Boireau, S.; Maiuri, P.; Basyuk, E.; De La Mata, M.; Knezevich, A.; Pradet-Balade, B.; Bäcker, V.; Kornblihtt, A.; Marcello, A.; Bertrand, E. "The transcriptional cycle of HIV-1 in real-time and live cells" (2007) Journal of Cell Biology. 179(2):291-304
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RNA polymerase II (RNAPII) is a fundamental enzyme, but few studies have analyzed its activity in living cells. Using human immunodeficiency virus (HIV) type 1 reporters, we study real-time messenger RNA (mRNA) biogenesis by photobleaching nascent RNAs and RNAPII at specific transcription sites. Through modeling, the use of mutant polymerases, drugs, and quantitative in situ hybridiza tion, we investigate the kinetics of the HIV-1 transcription cycle. Initiation appears efficient because most polymerases demonstrate stable gene association. We calculate an elongation rate of approximately 1.9 kb/min, and, surprisingly, polymerases remain at transcription sites 2.5 min longer than nascent RNAs. With a total polymerase residency time estimated at 333 s, 114 are assigned to elongation, and 63 are assigned to 3′-end processing and/or transcript release. However, mRNAs were released seconds after polyadenylation onset, and analysis of polymerase density by chromatin immunoprecipitation suggests that they pause or lose processivity after passing the polyA site. The strengths and limitations of this kinetic approach to analyze mRNA biogenesis in living cells are discussed. © The Rockefeller University Press.


Documento: Artículo
Título:The transcriptional cycle of HIV-1 in real-time and live cells
Autor:Boireau, S.; Maiuri, P.; Basyuk, E.; De La Mata, M.; Knezevich, A.; Pradet-Balade, B.; Bäcker, V.; Kornblihtt, A.; Marcello, A.; Bertrand, E.
Filiación:Institute of Molecular Genetics of Montpellier, Unité Mixte de Recherche 5535, 34293 Montpellier, France
Montpellier Rio Imaging, Centre National de la Recherche Scientifique, 34293 Montpellier, France
Laboratory of Molecular Virology, International Centre for Genetic Engineering and Biotechnology, 34012 Trieste, Italy
Laboratorio de Fisiología Y Biología Molecular, Departamento de Fisiología, Biología Molecular, Y Celular, Ciudad Universitaria, (C1428EHA) Buenos Aires, Argentina
Palabras clave:camptothecin; messenger RNA; RNA polymerase II; transactivator protein; virus RNA; article; biogenesis; chromatin immunoprecipitation; enzyme activity; fluorescence recovery after photobleaching; human; human cell; Human immunodeficiency virus 1; Human immunodeficiency virus 1 infection; in situ hybridization; kinetics; life cycle; nonhuman; nucleotide sequence; priority journal; promoter region; transcription initiation; virus cell interaction; virus transcription; Cell Line, Tumor; Cell Survival; Computer Simulation; Fluorescence Recovery After Photobleaching; Gene Expression Regulation, Viral; Genes, Reporter; HIV-1; Humans; In Situ Hybridization; Kinetics; Models, Genetic; Mutation; Photobleaching; Polyadenylation; RNA 3' End Processing; RNA Polymerase II; RNA, Messenger; RNA, Viral; Time Factors; Transcription, Genetic; Human immunodeficiency virus 1
Página de inicio:291
Página de fin:304
Título revista:Journal of Cell Biology
Título revista abreviado:J. Cell Biol.
CAS:camptothecin, 7689-03-4; RNA Polymerase II, EC 2.7.7.-; RNA, Messenger; RNA, Viral


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---------- APA ----------
Boireau, S., Maiuri, P., Basyuk, E., De La Mata, M., Knezevich, A., Pradet-Balade, B., Bäcker, V.,..., Bertrand, E. (2007) . The transcriptional cycle of HIV-1 in real-time and live cells. Journal of Cell Biology, 179(2), 291-304.
---------- CHICAGO ----------
Boireau, S., Maiuri, P., Basyuk, E., De La Mata, M., Knezevich, A., Pradet-Balade, B., et al. "The transcriptional cycle of HIV-1 in real-time and live cells" . Journal of Cell Biology 179, no. 2 (2007) : 291-304.
---------- MLA ----------
Boireau, S., Maiuri, P., Basyuk, E., De La Mata, M., Knezevich, A., Pradet-Balade, B., et al. "The transcriptional cycle of HIV-1 in real-time and live cells" . Journal of Cell Biology, vol. 179, no. 2, 2007, pp. 291-304.
---------- VANCOUVER ----------
Boireau, S., Maiuri, P., Basyuk, E., De La Mata, M., Knezevich, A., Pradet-Balade, B., et al. The transcriptional cycle of HIV-1 in real-time and live cells. J. Cell Biol. 2007;179(2):291-304.