  
  • Song, M., Basu, V. P., Hanson, M. N., Roques, B. P., Bambara, R. A. (2008). Proximity and Branch Migration Mechanisms in HIV-1 Minus Strand Strong Stop DNA Transfer. J. Biol. Chem. 283: 3141-3150 [Abstract] [Full Text]  
  • Sinck, L., Richer, D., Howard, J., Alexander, M., Purcell, D. F.J., Marquet, R., Paillart, J.-C. (2007). In vitro dimerization of human immunodeficiency virus type 1 (HIV-1) spliced RNAs. RNA 13: 2141-2150 [Abstract] [Full Text]  
  • Purohit, V., Roques, B. P., Kim, B., Bambara, R. A. (2007). Mechanisms That Prevent Template Inactivation by HIV-1 Reverse Transcriptase RNase H Cleavages. J. Biol. Chem. 282: 12598-12609 [Abstract] [Full Text]  
  • Wu, H.-Y., Brian, D. A. (2007). 5'-Proximal Hot Spot for an Inducible Positive-to-Negative-Strand Template Switch by Coronavirus RNA-Dependent RNA Polymerase. J. Virol. 81: 3206-3215 [Abstract] [Full Text]  
  • Gao, L., Balakrishnan, M., Roques, B. P., Bambara, R. A. (2007). Insights into the Multiple Roles of Pausing in HIV-1 Reverse Transcriptase-promoted Strand Transfers. J. Biol. Chem. 282: 6222-6231 [Abstract] [Full Text]  
  • Operario, D. J., Balakrishnan, M., Bambara, R. A., Kim, B. (2006). Reduced dNTP Interaction of Human Immunodeficiency Virus Type 1 Reverse Transcriptase Promotes Strand Transfer. J. Biol. Chem. 281: 32113-32121 [Abstract] [Full Text]  
  • Lanciault, C., Champoux, J. J. (2006). Pausing during Reverse Transcription Increases the Rate of Retroviral Recombination. J. Virol. 80: 2483-2494 [Abstract] [Full Text]  
  • Galetto, R., Giacomoni, V., Veron, M., Negroni, M. (2006). Dissection of a Circumscribed Recombination Hot Spot in HIV-1 after a Single Infectious Cycle. J. Biol. Chem. 281: 2711-2720 [Abstract] [Full Text]  
  • Buxton, P., Tachedjian, G., Mak, J. (2005). Analysis of the Contribution of Reverse Transcriptase and Integrase Proteins to Retroviral RNA Dimer Conformation. J. Virol. 79: 6338-6348 [Abstract] [Full Text]  
  • Rhodes, T. D., Nikolaitchik, O., Chen, J., Powell, D., Hu, W.-S. (2005). Genetic Recombination of Human Immunodeficiency Virus Type 1 in One Round of Viral Replication: Effects of Genetic Distance, Target Cells, Accessory Genes, and Lack of High Negative Interference in Crossover Events. J. Virol. 79: 1666-1677 [Abstract] [Full Text]  
  • Lanciault, C., Champoux, J. J. (2005). Effects of Unpaired Nucleotides within HIV-1 Genomic Secondary Structures on Pausing and Strand Transfer. J. Biol. Chem. 280: 2413-2423 [Abstract] [Full Text]  
  • Diamond, T. L., Roshal, M., Jamburuthugoda, V. K., Reynolds, H. M., Merriam, A. R., Lee, K. Y., Balakrishnan, M., Bambara, R. A., Planelles, V., Dewhurst, S., Kim, B. (2004). Macrophage Tropism of HIV-1 Depends on Efficient Cellular dNTP Utilization by Reverse Transcriptase. J. Biol. Chem. 279: 51545-51553 [Abstract] [Full Text]  
  • Galetto, R., Moumen, A., Giacomoni, V., Veron, M., Charneau, P., Negroni, M. (2004). The Structure of HIV-1 Genomic RNA in the gp120 Gene Determines a Recombination Hot Spot in Vivo. J. Biol. Chem. 279: 36625-36632 [Abstract] [Full Text]  
  • Nikolenko, G. N., Svarovskaia, E. S., Delviks, K. A., Pathak, V. K. (2004). Antiretroviral Drug Resistance Mutations in Human Immunodeficiency Virus Type 1 Reverse Transcriptase Increase Template-Switching Frequency. J. Virol. 78: 8761-8770 [Abstract] [Full Text]  
  • Andersen, E. S., Contera, S. A., Knudsen, B., Damgaard, C. K., Besenbacher, F., Kjems, J. (2004). Role of the Trans-activation Response Element in Dimerization of HIV-1 RNA. J. Biol. Chem. 279: 22243-22249 [Abstract] [Full Text]  
  • Moriarty, T. J., Marie-Egyptienne, D. T., Autexier, C. (2004). Functional Organization of Repeat Addition Processivity and DNA Synthesis Determinants in the Human Telomerase Multimer. Mol. Cell. Biol. 24: 3720-3733 [Abstract] [Full Text]  
  • Mikkelsen, J. G., Rasmussen, S. V., Pedersen, F. S. (2004). Complementarity-directed RNA dimer-linkage promotes retroviral recombination in vivo. Nucleic Acids Res 32: 102-114 [Abstract] [Full Text]  
  • Rhodes, T., Wargo, H., Hu, W.-S. (2003). High Rates of Human Immunodeficiency Virus Type 1 Recombination: Near-Random Segregation of Markers One Kilobase Apart in One Round of Viral Replication. J. Virol. 77: 11193-11200 [Abstract] [Full Text]  
  • Chen, Y., Balakrishnan, M., Roques, B. P., Bambara, R. A. (2003). Steps of the Acceptor Invasion Mechanism for HIV-1 Minus Strand Strong Stop Transfer. J. Biol. Chem. 278: 38368-38375 [Abstract] [Full Text]  
  • Roda, R. H., Balakrishnan, M., Hanson, M. N., Wohrl, B. M., Le Grice, S. F. J., Roques, B. P., Gorelick, R. J., Bambara, R. A. (2003). Role of the Reverse Transcriptase, Nucleocapsid Protein, and Template Structure in the Two-step Transfer Mechanism in Retroviral Recombination. J. Biol. Chem. 278: 31536-31546 [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»