生物谷報道：科學家們親眼目睹了活組織中的基因轉錄過程，這一最新的研究成果在線發(fā)表在8月號的《自然—結構和分子生物學》期刊上,。
RNA聚合酶II是一種將DNA信息轉錄到信使RNA的酶,，由這種酶所實施的轉錄是基因表達的核心，也是生物調控機制的關鍵所在,。以前,，絕大部分對這種轉錄過程的了解來自于用純化物質在生物體外所做的試管實驗，科學家們對RNA聚合酶在活體中的作用過程知之甚少,。
利用高級熒光成像技術,，Robert Singer和合作者量化測量了RNA聚合酶在活哺乳類動物細胞中實施遺傳轉錄的動力學過程，從而將更高級生物中的基因轉錄機理分析提高到了一個新水平,。這項工作的最終目標是建立活體中基因轉錄的量化模型,。在活體組織的轉錄中，他們發(fā)現(xiàn)了全新和意料之外的特征,。
首先,，他們認為只有大約1%的與基因相連結的RNA聚合酶參加了基因轉錄過程，并產(chǎn)生出信使RNA,。其次,，他們發(fā)現(xiàn)RNA聚合酶轉錄的速度比想象的更快，而且通常只是在周期延長時才暫時停止,。
在量化認識單個活體細胞中的轉錄機理過程中,，該新研究是一個里程碑性的貢獻。（科學時報）
原始出處:
Nature Structural & Molecularbiology
Published online: 5 August 2007; | doi:10.1038/nsmb1280
In vivo dynamics of RNA polymerase II transcription
Xavier Darzacq1, 2, Yaron Shav-Tal1, 3, Valeria de Turris1, Yehuda Brody3, Shailesh M Shenoy1, Robert D Phair4 & Robert H Singer1
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.
3 The Mina & Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900, Israel.
4 Integrative Bioinformatics, Inc., Los Altos, California 94024, USA.
Correspondence should be addressed to Robert H Singer [email protected]
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.
