Ray B L, White C I, Haber J E
Rosentiel Basic Medical Research Center, Waltham, Massachusetts.
Mol Cell Biol. 1991 Oct;11(10):5372-80. doi: 10.1128/mcb.11.10.5372-5380.1991.
We sequenced two alleles of the MATa locus of Saccharomyces cerevisiae that reduce homothallic switching and confer viability to HO rad52 strains. Both the MATa-stk (J. E. Haber, W. T. Savage, S. M. Raposa, B. Weiffenbach, and L. B. Rowe, Proc. Natl. Acad. Sci. USA 77:2824-2828, 1980) and MATa-survivor (R. E. Malone and D. Hyman, Curr. Genet. 7:439-447, 1983) alleles result from a T----A base change at position Z11 of the MAT locus. These strains also contain identical base substitutions at HMRa, so that the mutation is reintroduced when MAT alpha switches to MATa. Mating-type switching in a MATa-stk strain relative to a MATa Z11T strain is reduced at least 50-fold but can be increased by expression of HO from a galactose-inducible promoter. We confirmed by Southern analysis that the Z11A mutation reduced the efficiency of double-strand break formation compared with the Z11T variant; the reduction was more severe in MAT alpha than in MATa. In MAT alpha, the Z11A mutation also creates a mat alpha 1 (sterile) mutation that distinguishes switches of MATa-stk to either MAT alpha or mat alpha 1-stk. Pedigree analysis of cells induced to switch in G1 showed that MATa-stk switched frequently (23% of the time) to produce one mat alpha 1-stk and one MAT alpha progeny. This postswitching segregation suggests that Z11 was often present in heteroduplex DNA that was not mismatch repaired. When mismatch repair was prevented by deletion of the PMS1 gene, there was an increase in the proportion of mat alpha 1-stk/MAT alpha sectors (59%) and in pairs of switched cells that both retained the stk mutation (27%). We conclude that at least one strand of DNA only 4 bp from the HO cut site is not degraded in most of the gene conversion events that accompany MAT switching.
我们对酿酒酵母MATa基因座的两个等位基因进行了测序，这两个等位基因可降低同宗配合转换并赋予HO rad52菌株生存能力。MATa - stk（J. E. 哈伯、W. T. 萨维奇、S. M. 拉波萨、B. 韦芬巴赫和L. B. 罗，《美国国家科学院院刊》77:2824 - 2828，1980）和MATa - 存活者（R. E. 马龙和D. 海曼，《当代遗传学》7:439 - 447，1983）等位基因均源于MAT基因座Z11位置的T→A碱基变化。这些菌株在HMRa处也含有相同的碱基替换，因此当MATα转换为MATa时，该突变会再次引入。与MATa Z11T菌株相比，MATa - stk菌株中的交配型转换至少降低了50倍，但可通过从半乳糖诱导型启动子表达HO来增加。我们通过Southern分析证实，与Z11T变体相比，Z11A突变降低了双链断裂形成的效率；在MATα中这种降低比在MATa中更严重。在MATα中，Z11A突变还产生了一个matα1（不育）突变，这可区分MATa - stk向MATα或matα1 - stk的转换。对在G1期诱导转换的细胞进行谱系分析表明，MATa - stk频繁转换（23%的时间），产生一个matα1 - stk和一个MATα后代。这种转换后分离表明Z11经常存在于未进行错配修复的异源双链DNA中。当通过缺失PMS1基因阻止错配修复时，matα1 - stk/MATα扇形区的比例增加（59%），并且在两个都保留stk突变的转换细胞对中也增加（27%）。我们得出结论，在伴随MAT转换的大多数基因转换事件中，距HO切割位点仅4个碱基对的DNA的至少一条链未被降解。
