Ingles C J
Proc Natl Acad Sci U S A. 1978 Jan;75(1):405-9. doi: 10.1073/pnas.75.1.405.
Mutant Chinese hamster ovary cell lines temperature-sensitive (TS) for growth and containing TS mutations in RNA polymerase II (nucleosidetriphosphate:RNA nucleotidyltransferase, EC 2.7.7.6) have been isolated. Wild-type cells were treated with the mutagen N-methyl-N'-nitro-N-nitrosoguanidine and a population of cells possessing mutations in RNA polymerase II was initially selected by isolating alpha-amanitin-resistant clones at 34 degrees . Of 168 such alpha-amanitin-resistant isolates screened for temperature sensitivity, nine were TS for growth at 39.5 degrees . By examining the behavior of the alpha-amanitin resistance of these TS cell lines in somatic cell hybrids, the TS mutation in a number of them was shown to be in RNA polymerase II. Hybrid cells obtained by the fusion of the TS and alpha-amanitin-resistant cells with cells possessing alpha-amanitin-sensitive polymerase II grew at both 34 degrees and 39.5 degrees ; the TS mutations were recessive. At 34 degrees all the hybrids were alpha-amanitin-resistant and possessed a mixture of alpha-amanitin-resistant and sensitive polymerase II. At 39.5 degrees the alpha-amanitin-resistant polymerase II activities in hybrids of four of the TS cell lines were lost; these four lines were alpha-amanitin-sensitive and possessed only alpha-amanitin-sensitive polymerase II. Temperature-insensitive revertants of two of these mutants were isolated. Reversion of the TS phenotype for mutants TsAma(R)-1 and TsAma(R)-8 was accompanied by an alteration in the level of alpha-amanitin resistance of the RNA polymerase II activities in the revertant cells. Together these data provide convincing evidence that TS mutations in RNA polymerase II can be coselected with alpha-amanitin resistance.
已分离出对生长温度敏感（TS）且RNA聚合酶II（核苷三磷酸：RNA核苷酸转移酶，EC 2.7.7.6）含有TS突变的突变型中国仓鼠卵巢细胞系。野生型细胞用诱变剂N-甲基-N'-硝基-N-亚硝基胍处理，最初通过在34℃下分离抗α-鹅膏蕈碱克隆来选择一群在RNA聚合酶II中具有突变的细胞。在筛选温度敏感性的168个此类抗α-鹅膏蕈碱分离株中，有9个在39.5℃下生长呈TS型。通过检查这些TS细胞系在体细胞杂种中抗α-鹅膏蕈碱的行为，发现其中一些细胞系的TS突变存在于RNA聚合酶II中。TS和抗α-鹅膏蕈碱细胞与具有α-鹅膏蕈碱敏感聚合酶II的细胞融合得到的杂种细胞在34℃和39.5℃下均能生长；TS突变是隐性的。在34℃时，所有杂种细胞均抗α-鹅膏蕈碱，并具有抗α-鹅膏蕈碱和敏感聚合酶II的混合物。在39.5℃时，四个TS细胞系杂种中的抗α-鹅膏蕈碱聚合酶II活性丧失；这四个细胞系对α-鹅膏蕈碱敏感，仅具有α-鹅膏蕈碱敏感聚合酶II。分离出了其中两个突变体的温度不敏感回复体。突变体TsAma(R)-1和TsAma(R)-8的TS表型回复伴随着回复体细胞中RNA聚合酶II活性的α-鹅膏蕈碱抗性水平的改变。这些数据共同提供了令人信服的证据，表明RNA聚合酶II中的TS突变可与抗α-鹅膏蕈碱共同选择。
