Recruitment timing and dynamics of transcription factors at the Hsp70 loci in living cells.

Recruitment timing and dynamics of transcription factors at the Hsp70 loci in living cells.
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DOI:
10.1016/j.molcel.2010.11.022
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发表时间:
2010-12-22
期刊:
影响因子:
16
通讯作者:
Lis JT
Lis JT
中科院分区:
生物学1区
文献类型:
--
作者:
Zobeck KL;Buckley MS;Zipfel WR;Lis JT

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染色质免疫沉淀(ChIP)研究提供了细胞群中染色质因子的快照。在这里,我们使用活细胞成像技术在高时间分辨率下研究了转录因子在个体果蝇唾液腺核中诱导Hsp70位点的募集和动态。主调控因子HSF的募集在基因激活后20秒内首先被检测到,其募集的时间从RNA聚合酶II和P-TEFb中确定,这些因子从Spt6和Topo i中确定。值得注意的是,每个因子的募集在不同细胞之间是高度同步的。此外,荧光漂白后恢复(Fluorescence Recovery after Photobleaching, FRAP)分析显示,多个因子的进出受到基因激活的逐渐限制,表明转录室逐渐形成。此外,我们证明了polyadp -核糖(PAR)聚合酶活性是维持转录室所必需的。我们提出PAR聚合物在转录室中局部保留因子。
Chromatin immunoprecipitation (ChIP) studies provide snapshots of factors on chromatin in cell populations. Here, we use live cell imaging to examine at high temporal resolution the recruitment and dynamics of transcription factors to the inducible Hsp70 loci in individual Drosophila salivary gland nuclei. Recruitment of the master regulator, HSF, is first detected within 20 sec of gene activation and the timing of its recruitment resolves from RNA polymerase II and P-TEFb, and these factors resolve from Spt6 and Topo I. Remarkably, the recruitment of each factor is highly synchronous between different cells. In addition, Fluorescence Recovery after Photobleaching (FRAP) analyses show that the entry and exit of multiple factors are progressively constrained upon gene activation, suggesting the gradual formation of a transcription compartment. Furthermore, we demonstrate that PolyADP-Ribose (PAR) Polymerase activity is required to maintain the transcription compartment. We propose that PAR polymers locally retain factors in a transcription compartment.
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