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 Van Maele, B.
Right arrow Articles by Debyser, Z.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Van Maele, B.
Right arrow Articles by Debyser, Z.

 Previous Article  |  Next Article 

Journal of Virology, April 2003, p. 4685-4694, Vol. 77, No. 8
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.8.4685-4694.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Impact of the Central Polypurine Tract on the Kinetics of Human Immunodeficiency Virus Type 1 Vector Transduction

Bénédicte Van Maele, Jan De Rijck, Erik De Clercq, and Zeger Debyser*

Rega Institute for Medical Research, Leuven, Flanders, Belgium

Received 28 October 2002/ Accepted 16 January 2003

Lentiviral vectors derived from human immunodeficiency virus type 1 (HIV-1) show great promise as gene carriers for future gene therapy. Insertion of a fragment containing the central polypurine tract (cPPT) in HIV-1 vector constructs is known to enhance transduction efficiency drastically, reportedly by facilitating the nuclear import of HIV-1 cDNA through a central DNA flap. We have studied the impact of the cPPT on the kinetics of HIV-1 vector transduction by real-time PCR. The kinetics of total HIV-1 DNA, two-long-terminal-repeat (2-LTR) circles, and, by an Alu-PCR, integrated proviral DNA were monitored. About 6 to 12 h after transduction, the total HIV-1 DNA reached a maximum level, followed by a steep decrease. The 2-LTR circles peaked after 24 to 48 h and were diluted upon cell division. Integration of HIV-1 DNA was first detected at 12 h postinfection. When HIV-1 vectors that contained the cPPT were used, DNA synthesis was similar but a threefold higher amount of 2-LTR circles was detected, confirming the impact on nuclear import. Moreover, a 10-fold increase in the amount of integrated DNA was observed in the presence of the cPPT. Only in the absence of the cPPT was a saturation in 2-LTR circle formation seen at a high multiplicity of infection, suggesting a role for the cPPT in overcoming a barrier to the nuclear import of HIV-1 DNA. A major effect of the central DNA flap on the juxtaposition of both LTRs is unlikely, since transduction with HIV-1 vectors containing ectopic cPPT fragments resulted in increased amounts of 2-LTR circles as well as integrated DNA. Inhibitors of transduction by cPPT-containing HIV vectors were also studied by real-time PCR. The reverse transcriptase inhibitor azidothymidine (AZT) and the nonnucleoside reverse transcriptase inhibitor {alpha}-APA clearly inhibited viral DNA synthesis, whereas integrase inhibitors such as the diketo acid L-708,906 and the pyranodipyrimidine V-165 specifically inhibited integration.

* Corresponding author. Mailing address: Rega Institute for Medical Research, Minderbroedersstraat 10, B-3000 Leuven, Flanders, Belgium. Phone: 32 16 33 21 83. Fax: 32 16 33 21 31. E-mail: zeger.debyser{at}uz.kuleuven.ac.be.

