A POSITIVE REGULATORY GENE IS REQUIRED FOR ACCUMULATION OF THE FUNCTIONAL MESSENGER-RNA FOR THE GLUCOSE-REPRESSIBLE ALCOHOL-DEHYDROGENASE FROM SACCHAROMYCES-CEREVISIAE
A POSITIVE REGULATORY GENE IS REQUIRED FOR ACCUMULATION OF THE FUNCTIONAL MESSENGER-RNA FOR THE GLUCOSE-REPRESSIBLE ALCOHOL-DEHYDROGENASE FROM SACCHAROMYCES-CEREVISIAE
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DOI：
10.1016/0022-2836(81)90181-9
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发表时间：
1981-01-01
影响因子：
5.6
通讯作者：
YOUNG, ET
中科院分区：
文献类型：
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作者：
DENIS, CL;CIRIACY, M;YOUNG, ET
葡萄糖抑制性醇脱氢酶（ADH）的量受其功能mRNA的量调节。通过放射免疫分析检测ADHII蛋白，并通过金黄色葡萄球菌蛋白酶的有限蛋白水解将其与经典的ADH同工酶ADHI区分开来。当酵母含有野生型等位基因ADR 2（ADHII结构基因座）和ADR 1（其正调控基因）的脉冲标记与[35 S]蛋氨酸在去阻遏，放射性标记积累在抗体沉淀的ADHII coterminously与ADHII活性的外观。动力学的功能ADHII mRNA的外观在该菌株的去阻遏过程中被证明是相同的ADHII蛋白质的合成在体内时，RNA，从去阻遏的细胞中提取的，在小麦胚芽无细胞翻译系统中翻译。使用该位点突变的2株菌株分析了正调节基因ADR 1在ADHII表达中的作用。含有adr 1 -1等位基因的酵母不能去抑制ADH II活性。当该菌株在去阻遏期间用[35 S]甲硫氨酸脉冲标记时，检测到的ADH II蛋白合成水平约为野生型菌株的1/10至1/20。当从含有adr 1 -1等位基因的细胞中提取RNA并在小麦胚芽无细胞系统中翻译时，几乎没有功能性的ADHII mRNA存在。使用含有ADR 1 -5c等位基因的菌株进一步分析ADR 1基因的作用，该等位基因允许组成型合成ADHII活性。在葡萄糖抑制过程中。ADHII蛋白合成和功能mRNA的量处于与野生型菌株在完全去阻遏后所发现的水平相当的水平。类似的动力学的ADH II蛋白质合成和mRNA的积累在去阻遏过程中观察到携带ADR 1 -5c等位基因的菌株相比，携带ADR 1 -5c等位基因，但绝对量大3- 4倍，在细胞中含有ADR 1 -5c等位基因。ADR 1基因在去阻遏过程中明显地起作用以增加功能性ADHII mRNA的水平。
The amount of glucose-repressible alcohol dehydrogenase (ADH) is regulated by the amount of its functional mRNA. ADHII protein was detected by a radioimmune assay and differentiated from ADHI, the classical ADH isozyme, by limited proteolysis with Staphylococcus aureus protease. When yeast containing the wild-type alleles for ADR2 (the ADHII structural locus) and for ADR1 (its positive regulatory gene) were pulse-labeled with [35S]methionine during derepression, radioactive label accumulated in the antibody-precipitated ADHII coterminously with the appearance of ADHII activity. The kinetics of functional ADHII mRNA appearance during derepression in this strain were shown to be the same as those for ADHII protein synthesis in vivo when RNA, extracted from derepressed cells, was translated in a wheat germ cell-free translation system. The role of the positive regulatory gene, ADR1, in ADHII expression was analyzed using 2 strains mutated at that locus. Yeast containing the adr1-1 allele are incapable of derepressing ADHII activity. When this strain was pulse-labeled with [35S]methionine during derepression, approximately 1/10 to 1/20 the level of ADHII protein synthesis was detected as in the wild-type strain. When RNA was extracted during derepression from cells containing the adr1-1 allele and translated in a wheat germ cell-free system, little functional ADHII mRNA was present. The role of the ADR1 gene was further analyzed using a strain containing the ADR1-5c allele, which allows constitutive synthesis of ADHII activity. In this strain during glucose repression. ADHII protein synthesis and amount of functional mRNA were at levels comparable to those found for the wild-type strain after complete derepression. Similar kinetics of ADHII protein synthesis and of mRNA accumulation during derepression were observed in the strain carrying the ADR1-5c allele when compared to that carrying the ADR1 allele, but the absolute amounts were greater by 3- to 4-fold in cells containing the ADR1-5c allele. The ADR1 gene apparently acts to increase the level of functional ADHII mRNA during derepression.