MOLECULAR MECHANISMS OF DNA REPAIR IN YEAST

酵母有丝分裂细胞中DNA修复系统的鉴定 酿酒酵母正在接受检查：a)它们对细胞的保护作用 减数分裂，以及b)相应基因在正常减数分裂中的作用。我们 开发了独特的蔗糖梯度技术来检查修复后 低剂量紫外线或电离辐射并跟踪减数分裂DNA的变化 在减数分裂期间。 RAD50、RAD52和RAD57基因在DNA修复中是必不可少的 有丝分裂细胞中的双链断裂。它们还需要用于 减数分裂。这些基因的突变会取消正常的减数分裂重组；它 RAD50似乎在减数分裂的早期阶段起作用。罕见的单链 之后不久，在rad52和rad57菌株中观察到了中断(SSI) 减数分裂DNA合成的开始，这些似乎与 重组。温和的隔离技术使 DNA断裂的SSI特征；许多SSIS具有3‘OH和5’PO4 终极迷你。SSI似乎不是随机分布的，基于 涉及特定染色体区域的探针的实验表明 参与正常减数分裂重组的特定位点或区域。 而rad52和rad57突变体在减数分裂重组中存在缺陷， 在承诺进行减数分裂之前，可以回收重组体。 在rad52突变体中的频率比在RAD菌株中低得多； 在rad57突变体中，频率较低。重组体也是 与RAD中的那些在性质上不同。当减数分裂停止时， RAD52或RAD57暴露在生长介质中，重组子不能 恢复了。这是因为重组子需要更长的时间来 形式，并建议在中间步骤阻止rad52和rad57 在重组中。重组中间体缓慢分解，并且 在分解之前的生长阻止了重组体的出现。

DNA repair systems identified in mitotic cells of the yeast Saccharomyces cerevisiae are being examined for a) their protection of cells undergoing meiosis, and b) the role of the corresponding genes in normal meiosis. We have developed unique sucrose gradient techniques to examine repair after low doses of UV or ionizing radiation and to follow changes in meiotic DNA during meiosis. The RAD50, RAD52 and RAD57 genes are essential in the repair of DNA double-strand breaks in mitotic cells. They are also required for meiosis. Mutations in these genes abolish normal meiotic recombination; it appears that RAD50 acts at an early step in meiosis. Rare single-strand interruptions (SSIs) were observed in rad52 and rad57 strains shortly after the beginning of meiotic DNA synthesis and these appear to be related to recombination. Gentle isolation techniques have allowed the characterization of SSIs as breaks in DNA; many have 3' OH and 5' PO4 termini. The SSIs do not appear to be randomly distributed, based on experiments involving probes for specific chromosomal regions, suggesting specific sites or regions involved in normal meiotic recombination. While rad52 and rad57 mutants are defective in meiotic recombination, recombinants can be recovered prior to commitment to reductional division. The frequency in rad52 mutants is much lower than in RAD+ strains; the frequency is somewhat lower in rad57 mutants. The recombinants are also qualitatively different from those in RAD+. When meiosis is arrested and rad52 or rad57 are exposed to growth medium, recombinants are not recovered. This is due to the extended time necessary for recombinants to form, and suggests that rad52 and rad57 are blocked at an intermediate step in recombination. The recombination intermediates are resolved slowly, and growth prior to resolution prevents the appearance of recombinants.