Dean N, Berk A J
Molecular Biology Institute, University of California, Los Angeles 90024.
Mol Cell Biol. 1988 Aug;8(8):3017-25. doi: 10.1128/mcb.8.8.3017-3025.1988.
The separation of the mammalian class III transcription factor TFIIIC into two functional components, termed TFIIIC1 and TFIIIC2, enabled an analysis of their functions in transcription initiation. Template competition assays were used to define the order with which these factors interact in vitro to form stable preinitiation complexes on the adenovirus VAI and Drosophila melanogaster tRNA(Arg) genes. The interaction between these genes and TFIIIC2, the factor that binds with high affinity to the B block, was both necessary and sufficient for template commitment. When either the VAI or tRNA(Arg) gene was preincubated with TFIIIC2 alone, transcription of a second gene added subsequently was excluded, indicating that TFIIIC2 bound stably to the first template. Furthermore, the interaction between TFIIIC2 and these genes must occur prior to that of TFIIIC1 or TFIIIB. Once TFIIIC2 was bound, TFIIIC1 could bind to the tRNA(Arg) and VAI genes, although its interaction with the VAI gene was less stable than that with the tRNA(Arg) gene. TFIIIB activity bound stably to the complex of both genes with TFIIIC2. These results demonstrate that TFIIIC2 is the first transcription factor to bind to these genes and that TFIIIB and TFIIIC1 can then interact in either order to form a preinitiation complex.
哺乳动物III类转录因子TFIIIC可分离为两个功能组分，即TFIIIC1和TFIIIC2，这使得对它们在转录起始中的功能进行分析成为可能。采用模板竞争试验来确定这些因子在体外相互作用的顺序，以便在腺病毒VAI和果蝇tRNA（Arg）基因上形成稳定的起始前复合物。这些基因与TFIIIC2（即与B区具有高亲和力结合的因子）之间的相互作用对于模板确定而言既是必要的也是充分的。当单独将VAI或tRNA（Arg）基因与TFIIIC2预孵育时，随后添加的第二个基因的转录被排除，这表明TFIIIC2稳定地结合到第一个模板上。此外，TFIIIC2与这些基因之间的相互作用必须在TFIIIC1或TFIIIB之前发生。一旦TFIIIC2结合，TFIIIC1就可以结合到tRNA（Arg）和VAI基因上，尽管它与VAI基因的相互作用比与tRNA（Arg）基因的相互作用更不稳定。TFIIIB活性稳定地结合到两个基因与TFIIIC2的复合物上。这些结果表明，TFIIIC2是第一个结合到这些基因上的转录因子，然后TFIIIB和TFIIIC1可以以任意顺序相互作用以形成起始前复合物。
