DNA MISMATCH REPAIR AND CANCER IN MURINE MODELS

超出所提供的空间。DNA错配修复（MMR）系统对于维持哺乳动物基因组的完整性是必不可少的，并且哺乳动物MMR基因中的突变导致癌症易感性增加和减数分裂失败。真核MMR是一个复杂的系统，需要几个MutS和MutL蛋白的相互作用来启动修复反应。最近的研究结果表明，哺乳动物MMR蛋白在许多过程中发挥作用，这些过程不仅包括碱基错配的修复，而且还包括响应DNA损伤的凋亡信号以及抑制不相同序列之间的重组。由于这些活动中的每一种都可能对预防基因组不稳定和癌症有不同的贡献，因此有必要研究其个体损失的影响。为了实现这一目标，我们建议创建携带致病性错义突变的小鼠品系，这些突变将破坏特定基序的功能，但不会完全破坏蛋白质。我们推测，有丝分裂MutS同源物在错配修复、凋亡和抗重组中的每一种不同功能都协同抑制肿瘤发生。该提案的具体目标是：1.生成在Msh2、Msh3和Msh6的ATP酶结构域中携带敲入突变的小鼠品系，并分析所得癌症易感性表型。该分析将阐明该基序在肿瘤抑制中的重要性，并且还用作产生其他Msh2、Msh3和Msh6敲入等位基因的模型。2.分析Msh基因敲入突变对体外DNA修复功能的影响，并确定小鼠组织中产生的体内突变表型。3.研究Msh 2、Msh 6和Msh 3基因敲入小鼠的DNA损伤反应。我们建议在小鼠胚胎成纤维细胞系和Msh基因敲入小鼠系的胃肠道中研究暴露于DNA损伤剂后的凋亡反应。此外，我们建议在无胸腺裸小鼠异种移植模型中分析来自基因敲入小鼠的胚胎干细胞和肿瘤细胞系对顺铂和其他化疗药物的敏感性。4.研究Msh2、Msh3和Msh6点突变对MutS复合物抗重组功能的影响。我们将研究DNA双链断裂诱导的胚胎干细胞染色体重组，以测试敲入突变是否导致小鼠DNA修复和抗重组功能的分离，并评估这种功能丧失对肿瘤发生的后果。性能现场=

EXCEED THE SPACE PROVIDED. The DNA mismatch repair (MMR) system is essential for maintaining the integrity of the mammalian genome and mutations in the mammalian MMR genes result in increased cancer susceptibility and meiotic failure. Eukaryotic MMR is a complex system that requires the interaction of several MutS and MutL proteins for the initiation of the repair reaction. Recent results suggest that the mammalian MMR proteins function in a number of processes which not only include the repair of base mismatches, but also the signaling of apoptosis in response to DNA damage as well as the suppression of recombination between non-identical sequences. Because it is likely that each of these activities contributes differentially to the prevention of genomic instability and cancer, it is necessary to study the impact of their individual loss. To accomplish this, we propose to create mouse lines that carry pathogenic missense mutations which will disrupt the function of a particular motif but not completely inactivate the protein. We hypothesize that each of the distinct functions of the mitotic MutS homologs in mismatch repair, apoptosis and anti-recombination cooperate to suppress tumorigenesis. The specific aims of this proposal are: 1. To generate mouse lines that carry knock-in mutations in the ATPase domains of Msh2, Msh3 and Msh6 and analyze the resulting cancer susceptibility phenotype. This analysis will elucidate the significance of this motif in tumor suppression and also serve as a model for the generation of other Msh2, Msh3 and Msh6 knock-in alleles. 2. To analyze the effect of the Msh knock-in mutations on DNA repair functions in vitro and determine the resulting in vivo mutator phenotype in mouse tissues. 3. To study the DNA damage response in the Msh2-, Msh-6 and Msh3- knock-in mouse lines. We propose to study the apoptotic response after exposure to DNA damaging agents in mouse embryonic fibroblast lines and in the in the gastrointestinal tract of the Msh knock-in mouse lines. In addition, we propose to analyze the sensitivity of embryonic stem cells and tumor cell lines derived from the knock-in mice to cisplatin and other chemotherapeutic drugs in a xenograft model in athymic nude mice. 4. To investigate the impact of the Msh2, Msh3 and Msh6 point mutations on the anti-recombination function of the MutS complexes. We will investigate DNA double strand break induced recombination in the chromosomes of embryonic stem cells to test whether the knock-in mutations result in a separation of DNA repair and anti-recombination functions in mice and assess the consequence of loss of this function on tumorigenesis. PERFORMANCE SITE ========================================Section End===========================================