Journal of Virology, April 2003, p. 4685-4694, Vol. 77, No. 8
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.8.4685-4694.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Arfi, V., Lienard, J., Nguyen, X.-N., Berger, G., Rigal, D., Darlix, J.-L., Cimarelli, A. (2009). Characterization of the Behavior of Functional Viral Genomes during the Early Steps of Human Immunodeficiency Virus Type 1 Infection. J. Virol. 83: 7524-7535 [Abstract] [Full Text]  
  • Wiktorowicz, T., Peters, K., Armbruster, N., Steinert, A. F., Rethwilm, A. (2009). Generation of an improved foamy virus vector by dissection of cis-acting sequences. J. Gen. Virol. 90: 481-487 [Abstract] [Full Text]  
  • Pizzato, M., Popova, E., Gottlinger, H. G. (2008). Nef Can Enhance the Infectivity of Receptor-Pseudotyped Human Immunodeficiency Virus Type 1 Particles. J. Virol. 82: 10811-10819 [Abstract] [Full Text]  
  • Hombrouck, A., Van Remoortel, B., Michiels, M., Noppe, W., Christ, F., Eneroth, A., Sahlberg, B. L., Benkestock, K., Vrang, L., Johansson, N. G., Barreca, M. L., De Luca, L., Ferro, S., Chimirri, A., Debyser, Z., Witvrouw, M. (2008). Preclinical Evaluation of 1H-Benzylindole Derivatives as Novel Human Immunodeficiency Virus Integrase Strand Transfer Inhibitors. Antimicrob. Agents Chemother. 52: 2861-2869 [Abstract] [Full Text]  
  • Skasko, M., Kim, B. (2008). Compensatory Role of Human Immunodeficiency Virus Central Polypurine Tract Sequence in Kinetically Disrupted Reverse Transcription. J. Virol. 82: 7716-7720 [Abstract] [Full Text]  
  • Park, F. (2007). Lentiviral vectors: are they the future of animal transgenesis?. Physiol. Genomics 31: 159-173 [Abstract] [Full Text]  
  • Hombrouck, A., Hantson, A., van Remoortel, B., Michiels, M., Vercammen, J., Rhodes, D., Tetz, V., Engelborghs, Y., Christ, F., Debyser, Z., Witvrouw, M. (2007). Selection of human immunodeficiency virus type 1 resistance against the pyranodipyrimidine V-165 points to a multimodal mechanism of action. J Antimicrob Chemother 59: 1084-1095 [Abstract] [Full Text]  
  • Marsden, M. D., Zack, J. A. (2007). Human Immunodeficiency Virus Bearing a Disrupted Central DNA Flap Is Pathogenic In Vivo. J. Virol. 81: 6146-6150 [Abstract] [Full Text]  
  • Goffinet, C., Allespach, I., Keppler, O. T. (2007). HIV-susceptible transgenic rats allow rapid preclinical testing of antiviral compounds targeting virus entry or reverse transcription. Proc. Natl. Acad. Sci. USA 104: 1015-1020 [Abstract] [Full Text]  
  • De Rijck, J., Vandekerckhove, L., Gijsbers, R., Hombrouck, A., Hendrix, J., Vercammen, J., Engelborghs, Y., Christ, F., Debyser, Z. (2006). Overexpression of the Lens Epithelium-Derived Growth Factor/p75 Integrase Binding Domain Inhibits Human Immunodeficiency Virus Replication. J. Virol. 80: 11498-11509 [Abstract] [Full Text]  
  • Bona, R., Andreotti, M., Buffa, V., Leone, P., Galluzzo, C. M., Amici, R., Palmisano, L., Mancini, M. G., Michelini, Z., Di Santo, R., Costi, R., Roux, A., Pommier, Y., Marchand, C., Vella, S., Cara, A. (2006). Development of a human immunodeficiency virus vector-based, single-cycle assay for evaluation of anti-integrase compounds.. Antimicrob. Agents Chemother. 50: 3407-3417 [Abstract] [Full Text]  
  • Arhel, N., Munier, S., Souque, P., Mollier, K., Charneau, P. (2006). Nuclear import defect of human immunodeficiency virus type 1 DNA flap mutants is not dependent on the viral strain or target cell type.. J. Virol. 80: 10262-10269 [Abstract] [Full Text]  
  • Jamburuthugoda, V. K., Chugh, P., Kim, B. (2006). Modification of Human Immunodeficiency Virus Type 1 Reverse Transcriptase to Target Cells with Elevated Cellular dNTP Concentrations. J. Biol. Chem. 281: 13388-13395 [Abstract] [Full Text]  
  • Vandekerckhove, L., Christ, F., Van Maele, B., De Rijck, J., Gijsbers, R., Van den Haute, C., Witvrouw, M., Debyser, Z. (2006). Transient and Stable Knockdown of the Integrase Cofactor LEDGF/p75 Reveals Its Role in the Replication Cycle of Human Immunodeficiency Virus. J. Virol. 80: 1886-1896 [Abstract] [Full Text]  
  • Haffar, O., Dubrovsky, L., Lowe, R., Berro, R., Kashanchi, F., Godden, J., Vanpouille, C., Bajorath, J., Bukrinsky, M. (2005). Oxadiazols: a New Class of Rationally Designed Anti-Human Immunodeficiency Virus Compounds Targeting the Nuclear Localization Signal of the Viral Matrix Protein. J. Virol. 79: 13028-13036 [Abstract] [Full Text]  
  • Sancho, R., Marquez, N., Gomez-Gonzalo, M., Calzado, M. A., Bettoni, G., Coiras, M. T., Alcami, J., Lopez-Cabrera, M., Appendino, G., Munoz, E. (2004). Imperatorin Inhibits HIV-1 Replication through an Sp1-dependent Pathway. J. Biol. Chem. 279: 37349-37359 [Abstract] [Full Text]  
  • Ao, Z., Yao, X., Cohen, E. A. (2004). Assessment of the Role of the Central DNA Flap in Human Immunodeficiency Virus Type 1 Replication by Using a Single-Cycle Replication System. J. Virol. 78: 3170-3177 [Abstract] [Full Text]  
  • Maertens, G., Cherepanov, P., Pluymers, W., Busschots, K., De Clercq, E., Debyser, Z., Engelborghs, Y. (2003). LEDGF/p75 Is Essential for Nuclear and Chromosomal Targeting of HIV-1 Integrase in Human Cells. J. Biol. Chem. 278: 33528-33539 [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»