Roles of Mre11 in lymphocyte development and DNA repair

描述（由申请人提供）：遗传Mre11基因突变的人对电离辐射极度敏感，发生小脑变性，循环淋巴细胞染色体易位频繁。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修复和对DNA损伤的整体细胞反应的复合体的成员。3种蛋白质组成这个复合体；Mre11， Rad50和NBS。从细菌到后生动物，Mre11和Rad50的同源物都是保守的。NBS（酵母中的XRS2）在氨基酸水平上保守性较差，但仍是必需的。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。该蛋白是模块化的，因为所有的核酸酶活性都局限于N端区域，而双链DNA结合则局限于C端。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，零等位基因的细胞致命性阻碍了研究，并且认识到ATLD患者含有维持重要蛋白质功能的突变等位基因。由于缺乏任何现有的哺乳动物突变，使Mre11的内切酶和外切酶活性失活，这一问题尤其突出，Mre11, Rad50， NBS复合物的最重要功能之一是失活。本文描述了第一只缺乏Mre11核酸酶活性的小鼠的完整构建。本文提出的实验将回答有关高度保守的Mre11核酸酶在正常细胞生长、DNA损伤反应以及淋巴细胞和配子细胞特异性重组事件中的作用的许多问题。这些研究可能有助于我们了解多功能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.