Enzymology Of Eukaryotic Mismatch Repair

描述(由申请人提供)：DNA错配修复是一种关键的突变避免途径，有几个原因引起临床关注。错配修复失活是一种常见形式的遗传性结肠癌的原因，并与一组散发性肿瘤的发展有关。错配修复缺陷也对癌症治疗有影响，因为该途径的失活使细胞对某些抗肿瘤药物的细胞毒作用产生抗药性，这是系统参与DNA损伤反应的结果。也许令人惊讶的是，错配修复功能也是产生某些突变所必需的，例如(CAG)n重复序列的扩展，这是许多神经退行性疾病的主要原因。通过阐明人类错配修复的分子本质，我们希望了解它在控制突变发生中的作用。为此，我们提出了四项工作：(1)有关链导向的人类错配修复性质的现有信息表明，反应的过程由一组进化的蛋白质-蛋白质和蛋白质-DNA相互作用决定，错配识别活动MutS1和MutS2启动的修复事件在显著方面不同。通过分析选定的多蛋白和多蛋白-DNA复合体的性质，我们希望进一步阐明MutS1和MutS2启动的修复事件的机制。(2)(CAG)n神经退行性疾病的躯体扩张期可发生在有丝分裂后细胞中，这取决于错配修复活性MutS2和MutL1，提示修复DNA合成参与了这一过程。在提取和纯化系统中都将讨论人类错配修复系统处理(CAG)n重复元件的性质。(3)对SN1DNA甲基化子的检验点和凋亡反应需要错配修复功能。使用生化方法，我们将探索依赖MutS1和MutL1激活ATR损伤信号激酶的机制，以响应O6-甲基鸟嘌呤，O6-甲基鸟嘌呤是此类药物产生的主要细胞毒性损伤。(4)与Lorena Beese实验室的合作研究将解决人类错配识别系统识别和处理病变的结构基础。

DESCRIPTION (provided by applicant): DNA mismatch repair is a key mutation avoidance pathway that is of clinical interest for several reasons. Inactivation of mismatch repair is the cause of a common form of hereditary colon cancer and has been implicated in the development of a subset of sporadic tumors. Mismatch repair defects also have implications for cancer therapy because inactivation of the pathway renders cells resistant to the cytotoxic effects of certain anti-tumor drugs, a consequence of participation of the system in the DNA damage response. Perhaps surprisingly, mismatch repair function is also required for the production of certain mutations, such as the expansion of (CAG)n repeat sequences, the primary cause of a number of neurodegenerative diseases. By elucidating the molecular nature of human mismatch repair, we hope to understand its roles in controlling the occurrence of mutation. To this end we propose four lines of work: (1) Available information on the nature of strand-directed human mismatch repair indicates that the course of the reaction is dictated by an evolving set of protein-protein and protein-DNA interactions, and that repair events initiated by the mismatch recognition activities MutS1 and MutS2 differ in significant ways. By analyzing the nature of selected multi-protein and multi-protein-DNA complexes, we hope to further clarify the mechanisms of MutS1- and MutS2-initiated repair events. (2) The somatic expansion stage of (CAG)n neurodegenerative diseases, which depends on the mismatch repair activities MutS2 and MutL1, can occur in postmitotic cells, suggesting involvement of repair DNA synthesis in this process. The nature of processing of (CAG)n repeat elements by the human mismatch repair system will be addressed in both extract and purified systems. (3) Mismatch repair function is required for checkpoint and apoptotic responses to SN1 DNA methylators. Using a biochemical approach, we will pursue the mechanisms of MutS1- and MutL1-dependent activation of the ATR damage-signaling kinase in response to O6-methylguanine, the primary cytotoxic lesion produced by this class of drug. (4) Collaborative studies with the laboratory of Lorena Beese will address the structural basis of lesion recognition and processing by the human mismatch recognition system.