Roles of Mre11 in lymphocyte development and DNA repair

描述(申请人提供)：遗传Mre11基因突变的人类对电离辐射极其敏感，发展为小脑变性，并在循环中的淋巴细胞中有频繁的染色体易位。此外，已发现Mre11的体细胞突变与多种肿瘤有关。Mre11蛋白是参与DNA修复和整个细胞对DNA损伤反应的复合体的成员。该复合体由3种蛋白质组成：Mre11、Rad50和Nbs。Mre11和Rad50的同源物从细菌到后生动物都是保守的。NBS(酵母中的XRS2)在氨基酸水平上不太保守，但仍然是必需的。体外生化研究和酵母体内分析表明，Mre11具有单链DNA内切酶、单链DNA外切酶和双链DNA结合活性。该蛋白质是模块化的，因为所有的核酸酶活性都限制在N末端，而双链DNA结合则限制在C末端。关于哺乳动物中的Mre11仍有许多有待了解的地方，因为在常见的实验系统中，零等位基因对细胞的致命性，以及对ATLD患者含有维持显著蛋白质功能的突变等位基因的认识，阻碍了研究。这个问题尤其突出的是，缺乏任何现有的哺乳动物突变，使Mre11的内切和外切核酸酶活性失活，这是Mre11、Rad50、NBS复合体最重要的功能之一。在此，描述了第一只缺乏mre11核酸酶活性的小鼠的完整构建。提出的实验将回答许多问题，涉及高度保守的Mre11核酸酶在正常细胞生长过程中的作用，对DNA损伤的反应，以及在淋巴细胞和配子细胞中的特殊重组事件中的作用。这些研究可能有助于我们理解多功能的Mre11蛋白及其在人类健康和疾病中的作用。

DESCRIPTION (provided by applicant): Humans that inherit mutations in the Mre11 gene suffer from extreme sensitivity to ionizing radiation, develop cerebellar degeneration, and have frequent chromosomal translocations in circulating lymphocytes. In addition, somatic mutation of Mre11 has been found in association with various tumors. The Mre11 protein is a member of a complex involved in DNA repair and in the overall cellular responses to DNA damage. 3 proteins comprise this complex; Mre11, Rad50 and NBS. Homologues of Mre11 and Rad50 are conserved from bacteria to metazoans. NBS (XRS2 in yeast) is less well conserved at the amino acid level, but is still required. In vitro biochemical studies and in vivo analyses in yeast have demonstrated that Mre11 possesses single strand DNA endonuclease, single strand DNA exonuclease, and double strand DNA binding activities. The protein is modular in that all nuclease activities are confined to the N terminal region while double strand DNA binding is confined to the C terminus. Much remains to be learned about Mre11 in mammals because studies have been hampered by the cellular lethality of null alleles in common experimental systems, and the realization that ATLD patients harbor mutant alleles that maintain significant protein function. This problem is especially highlighted by the lack of any existing mammalian mutation that inactivates the endo-and exonuclease activities of Mre11 arguably 1 of the most important functions of the Mre11, Rad50, NBS complex. Herein is described the completed construction of the first mouse deficient for the nuclease activities of Mre11. Experiments are proposed which will answer many questions regarding the roles of the highly conserved Mre11 nuclease during normal cell growth, in response to DNA damage, and in specialized recombination events in lymphocytes and gametocytes. These studies may contribute significantly to our understanding of the multifunctional Mre11 protein and its roles in human health and disease.