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首页> 外文期刊>Molecular and Cellular Biology >Requirement of TRAP/Mediator for Both Activator-Independent and Activator-Dependent Transcription in Conjunction with TFIID-Associated TAFIIs
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Requirement of TRAP/Mediator for Both Activator-Independent and Activator-Dependent Transcription in Conjunction with TFIID-Associated TAFIIs

机译：TRAP /介体对与TFIID相关的TAFII结合的非激活子和激活子转录的要求

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The multiprotein human TRAP/Mediator complex, which is phylogenetically related to the yeast SRB/Mediator coactivator, facilitates activation through a wide variety of transcriptional activators. However, it remains unclear how TRAP/Mediator functions in the context of other coactivators. Here we have identified a previously uncharacterized integral subunit (TRAP25) of the complex that is apparently metazoan specific. An antibody that is specific for TRAP25 allowed quantitative immunodepletion of essentially all TRAP/Mediator components from HeLa nuclear extract, without detectably affecting levels of RNA polymerase II and corresponding general transcription factors. Surprisingly, the TRAP/Mediator-depleted nuclear extract displayed severely reduced levels of both basal and activator-dependent transcription from DNA templates. Both activities were efficiently restored upon readdition of purified TRAP/Mediator. Moreover, restoration of basal and activator-dependent transcription to extracts that were simultaneously depleted of TRAP/Mediator and TFIID (TBP plus the major TAFIIs) required addition of both TBP and associated TAFIIs, as well as TRAP/Mediator. These observations indicate that TAFIIs and Mediator are jointly required for both basal and activated transcription in the context of a more physiological complement of nuclear proteins. We propose a close mechanistic linkage between these components that most likely operates at the level of combined effects on the general transcription machinery and, in addition, a direct role for Mediator in relaying activation signals to this machinery.
机译：与酵母SRB /介体共激活剂在系统发育上相关的多蛋白人TRAP /介体复合物，通过多种转录激活剂促进了激活。但是，尚不清楚TRAP /介体在其他共激活剂的背景下如何发挥作用。在这里，我们确定了该复合物的一个先前未表征的整体亚基（TRAP25），该亚基显然是后生的。对TRAP25特异的抗体可对HeLa核提取物中的所有TRAP /介体成分进行定量免疫消除，而不会检测到RNA聚合酶II和相应的一般转录因子的水平。出乎意料的是，TRAP /介体耗尽的核提取物显示出DNA模板中基础和激活子依赖性转录的水平大大降低。重新纯化纯化的TRAP /介体后，两种活性均得以有效恢复。此外，要恢复基础和活化剂依赖性转录至同时耗尽TRAP /介体和TFIID（TBP加上主要TAF II ）的提​​取物，则需要同时添加TBP和相关的TAF II 以及TRAP /介体。这些观察结果表明，在更生理地补充核蛋白的情况下，基础转录和活化转录共同需要TAF II s和介体。我们建议这些组件之间建立紧密的机械联系，最有可能在对通用转录机制产生综合影响的水平上发挥作用，此外，介体在将激活信号中继至该机制中也起直接作用。

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