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: 2654914
• 负责人: Jirair K Bedoyan
• 金额: $ 2.62万
• 依托单位: FRED HUTCHINSON CANCER RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-30 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2654914
• 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 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 1) 分离并鉴定晚期粟酒裂殖酵母缺陷突变体 减数分裂重组的步骤，例如霍利迪结构的解析 在同源染色体减数分裂过程中形成，2) 使用分子测定法确定粟酒裂殖酵母减数分裂重组 中间体和产品。 遗传和生化方法 用于实现这些目标。 在遗传方法中，筛选 提议将在后期识别有缺陷的粟酒裂殖酵母突变体 减数分裂重组的步骤。 在这个屏幕中，一个突变体参与了 重组的后期步​​骤并产生不可存活的孢子被拯救 一种额外的早期重组突变体表型，可防止 交换从开始。 在生化方法中，分子 测定法用于检测和分析粟酒裂殖酵母减数分裂 重组中间体（例如，联合分子、切口和双 链断裂）和成熟的产品。 这些方法使用两个 与补骨脂素结合的三维 DNA 分离测定 DNA 交联，一种引物延伸测定法，用于确定 含有霍利迪结构的中间体，以及分离的突变体 来自前面提到的遗传方法。 在真核生物中，遗传 重组在以下方面发挥着核心作用：a）在两个方面产生多样性 细胞和有机体水平，b) 修复受损染色体，以及 c) 减数分裂过程中同源染色体的忠实分离。 重组畸变会产生 DNA 突变（例如， 取代和缺失）和染色体重排（例如， 易位）和丢失（例如，非整倍体）。 此类异常现象经常发生 临床表现为人类癌症和出生缺陷。 粟酒裂殖酵母 在这里用作模型真核生物，以进一步我们对 减数分裂重组的分子机制。 知识点 在这里获得的知识将使我们对几个原因有更多的了解 人类遗传性疾病（例如癌症）并可能导致 深入了解未来预防这些问题的方法。