Genome maintenance by the mouse Hus1 checkpoint gene

描述（由申请人提供）：细胞持续经历基因组损伤，如果不适当处理，可能导致发育缺陷，早衰和增加癌症易感性。DNA损伤检查点通过调节细胞周期转变、细胞凋亡和DNA修复来抵御基因组不稳定的这些后果。在哺乳动物中，检查点蛋白Hus1是响应复制应激和各种DNA损伤的途径的关键组成部分。Hus1依赖通路对胚胎发育至关重要，因此其在动物中的许多生物学功能尚不明确。本研究的长期目标是了解hus1依赖性检查点通路如何保护基因组完整性，并确定该机制中的缺陷如何影响肿瘤发生和生理DNA损伤反应。提出的研究将通过在小鼠Hus1基因组位点上进行独特的靶向修饰来推进这些重要目标。目的一解决部分或完全Hus1失活如何影响激活癌基因诱导的肿瘤发展，以解决Hus1作为候选肿瘤抑制因子和癌细胞生存和增殖所需因子的对立作用。在第二项研究中，在体内DNA损伤反应中对Hus1的需求将通过测量Hus1表达降低的小鼠的组织特异性基因毒性应激反应来确定，部分目的是为了获得检查点功能如何影响对DNA损伤抗癌治疗的敏感性的新见解。在前两个目标中对检查点功能障碍的生理后果的分析将被第三个目标中对Hus1分子机制的研究所补充，以努力定义对基因毒性应激生存至关重要的Hus1功能，并确定Hus1序列变异如何影响基因组维持。总之，拟议的研究将定义一个基本的检查点机制如何在肿瘤发生和组织特异性DNA损伤反应中起作用，并将为自发突变、自然多态性或药物抑制引起的检查点功能障碍的生物医学意义提供见解。公共卫生相关性：通过防御基因组损伤，DNA损伤检查点对正常胚胎发育、对环境压力和DNA损伤化疗的保护性反应以及成年生物体的无肿瘤存活至关重要。本实验利用新型基因工程小鼠品系分析了重要的DNA损伤检查点基因Hus1的生物学功能。这些研究对于提高对癌症如何发展，正常细胞和肿瘤细胞如何对抗癌治疗作出反应，以及这些过程如何受到Hus1序列变异（包括自发突变和自然多态性）的影响的理解具有很大的希望。

DESCRIPTION (provided by applicant): Cells continuously experience genome damage that, if not properly attended to, can cause developmental defects, premature aging, and increased cancer predisposition. DNA damage checkpoints defend against these consequences of genomic instability by regulating cell cycle transitions, apoptosis, and DNA repair. In mammals, the checkpoint protein Hus1 is a critical component of a pathway that responds to replication stress and a variety of DNA lesions. The Hus1- dependent pathway is essential for embryonic development, and consequently many of its biological functions in animals are not well-defined. The long-term goals of the research described in this proposal are to understand how the Hus1-dependent checkpoint pathway protects genomic integrity and to determine how defects in this mechanism affect tumorigenesis and physiological DNA damage responses. The proposed studies will advance these important objectives using a unique collection of targeted modifications at the mouse Hus1 genomic locus. Aim one addresses how partial or complete Hus1 inactivation affects tumor development induced by activated oncogenes in order to resolve the opposing roles of Hus1 as a candidate tumor suppressor and a factor required for cancer cell survival and proliferation. In Aim Two, the requirements for Hus1 during in vivo DNA damage responses will be determined by measuring the tissue-specific genotoxic stress responses of mice with reduced Hus1 expression, in part to gain new insights into how checkpoint function impacts sensitivity to DNA damaging anti-cancer therapies. These analyses of the physiological consequences of checkpoint dysfunction in the first two aims will be complemented by the investigation of Hus1 molecular mechanisms in Aim Three, in an effort to define the Hus1 functions that are essential for the survival of genotoxic stress and to establish how Hus1 sequence variations affect genome maintenance. Together, the proposed studies will define how an essential checkpoint mechanism functions during tumorigenesis and tissue-specific DNA damage responses, and will provide insights into the biomedical implications of checkpoint dysfunction stemming from spontaneous mutations, natural polymorphisms, or pharmacological inhibition. PUBLIC HEALTH RELEVANCE: By defending against damage to the genome, DNA damage checkpoints are critical for normal embryonic development, protective responses to environmental stresses and DNA damaging chemotherapeutics, and the tumor-free survival of adult organisms. The experiments of this proposal make use of novel genetically-engineered mouse strains to analyze the biological functions of the essential DNA damage checkpoint gene Hus1. These studies hold great promise for improving the understanding of how cancers develop and how normal and neoplastic cells respond to anti-cancer therapies, as well as how these processes are influenced by Hus1 sequence variations, including spontaneous mutations and natural polymorphisms.