MECHANISM OF 5S RNA SYNTHESIS

非洲爪蟾5S RNA基因转录控制的研究已经 建立了这个基因系统，作为理解 真核生物中转录的调节。 5S RNA基因被激活， 一种转录因子，命名为TFIIIA， 120个核苷酸长的基因。 这个转录因子， 锌指DNA结合蛋白的原型，具有新的能力， 与5S RNA和5S DNA特异性结合。 实验提出了 为了进一步研究含有5S的7S颗粒的详细结构， RNA和TFIIIA。 初步结果导致了7S粒子的模型 这些结构将在这些实验中直接测试。 结合 TFIIIA对该基因的作用只是转录激活的第一步。 转录因子TFIIIC与因子A-DNA复合物结合， 稳定的复合物，抵抗第二模板的竞争。 进一步 将进行爪蟾TFIIIC的纯化以确定 它的两个可分离的组件在结构和功能上类似于 人TFIIIC 1和TFIIIC 2。 DNA酶保护实验将在 进行以确定非洲爪蟾TFIIIC 2是否结合到 转录起始，正如对其人类对应物所建议的那样。 TFIIIIa之间间隔改变的突变5S RNA基因的集合 结合位点和起始位点（含有推测的TFIIIC 2 结合位点）将被研究以关联转录因子结合和 DNA酶保护稳定复合物。 一种新的蛋白酶保护剂 “足迹”技术将在稳定的综合体中发展。 三 计划进行一系列实验来研究RNA聚合酶III的作用。 首先，将采用化学标记程序来识别 在RNA聚合酶的活性位点中的多肽， 转录起始和早期延伸模式。 二是 RNA聚合酶III中活性蛋白激酶的意义将是 研究了 最后，RNA聚合酶II是否是 能够准确终止，而不需要La抗原作为 将重新调查终止因素。

Studies of the control of transcription of Xenopus 5S RNA genes have established this gene system as a leading model for understanding the regulation of transcription in eukaryotes. The 5S RNA gene is activated by the binding of a transcription factor, designated TFIIIA, to the enter of the 120 nucleotide long gene. This transcription factor, which is the prototype for a zinc-finger DNA binding protein, has the novel ability to bind specifically to 5S RNA as well as to 5S DNA. Experiments are proposed to further study the detailed structure of the 7S particle containing 5S RNA and TFIIIA. Preliminary results have led to a model for 7S particle structure that will be directly tested in these experiments. Binding of TFIIIA to the gene is only the first step in transcriptional activation. Transcription factor TFIIIC binds to the factor A-DNA complex to form a stable complex that resists competition by a second template. Further purification of Xenopus TFIIIC will be conducted to determine the extent to which its two separable components structurally and functionally resemble the human TFIIIC1 and TFIIIC2. DNase protection experiments will be conducted to determine whether the Xenopus TFIIIC2 binds to the site of transcription initiation, as has been suggested for its human counterpart. A collection of mutant 5S RNA genes with altered spacing between the TFIIIa binding site and the initiation site (containing the presumed TFIIIC2 binding site) will be studied to correlate transcription factor binding and DNAse protection stable complex. A novel protease-protection "footprinting" technique will be developed in the stable complex. Three series of experiments are planned to study the role of RNA polymerase III. First, a chemical labeling procedure will be adapted to identify the polypeptides in the active site of the RNA polymerase in both the initiation and early elongation modes of transcription. Second, the significance of an active protein kinase in RNA polymerase III will be investigated. Finally, the question of whether RNA polymerase II is capable of accurate termination without the action of La antigen as a termination factor will be reinvestigated.