Steinberg T H, Mathews D E, Durbin R D, Burgess R R
Department of Oncology, McArdle Laboratory for Cancer Research, University of Wisconsin-Madison.
J Biol Chem. 1990 Jan 5;265(1):499-505.
We demonstrate that tagetitoxin, a bacterial phytotoxin, preferentially inhibits eukaryotic RNA polymerase III. We used promoter-directed transcription of cloned genes in cell-free extracts to compare tagetitoxin inhibition of RNA polymerases from diverse sources. In HeLa cell extracts, accumulation of 5 S rRNA, and U6 snRNAs (transcribed by RNA polymerase III) was inhibited at 0.3-3.0 microM tagetitoxin but transcription from adenovirus 2 major late promoter (RNA polymerase II) was not significantly affected at concentrations below 30 microM. Tagetitoxin also inhibited promoter-directed RNA polymerase III transcription in cell-free extracts from Bombyx mori (pre-tRNA), and Saccharomyces cerevisiae (pre-tRNA) at 0.3-3.0 microM, concentrations that also inhibit chloroplast or bacterial promoter-directed transcription. In nonspecific transcription assays, partially purified B. mori RNA polymerase III was inhibited by tagetitoxin at concentrations that inhibit Escherichia coli RNA polymerase; purified calf thymus RNA polymerase II was not inhibited by tagetitoxin. Using injection into Xenopus laevis oocytes, we compared tagetitoxin effects on accumulation of U1 snRNA, hH2B mRNA (transcribed by RNA polymerase II), 5 S rRNA and U6 snRNA (RNA polymerase III), and 5.8 S rRNA (RNA polymerase I). In Xenopus oocytes, RNA polymerase III transcription was preferentially inhibited by tagetitoxin.
我们证明了细菌植物毒素塔格毒素优先抑制真核生物RNA聚合酶III。我们利用无细胞提取物中克隆基因的启动子指导转录，来比较塔格毒素对不同来源RNA聚合酶的抑制作用。在HeLa细胞提取物中，0.3 - 3.0微摩尔的塔格毒素可抑制5S rRNA和U6 snRNA（由RNA聚合酶III转录）的积累，但在浓度低于30微摩尔时，腺病毒2主要晚期启动子（RNA聚合酶II）的转录未受到显著影响。塔格毒素在0.3 - 3.0微摩尔时，也抑制了家蚕（前体tRNA）和酿酒酵母（前体tRNA）无细胞提取物中启动子指导的RNA聚合酶III转录，该浓度也抑制叶绿体或细菌启动子指导的转录。在非特异性转录试验中，部分纯化的家蚕RNA聚合酶III在抑制大肠杆菌RNA聚合酶的浓度下被塔格毒素抑制；纯化的小牛胸腺RNA聚合酶II未被塔格毒素抑制。通过注射到非洲爪蟾卵母细胞中，我们比较了塔格毒素对U1 snRNA、hH2B mRNA（由RNA聚合酶II转录）、5S rRNA和U6 snRNA（RNA聚合酶III）以及5.8S rRNA（RNA聚合酶I）积累的影响。在非洲爪蟾卵母细胞中，RNA聚合酶III转录优先被塔格毒素抑制。
