Escherichia coli cells expressing a mutant glyV (glycine tRNA) gene have a UVM-constitutive phenotype: implications for mechanisms underlying the mutA or mutC mutator effect.
Escherichia coli cells expressing a mutant glyV (glycine tRNA) gene have a UVM-constitutive phenotype: implications for mechanisms underlying the mutA or mutC mutator effect.
复制标题
表达突变 glyV（甘氨酸 tRNA）基因的大肠杆菌细胞具有 UVM 组成型表型：对 mutA 或 mutC 突变效应潜在机制的影响。
DOI：
10.1128/jb.179.23.7507-7514.1997
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发表时间：
1997
影响因子：
3.2
通讯作者：
Humayun,MZ
中科院分区：
文献类型：
--
作者：
Murphy,HS;Humayun,MZ
将携带3，n4 -乙基胞嘧啶病变的M13单链病毒DNA转染到经紫外线预处理的大肠杆菌细胞中，与未经处理的细胞相比，病变部位的诱变率显著升高。这种反应被称为UVM，即紫外线诱变调制，由多种DNA损伤剂诱导，不同于已知的细胞对DNA损伤的反应，包括SOS反应。本报告描述了我们的观察结果，作为我们对UVM现象的调查的一部分，即携带突变a或突变c等位基因的大肠杆菌细胞显示出UVM构成表型。这些突变等位基因最近被定位（M. M. Slupska， C. Baikalov， R. Lloyd, and J. H. Miller, Proc. Natl）。学会科学。glyV （mutA）和glyW (mutC) tRNA基因。每个突变等位基因都是由反密码子序列的相同突变引起的，因此突变的trna原则上可以将mRNA中的天冬氨酸密码子误译为低水平的甘氨酸。由于tRNA基因突变导致的uvm构成表型是意料之外的，因此我们进行了一系列实验，旨在测试该表型是否确实由突变的甘氨酸tRNA表达介导。我们将野生型或突变型glyV基因置于质粒载体上的异源诱导启动子的控制下。表达突变glyV基因的大肠杆菌细胞表现出以下所有三种表型：(i)测试等位基因的错义抑制，（ii）通过突变对利福平抗性测量的突变表型，以及（iii） uvm构成表型。这些表型与表达野生型glyV基因的细胞或存在突变等位基因但未被转录诱导的细胞无关。这些观察结果为突变tRNA的表达可以赋予突变表型的观点提供了强有力的支持，包括在mutA和mutC细胞中观察到的uvm构成表型。然而，我们的数据表明聚合酶III的epsilon亚基的低水平误译可能不能解释观察到的uvm构成表型。我们的研究结果还表明，mutA δ tareca双突变体显示正常的UVM表型，表明mutA效应依赖于recA。这里报道的观察结果提出了一些有趣的问题，并提出了UVM反应是通过复制环境的短暂改变来介导的可能性。
Transfection of M13 single-stranded viral DNA bearing a 3,N4-ethenocytosine lesion into Escherichia coli cells pretreated with UV results in a significant elevation of mutagenesis at the lesion site compared to that observed in untreated cells. This response, termed UVM, for UV modulation of mutagenesis, is induced by a variety of DNA-damaging agents and is distinct from known cellular responses to DNA damage, including the SOS response. This report describes our observation, as a part of our investigation of the UVM phenomenon, that E. coli cells bearing a mutA or mutC allele display a UVM-constitutive phenotype. These mutator alleles were recently mapped (M. M. Slupska, C. Baikalov, R. Lloyd, and J. H. Miller, Proc. Natl. Acad. Sci. USA 93:4380-4385, 1996) to the glyV (mutA) and glyW (mutC) tRNA genes. Each mutant allele was shown to arise by an identical mutation in the anticodon sequence such that the mutant tRNAs could, in principle, mistranslate aspartate codons in mRNA as glycine at a low level. Because a UVM-constitutive phenotype resulting from a mutation in a tRNA gene was unexpected, we undertook a series of experiments designed to test whether the phenotype was indeed mediated by the expression of mutant glycine tRNAs. We placed either a wild-type or a mutant glyV gene under the control of a heterologous inducible promoter on a plasmid vector. E. coli cells expressing the mutant glyV gene displayed all three of the following phenotypes: (i) missense suppression of a test allele, (ii) a mutator phenotype measured by mutation to rifampin resistance, and (iii) a UVM-constitutive phenotype. These phenotypes were not associated with cells expressing the wild-type glyV gene or with cells in which the mutant allele was present but was not transcriptionally induced. These observations provide strong support for the idea that expression of mutant tRNA can confer a mutator phenotype, including the UVM-constitutive phenotype observed in mutA and mutC cells. However, our data imply that low-level mistranslation of the epsilon subunit of polymerase III probably does not account for the observed UVM-constitutive phenotype. Our results also indicate that mutA deltarecA double mutants display a normal UVM phenotype, suggesting that the mutA effect is recA dependent. The observations reported here raise a number of intriguing questions and raise the possibility that the UVM response is mediated through transient alteration of the replication environment.