Machado A K, Morgan B A, Merrill G F
Department of Biochemistry and Biophysics and Center for Gene Research and Biotechnology, Oregon State University, Corvallis, Oregon 97331, USA.
J Biol Chem. 1997 Jul 4;272(27):17045-54. doi: 10.1074/jbc.272.27.17045.
Mlu1 cell cycle box (MCB) elements are found near the start site of yeast genes expressed at G1/S. Basal promoters dependent on the elements for upstream activating sequence activity are inactive in Deltaswi6 yeast. Yeast were screened for mutations that activated MCB reporter genes in the absence of Swi6. The mutations identified a single complementation group. Functional cloning revealed the mutations were alleles of the TRR1 gene encoding thioredoxin reductase. Although deletion of TRR1 activated MCB reporter genes, high copy expression did not suppress reporter gene activity. The trr1 mutations strongly (20-fold) stimulated MCB- and SCB (Swi4/Swi6 cell cycle box)-containing reporter genes, but also weakly (3-fold) stimulated reporter genes that lacked these elements. The trr1 mutations did not affect the level or periodicity of three endogenous MCB gene mRNAs (TMP1, RNR1, and SWI4). Deletion of thioredoxin genes TRX1 and TRX2 recapitulated the stimulatory effect of trr1 mutations on MCB reporter gene activity. Conditions expected to oxidize thioredoxin (exposure to H2O2) induced MCB gene expression, whereas conditions expected to conserve thioredoxin (exposure to hydroxyurea) inhibited MCB gene expression. The results suggest that thioredoxin oxidation contributes to MCB element activation and suggest a link between thioredoxin-oxidizing processes such as ribonucleotide reduction and cell cycle-specific gene transcription.
Mlu1细胞周期框（MCB）元件位于在G1/S期表达的酵母基因起始位点附近。依赖这些元件发挥上游激活序列活性的基础启动子在缺失Swi6的酵母中无活性。对酵母进行筛选，寻找在缺失Swi6时能激活MCB报告基因的突变。这些突变确定了一个单一的互补群。功能克隆显示这些突变是编码硫氧还蛋白还原酶的TRR1基因的等位基因。虽然缺失TRR1能激活MCB报告基因，但高拷贝表达并不能抑制报告基因活性。trr1突变强烈（20倍）刺激含有MCB和SCB（Swi4/Swi6细胞周期框）的报告基因，但也微弱（3倍）刺激缺乏这些元件的报告基因。trr1突变不影响三种内源性MCB基因mRNA（TMP1、RNR1和SWI4）的水平或周期性。缺失硫氧还蛋白基因TRX1和TRX2重现了trr1突变对MCB报告基因活性的刺激作用。预期会氧化硫氧还蛋白的条件（暴露于H2O2）诱导MCB基因表达，而预期会保存硫氧还蛋白的条件（暴露于羟基脲）抑制MCB基因表达。结果表明硫氧还蛋白氧化有助于MCB元件激活，并表明硫氧还蛋白氧化过程（如核糖核苷酸还原）与细胞周期特异性基因转录之间存在联系。
