Regulation of the Topoiomerase II-Dependent G2 Decatenation Checkpoint

描述（由申请人提供）：乳腺癌是美国女性中最常见的癌症类型，也是癌症相关死亡的第二大原因。最近的证据表明，存在几种不同的乳腺癌亚型，每种类型都是一种单独的疾病，有特定的治疗过程。预后最差的亚型是基底样亚型，包括那些携带BRCA1突变的亚型。BRCA1被认为在细胞周期控制的几乎所有方面都起作用，并在维持基因组稳定性方面起关键作用，这表明它的信号机制是防止恶性进展所必需的。BRCA1是decatenation检查点的一个组成部分，该检查点阻止有丝分裂进入，直到纠缠的姐妹染色单体被充分decatenation，这一过程由拓扑异构酶Ila酶活性介导。这个检查点的失败会导致染色单体断裂和/或非整倍体，从而导致癌细胞的产生。本研究的长期目标是确定负责人类十十烷化检查点和拓扑异构酶Ila调节的主要信号分子。该应用的具体目标包括开发一种遗传系统，在该遗传系统中可以在人乳腺上皮细胞中研究十十烷化检查点，并随后研究BRCA1/BARD1 E3泛素连接酶复合物在十十烷化检查点信号传导中的作用。我们的假设认为，十烷化检查点的失败可能导致非整倍体和染色单体断裂，而拓扑异构酶lla蛋白和BRCA1/BARD1复合体是这一过程的重要组成部分。免疫荧光显微镜、中期准备、免疫印迹、流式细胞术和siRNA消耗技术将被用于分析核结构、染色体结构和有丝分裂进入，以确定这些信号蛋白对十烯二化检查点的影响。这些实验将提供一个遗传系统，用于研究与十十烷化检查点有关的信号通路，并为十烷化检查点的机制提供可靠的遗传证据，特别是基底样乳腺癌。相关性：乳腺癌是美国女性最常见的癌症类型。该提案旨在了解可能负责启动导致乳腺癌的分子变化的信号机制，并有助于设计可能有助于预防乳腺癌进展的特定化疗药物。

DESCRIPTION (provided by applicant): Breast cancer is the most common type of cancer occurring among women in the United States and the second highest cause of cancer-related deaths. Recent evidence has suggested that several different subtypes of breast cancer exist and that each type presents itself as an individual disease with a specific course of treatment. The subtype with the least favorable prognosis is the basal-like subtype which includes those harboring a BRCA1 mutation. BRCA1 is thought to function in almost every aspect of cell cycle control and plays a pivotal role in maintaining genomic stability, suggesting that its signaling mechanisms are required to prevent malignant progression. BRCA1 is an integral part of the decatenation checkpoint, which prevents entry into mitosis until entangled sister chromatids are sufficiently decatenated, a process mediated by topoisomerase Ila enzymatic activity. Failure of this checkpoint can result in chromatid breakage and/or aneuploidy and thus contribute to the generation of cancer cells. The long term goal of this research is to define the major signaling molecules responsible for the human decatenation checkpoint and topoisomerase Ila regulation. Specific aims of this application include the development of a genetic system in which the decatenation checkpoint can be studied in human mammary epithelial cells and subsequent investigation of the role of the BRCA1/BARD1 E3 ubiquitin ligase complex in decatenation checkpoint signaling. Our hypothesis states that a failure of the decatenation checkpoint may lead to aneuploidy and chromatid breakage, and that the topoisomerase lla protein and BRCA1/BARD1 complex are essential components of this process. Immunofluorescence microscopy, metaphase preparations, western immunoblotting, flow cytometry, and siRNA depletion techniques will be employed to analyze nuclear structure, chromosomal structure, and mitotic entry to determine the effects of these signaling proteins on the decatenation checkpoint. These experiments will provide a genetic system for the study of the signaling pathways involved in the decatenation checkpoint and generate solid genetic evidence regarding the mechanisms of the decatenation checkpoint with a specific focus on basal-like breast cancers. RELEVANCE: Breast cancer is the most common type of cancer afflicting women in the United States. This proposal seeks to understand signaling mechanisms that may be responsible for initiating molecular changes leading to breast cancer and contribute to the design of specific chemotherapeutic drugs that may help to prevent breast cancer progression.