Drosophila Genes Affecting Chromosome Segregation

影响染色体分离的果蝇基因

• 批准号: 7052762
• 负责人: MICHAEL L GOLDBERG
• 金额: $ 42.1万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-09-30 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7052762
• 关键词: Drosophilidae; Xenopus oocyte; affinity chromatography; antibody; arthropod genetics; cell cycle; centromere; chromosome movement; cytogenetics; developmental genetics; dynein ATPase; enzyme substrate; gene expression; gene mutation; laboratory rabbit; matrix assisted laser desorption ionization; protein protein interaction; protein structure function; serine threonine protein kinase; video microscopy; Drosophilidae; Xenopus oocyte; affinity chromatography; antibody; arthropod genetics; cell cycle; centromere; chromosome movement; cytogenetics; developmental genetics; dynein ATPase; enzyme substrate; gene expression; gene mutation; laboratory rabbit; matrix assisted laser desorption ionization; protein protein interaction; protein structure function; serine threonine protein kinase; video microscopy

DESCRIPTION (provided by applicant): Mistakes in chromosome segregation during meiosis or mitosis can lead to spontaneous abortion or to abnormalities such as Down syndrome. They have also been implicated in the processes leading to cancer and aging. Our laboratory has undertaken large-scale genetic screens using the organism Drosophila melanogaster to identify genes encoding proteins critical for ensuring proper chromosome segregation. In the first specific aim, we will investigate several novel components of the chromosomal kinetochores that were first found in our screens and that have been conserved among metazoans including humans. One area of focus in this aim is a multisubunit complex including the proteins ZW10, Rod, and Zwilch. This complex targets the molecular motor dynein to the kinetochore, and it is required in a currently unknown fashion for the operation of the spindle assembly checkpoint that regulates anaphase onset. We propose experiments to dissect the structure of this complex and to investigate its interactions with NudE, a dynein-associated protein whose role is currently not well understood. A second area of focus in this aim will explore the possibility that two new proteins identified in our genetic screens may together constitute a new subassembly at the kinetochore also needed for proper chromosome segregation. The second specific aim describes a new area of research for our laboratory. Mutations in greatwall, one of the Drosophila genes found in our screens, cause chromosome undercondensation because of delayed transit through prophase. The greatwall gene encodes a novel protein kinase with an unusual structure that is conserved in insects and vertebrates. We made antibody against the Xenopus Greatwall protein and used it to. deplete this kinase from oocyte extracts. To our surprise, Greatwall depletion prevented the extracts from entering or maintaining M phase. In the second specific aim, we will follow up these intriguing observations in order to understand Greatwall's role in mitotic progression. We propose experiments to define Greatwall's substrates as well as the upstream kinases involved in Greatwall activation during mitosis. Additional experiments will investigate Greatwall's possible developmental role in oocyte maturation and will search for other proteins with which this kinase associates. The proposed studies on Greatwall have the potential to offer unique mechanistic insights concerning mitotic entry in vertebrate cells.

描述（由申请人提供）：减数分裂或有丝分裂期间染色体分离的错误可能会导致自发流产或唐氏综合症等异常。它们也与导致癌症和衰老的过程有关。我们的实验室已经使用果蝇果蝇（Melanogaster）进行了大规模的遗传筛选，以识别编码对确保正确染色体分离至关重要的蛋白质的基因。在第一个特定目的中，我们将研究染色体动物学的几个新成分，这些成分首先在我们的屏幕中发现，并且在包括人类在内的后代人中一直保守。该目标中的一个重点是一个多生育络合物，包括蛋白质ZW10，Rod和Zwilch。该复合物将分子运动动力蛋白靶向动力学，并以当前未知的方式需要调节后期开始的主轴组件检查点的操作。我们提出了实验，以剖析这种复合物的结构，并研究其与裸体的相互作用，裸体是一种与动力蛋白相关的蛋白质，其作用目前尚不清楚。该目标中的第二个重点区域将探讨我们遗传筛选中鉴定出的两种新蛋白质可能会构成适当的染色体隔离所需的新蛋白。 第二个特定目的描述了我们实验室的新研究领域。在我们筛网中发现的果蝇基因之一的Greatwall突变，导致染色体的降压，这是由于通过预言的延迟而引起的。 Greatwall基因编码一种新型的蛋白激酶，具有在昆虫和脊椎动物中保守的异常结构。我们对爪蟾大壁蛋白进行了抗体，并使用了抗体。从卵母细胞提取物中耗尽这种激酶。令我们惊讶的是，Greatwall的耗竭阻止了提取物进入或保持M期。在第二个特定目标中，我们将跟进这些有趣的观察结果，以了解大沃尔在有丝分裂进程中的作用。我们提出了实验，以定义Greatwall的底物以及在有丝分裂过程中涉及大壁激活的上游激酶。其他实验将研究Greatwall在卵母细胞成熟中的发育作用，并将搜索与该激酶相关的其他蛋白质。关于Greatwall的拟议研究有可能提供有关脊椎动物细胞中有丝分裂进入的独特机械见解。