Cooperative roles for Atm and Hus1 in genome maintenance

项目摘要 共济失调-毛细血管扩张症是一种基因组不稳定综合征，其中DNA损伤突变 检查点蛋白Atm导致神经变性、癌症易感性、不育和免疫缺陷。 功能障碍Atm在细胞对双链DNA断裂（DSB）的反应中起核心作用， 而涉及相关检查点蛋白Atr的第二条途径响应复制应激， 大体积DNA损伤以及双链断裂这些途径之间的功能重叠程度是 部分原因是Atr途径的组成部分，其中还包括Chk 1 和Rad 9-Rad 1-Hus 1（911）复合物，对于生存力是必需的。的长期目标 本提案中所描述的研究旨在解决Atm和Atr途径之间的关系 并确定当Atm被激活时，Atr通路的活性如何影响疾病的发病机制。 有缺陷我们在小鼠中使用了Hus 1等位基因系列来鉴定Atm之间的遗传相互作用， 和ATR途径。当部分Hus 1损伤结合时，观察到合成致死率 与ATM的缺陷，建立一个必要的合作关系，两个主要的 哺乳动物DNA损伤检查点途径。尽管Hus 1表达的严重减少 与Atm缺失相结合是致命的，稍微高水平的Hus 1表达产生了存活的小鼠 频率低于预期。在目标一中，将确定胚胎致死率的基础 通过胚胎的形态学和组织学分析以及DNA损伤的检查， 在具有Atm和Hus 1缺陷的培养细胞中的信号传导。此外，存活的小鼠 同时的Atm和Hus 1缺陷将被测试神经退行性变，神经退行性变是一种突出的表型， 在Atm敲除小鼠中未观察到的共济失调-毛细血管扩张患者。在目标二中， 将评估在Atm缺陷小鼠中降低的Hus 1功能对肿瘤发展的影响。ATM 杂合性增加了人类患乳腺癌的风险，因此肿瘤发生将 也可以在具有部分Hus 1缺陷的杂合Atm小鼠中进行检查。总的来说， 拟议的研究将阐明两种主要哺乳动物DNA损伤之间的关系， 检查点途径和解决这些基因组维护机制的合作作用 在执行发育程序，预防神经退行性变， 肿瘤发生相关性 共济失调-毛细血管扩张症是一种毁灭性疾病，估计发生率为1/40，000。的 拟议中的研究将确定这种疾病的严重程度， 个体之间的相互作用受到相关通路活动的影响。这些研究还将澄清 负责维持基因组完整性的信号网络的组织， 为将来的转化研究产生更准确的共济失调-毛细血管扩张症小鼠模型。

PROJECT SUMMARY Ataxia-Telangiectasia is a genomic instability syndrome in which mutations in the DNA damage checkpoint protein Atm cause neurodegeneration, cancer predisposition, infertility, and immune dysfunction. Atm plays a central role in cellular responses to double-stranded DNA breaks (DSB), while a second pathway involving the related checkpoint protein Atr responds to replication stress and bulky DNA lesions, as well as DSB. The extent of functional overlap between these pathways is incompletely understood in part because components of the Atr pathway, which also includes Chk1 and the Rad9-Rad1-Hus1 (911) complex, are essential for viability. The long-term objectives of the research described in this proposal are to resolve the relationship between the Atm and Atr pathways and to determine how the activity of the Atr pathway affects disease pathogenesis when Atm is defective. We employed a Hus1 allelic series in mice to identify genetic interactions between the Atm and Atr pathways. Synthetic lethality was observed when partial Hus1 impairment was combined with Atm deficiency, establishing an essential cooperative relationship between the two primary mammalian DNA damage checkpoint pathways. Although a severe reduction in Hus1 expression was lethal in combination with Atm loss, a slightly higher level of Hus1 expression yielded viable mice at less than expected frequency. In aim one, the basis for the embryonic lethality will be determined by morphological and histological analysis of embryos as well as by examination of DNA damage signaling in cultured cells with both Atm and Hus1 defects. In addition, surviving mice with simultaneous Atm and Hus1 defects will be tested for neurodegeneration, a prominent phenotype of Ataxia-Telangiectasia patients that is not observed in Atm knockout mice. In aim two, the impact of reduced Hus1 function on tumor development in Atm-deficient mice will be assessed. Atm heterozygosity confers an increased risk of breast cancer in humans, and therefore tumorigenesis will also be examined in heterozygous Atm mice with a partial Hus1 defect. Taken together, the proposed studies will clarify the relationship between two primary mammalian DNA damage checkpoint pathways and resolve the cooperative roles for these genome maintenance mechanisms in executing developmental programs, preventing neurodegeneration, and suppressing tumorigenesis. RELEVANCE Ataxia-Telangiectasia is a devastating disease with an estimated frequency of 1 in 40,000 births. The proposed research will determine how the severity of this disease, which can vary significantly between individuals, is affected by the activity of a related pathway. The studies also will clarify the organization of the signaling networks responsible for the maintenance of genomic integrity and may produce a more accurate mouse model for Ataxia-Telangiectasia for future translational studies.