PIC Activation through Functional Interplay between Mediator and TFIIH.
PIC Activation through Functional Interplay between Mediator and TFIIH.
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通过介体和 TFIIH 之间的功能相互作用激活 PIC。
DOI：
10.1016/j.jmb.2016.11.026
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发表时间：
2017
影响因子：
5.6
通讯作者：
Xue,Zhu
中科院分区：
文献类型：
--
作者：
Malik,Sohail;Molina,Henrik;Xue,Zhu
多蛋白介体辅激活因子复合物的功能在很大程度上是通过控制启动子结合的前起始复合物（PIC）的形成和功能来实现的，该复合物由RNA聚合酶II和一般转录因子组成。然而，Mediator究竟如何影响PIC，特别是招聘后，仍然不清楚。在这里，我们研究了介体对纯化成分重建的动物体外转录系统中基础转录的影响。我们的研究结果揭示了在PIC发展的早期阶段，调解人和TFIIH之间的密切功能相互作用。我们发现，在转录通常不需要TFIIH的条件下，介体实际上抑制转录。众所周知，TFIIH向PIC的招募是由调解员促进的，然后它会采取行动来缓解调解员诱导的抑制，以产生活性形式的PIC。PIC的凝胶迁移率变动分析和携带突变XPB易位酶亚基的TFIIH制剂的表征进一步表明，这种抑制的缓解是通过ATP水解消耗能量来实现的，这表明它与TFIIH的既定启动子熔解活性偶联。我们对这些结果的解释是，中介功能作为一个组装因子，促进PIC成熟的各个阶段。尽管介体的总体作用是刺激基础转录，但其与PIC的初始接合产生转录惰性PIC中间体，这需要能量消耗来完成该过程。
The multiprotein Mediator coactivator complex functions in large part by controlling the formation and function of the promoter-bound preinitiation complex (PIC), which consists of RNA polymerase II and general transcription factors. However, precisely how Mediator impacts the PIC, especially post-recruitment, has remained unclear. Here, we have studied Mediator effects on basal transcription in anin vitrotranscription system reconstituted from purified components. Our results reveal a close functional interplay between Mediator and TFIIH in the early stages of PIC development. We find that under conditions when TFIIH is not normally required for transcription, Mediator actually represses transcription. TFIIH, whose recruitment to the PIC is known to be facilitated by the Mediator, then acts to relieve Mediator-induced repression to generate an active form of the PIC. Gel mobility shift analyses of PICs and characterization of TFIIH preparations carrying mutant XPB translocase subunit further indicate that this relief of repression is achieved through expending energy via ATP hydrolysis, suggesting that it is coupled to TFIIH's established promoter melting activity. Our interpretation of these results is that Mediator functions as an assembly factor that facilitates PIC maturation through its various stages. Whereas the overall effect of the Mediator is to stimulate basal transcription, its initial engagement with the PIC generates a transcriptionally inert PIC intermediate, which necessitates energy expenditure to complete the process.
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