Ossipow V, Fonjallaz P, Schibler U
Département de Biologie Moléculaire Sciences II, CH-1211 Geneva 4, Switzerland.
Mol Cell Biol. 1999 Feb;19(2):1242-50. doi: 10.1128/MCB.19.2.1242.
Transcription initiation of protein-encoding genes involves the assembly of RNA polymerase II and a number of general transcription factors at the promoter. A mammalian RNA polymerase II complex containing all of the components required for promoter-specific transcription initiation can be isolated by immunopurification with a monoclonal antibody directed against the cyclin-dependent kinase CDK7, a subunit of the general transcription factor TFIIH. In vitro transcription by this immunopurified RNA polymerase II complex is effectively stimulated by thyroid embryonic factor (TEF), a basic leucine zipper transcription factor. Thus, the RNA polymerase II complex must also contain components required for activated transcription that interact with the transactivation domain of TEF. This conjecture was verified by affinity selection experiments in which the TEF transcription activation domain was used as a bait. Indeed, an RNA polymerase II complex containing all of the accessory proteins required for transcription initiation can be enriched by its affinity to recombinant proteins containing the TEF transactivation domain. These results are compatible with a mechanism by which TEF can recruit an RNA polymerase II holoenzyme to the promoter in a single step.
蛋白质编码基因的转录起始涉及RNA聚合酶II和一些通用转录因子在启动子处的组装。通过用针对细胞周期蛋白依赖性激酶CDK7（通用转录因子TFIIH的一个亚基）的单克隆抗体进行免疫纯化，可以分离出一种包含启动子特异性转录起始所需所有成分的哺乳动物RNA聚合酶II复合物。甲状腺胚胎因子（TEF，一种碱性亮氨酸拉链转录因子）可有效刺激这种免疫纯化的RNA聚合酶II复合物的体外转录。因此，RNA聚合酶II复合物还必须包含与TEF反式激活结构域相互作用的、用于激活转录的成分。通过以TEF转录激活结构域为诱饵的亲和选择实验验证了这一推测。实际上，一种包含转录起始所需所有辅助蛋白的RNA聚合酶II复合物可以通过其与含有TEF转录激活结构域的重组蛋白的亲和力而得到富集。这些结果与一种机制相符，即TEF可以一步将RNA聚合酶II全酶招募到启动子处。
