In vivo dynamics of RNA polymerase II transcription.

@article{citeulike:1543646, abstract = {We imaged transcription in living cells using a locus-specific reporter system, which allowed precise, single-cell kinetic measurements of promoter binding, initiation and elongation. Photobleaching of fluorescent RNA polymerase II revealed several kinetically distinct populations of the enzyme interacting with a specific gene. Photobleaching and photoactivation of fluorescent MS2 proteins used to label nascent messenger RNAs provided sensitive elongation measurements. A mechanistic kinetic model that fits our data was validated using specific inhibitors. Polymerases elongated at 4.3 kilobases min(-1), much faster than previously documented, and entered a paused state for unexpectedly long times. Transcription onset was inefficient, with only 1\% of polymerase-gene interactions leading to completion of an mRNA. Our systems approach, quantifying both polymerase and mRNA kinetics on a defined DNA template in vivo with high temporal resolution, opens new avenues for studying regulation of transcriptional processes in vivo.}, address = {[1] Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA. [2] Laboratoire de G\'{e}n\'{e}tique Mol\'{e}culaire, Centre National de la Recherche Scientifique, UMR-8541, Ecole Normale Sup\'{e}rieure, 75005 Paris, France.}, author = {Darzacq, Xavier and Shav-Tal, Yaron and de Turris, Valeria and Brody, Yehuda and Shenoy, Shailesh M M. and Phair, Robert D D. and Singer, Robert H H. }, citeulike-article-id = {1543646}, doi = {10.1038/nsmb1280}, issn = {1545-9993}, journal = {Nat Struct Mol Biol}, keywords = {biophysics, rna, transcription, trp3}, month = {August}, posted-at = {2007-08-13 08:02:13}, priority = {4}, title = {In vivo dynamics of RNA polymerase II transcription.}, url = {http://dx.doi.org/10.1038/nsmb1280}, year = {2007} }

TY - JOUR ID - citeulike:1543646 L3 - citeulike-article-id:1543646 TI - In vivo dynamics of RNA polymerase II transcription. JF - Nat Struct Mol Biol AD - [1] Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA. [2] Laboratoire de Génétique Moléculaire, Centre National de la Recherche Scientifique, UMR-8541, Ecole Normale Supérieure, 75005 Paris, France. SN - 1545-9993 N2 - We imaged transcription in living cells using a locus-specific reporter system, which allowed precise, single-cell kinetic measurements of promoter binding, initiation and elongation. Photobleaching of fluorescent RNA polymerase II revealed several kinetically distinct populations of the enzyme interacting with a specific gene. Photobleaching and photoactivation of fluorescent MS2 proteins used to label nascent messenger RNAs provided sensitive elongation measurements. A mechanistic kinetic model that fits our data was validated using specific inhibitors. Polymerases elongated at 4.3 kilobases min(-1), much faster than previously documented, and entered a paused state for unexpectedly long times. Transcription onset was inefficient, with only 1% of polymerase-gene interactions leading to completion of an mRNA. Our systems approach, quantifying both polymerase and mRNA kinetics on a defined DNA template in vivo with high temporal resolution, opens new avenues for studying regulation of transcriptional processes in vivo. KW - biophysics KW - rna KW - transcription KW - trp3 AU - Darzacq, Xavier AU - Shav-Tal, Yaron AU - de Turris, Valeria AU - Brody, Yehuda AU - Shenoy, Shailesh M AU - Phair, Robert D AU - Singer, Robert H PY - 2007/08/05/ UR - http://dx.doi.org/10.1038/nsmb1280 ER -