MOLECULAR MECHANISMS OF S POMBE MEIOTIC RECOMBINATION

裂殖酵母减数分裂重组的分子机制

• 批准号: 2872617
• 负责人: Jirair K Bedoyan
• 金额: $ 3.25万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-02-01 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2872617
• 关键词: Schizosaccharomyces pombe; fungal genetics; gene complementation; gene mutation; genetic recombination; meiosis

The long-term objective of the proposed research is to determine the basic molecular mechanism of meiotic homologous recombination in the fission yeast Schizosaccharomyces pombe. The specific aims are to 1) isolate and characterize mutants of S. Pombe defective in late steps of meiotic recombination such as resolution of Holliday structures formed during meiosis between homologous chromosomes and 2) determine, using molecular assays, S. Pombe meiotic recombination intermediates and products. Both genetic and biochemical approaches are used to achieve these objectives. In the genetic approach, a screen is proposed which will identify S. Pombe mutants defective in late steps of meiotic recombination. In this screen, a mutant involved in a late step of recombination and produces inviable spores is rescued by an additional early recombination mutant phenotype which prevents exchange from starting. In the biochemical approaches, molecular assays are used for detecting and analyzing S. pombe meiotic recombination intermediates (e.g., joint molecules, nicks, and double strand breaks) and mature products. These approaches use a two- dimensional DNA separation assay in combination with psoralen crosslinking of DNA, a primer extension assay for determining intermediates containing Holliday structures, and the mutants isolated from the genetic approach mentioned earlier. In eukaryotes, genetic recombination plays central roles in a) generating diversity at both the cellular and organismal levels, b) repair of damaged chromosomes, and c) faithful segregation of homologous chromosomes during meiosis. Recombinational aberrations generate DNA mutations (e.g., substitutions and deletions), and chromosome rearrangements (e.g., translocations) and loss (e.g., aneuploidy). Such aberrancies are often clinically manifested as cancer and birth defects in humans. S. pombe is used here as a model eukaryote to further our understanding of the molecular mechanisms of meiotic recombination. The knowledge gained here will give use more understanding into the causes of several diseases of genetic origin in humans (e.g., cancer) and possibly lead to insight into ways for preventing them in the future.

拟议研究的长期目标是确定 水稻减数分裂同源重组的基本分子机制 裂殖酵母：裂殖酵母。具体目标是 对1)Pombe缺陷链霉菌后期突变株的分离和鉴定 减数分裂重组的步骤，如Holliday结构的分解 在同源染色体减数分裂过程中形成，2) 用分子检测技术检测Pombe链霉菌减数分裂重组 中间体和产品。遗传和生化两种方法 用来实现这些目标。在遗传学方法中，一个屏幕 将在晚些时候发现有缺陷的S.Pombe突变体 减数分裂重组的步骤。在这个屏幕上，一个突变体参与了一个 重组的后期步骤和产生无活性的孢子被拯救 一种额外的早期重组突变表型，可防止 交换从开始。在生化方法中，分子 检测方法用于检测和分析S.pombe减数分裂。 复合中间体(例如，关节分子、镍和双链 链断裂)和成熟的产品。这些方法使用两个- 补骨脂素与三维DNA分离法联合应用 DNA的交联法--一种检测DNA的引物延伸法 含有Holliday结构的中间体及其分离的突变体 与前面提到的遗传方法不同。在真核生物中，基因 重组在a)产生多样性方面发挥了核心作用 细胞和生物体水平，b)受损染色体的修复，以及 C)减数分裂过程中同源染色体的忠实分离。 重组像差产生DNA突变(例如， 替换和缺失)和染色体重排(例如， 易位)和丢失(例如，非整倍体)。这种反常现象经常是 临床上表现为人类的癌症和出生缺陷。庞贝乳杆菌 在这里用作真核生物的模型，以加深我们对 减数分裂重组的分子机制。《知识》 在这里获得的将有助于更多地理解几个 人类的遗传性疾病(如癌症)，并可能导致 洞察未来预防此类事件的方法。