DNA REPAIR AND REPLICATION RE START IN VIVO

遗传不稳定是导致 癌症、肿瘤进展和化疗药物耐药性。 涉及的染色体变化包括同源基因之间的体细胞重组 染色体，促进杂合性丢失，易位在 非同源染色体、基因扩增和其他DNA重排 除了突变。这些不同形式的遗传不稳定是由 通过DNA修复事件。许多与遗传有关的人类蛋白质 不稳定性在真细菌中有同源物，看起来功能相似， 其中许多参与DNA修复，包括双链修复 断裂(DSB)和单链间隙。导致损伤的药物 需要修复的是主要致癌物质，以及人类遗传性疾病 细菌修复蛋白的同源物被改变，显示出癌症的倾向。 尽管基因重排和DNA复制一直被认为是 独立的事件，并已被单独研究，最近的许多线条 来自几个生物体的证据现在暗示了一种深刻的联系--促进 DNA复制的双链DNA断裂修复(DSBR)。只有一次 细胞(非病毒)系统这种联系是否牢固而直接 证明：在大肠杆菌中DSBR发生的时间大约为 复制促进事件，另一半通过重组而不是 复制。在这项建议中，DSBR在大肠杆菌中的分子机制 研究模型系统的目的是阐明DNA的结构 过程中的中间体和产品，确定所有相关的 参与的蛋白质，并了解复制与 非复制机制。这些信息将与基因相关 癌症的不稳定性，在某些情况下，人类大肠杆菌的直系物 修复蛋白也将被检测。因为对DNA的直接物理分析， 在这个模型系统中，除了有复杂的遗传工具外， 将获得的分子机制的详细程度是其他 有机体。因为到目前为止鉴定出的蛋白质与人类同源 肿瘤蛋白(例如BLM、WRN、hRad51)，它与BRCA蛋白在 DSBR)，并且它们的功能是保守的，信息直接 适用于引起、促进和促进遗传不稳定的机制 制造抗药性的人类癌症。

Genetic instability is a primary cellular event leading to cancers, tumor progression, and resistance to chemotherapeutic drugs. Chromosomal changes involved include somatic recombination between homologous chromosomes, promoting loss of heterozygosity, translocations between non-homologous chromosomes, gene amplification, and other DNA rearrangements in addition to mutation. These various forms of genetic instability are provoked by DNA repair events. Many of the human proteins implicated in genetic instability have homologs in eubacteria which appear to function similarly, many of which participate in DNA repair including repair of double-strand breaks (DSBs) and single-strand gaps. Agents that cause the lesions that require repair are primary carcinogens, and human genetic diseases in which homologs of the bacterial repair proteins are altered display cancer-proneness. Although genetic rearrangements and DNA replication have been thought of as separate events, and have been studied separately, many lines of recent evidence from several organisms now imply a profound connection--the promotion of DNA replication by double-strand DNA break-repair (DSBR). In only one cellular (non-viral) system has this connection been firmly and directly demonstrated: DSBR in E. coli occurs roughly half the time as a replication-promoting event, and the other half via recombination without replication. In this proposal the molecular mechanism of DSBR in the E. coli model system is investigated with the goals of elucidating structures of DNA intermediates in and products of the process, identifying all of the relevant proteins involved, and understanding control of replicative versus non-replicative mechanisms. This information will be related to genetic instability in cancer, and, in some cases, the human orthologs of E. coli repair proteins will also be examined. Because direct physical analysis of DNA, as well as sophisticated genetic tools are available in this model system, the level of detail of molecular mechanism to be obtained is unparalleled in other organisms. Because the proteins so far identified are homologous with human cancer proteins (e.g. Blm, Wrn, hRad51 which functions with Brca proteins in DSBR), and their functions are conserved, the information is directly applicable to the mechanisms of genetic instability that cause, promote, and make drug-resistant, human cancers.