Steinberg T H, Burgess R R
McArdle Laboratory for Cancer Research, Department of Oncology, University of Wisconsin, Madison 53706.
J Biol Chem. 1992 Oct 5;267(28):20204-11.
In yeast nuclear extracts, tagetitoxin inhibition of RNA polymerase III promoter-directed single- and multiple-round transcription is characterized by pausing or stalling of the elongation complex at several discrete points on the template. Paused ternary complexes isolated from tagetitoxin-inhibited reactions can be elongated to produce full-length RNA. The distribution of "tagetitoxin-enhanced" pause sites is distinct for each of the class III genes we have examined. These tagetitoxin-enhanced pause sites may also be intrinsic pause sites for the elongation complex. Tagetitoxin inhibition of in vitro transcription of the yeast SUP4 and SUP6 tRNA(Tyr) genes demonstrates template dependence and indicates that inhibition may occur after UMP incorporation. Factor-independent transcription by purified yeast RNA polymerase III can also be inhibited by tagetitoxin, and the degree of inhibition is template-dependent. Tagetitoxin may be most effective as an inhibitor under conditions where polymerase III tends to pause on the template. We propose that differences in tagetitoxin inhibition among class III genes may reflect differences in the number of stability of these pause sites.
在酵母核提取物中，tagetitoxin对RNA聚合酶III启动子指导的单轮和多轮转录的抑制作用表现为延伸复合物在模板上的几个离散点处暂停或停滞。从tagetitoxin抑制反应中分离出的暂停三元复合物可以延伸以产生全长RNA。我们检测的每个III类基因的“tagetitoxin增强”暂停位点的分布都是不同的。这些tagetitoxin增强的暂停位点也可能是延伸复合物的固有暂停位点。tagetitoxin对酵母SUP4和SUP6 tRNA（Tyr）基因体外转录的抑制作用显示出模板依赖性，并表明抑制可能在UMP掺入后发生。纯化的酵母RNA聚合酶III的不依赖因子的转录也可被tagetitoxin抑制，且抑制程度取决于模板。在聚合酶III倾向于在模板上暂停的条件下，tagetitoxin可能作为一种抑制剂最为有效。我们认为III类基因之间tagetitoxin抑制作用的差异可能反映了这些暂停位点稳定性数量的差异。
