Mechanisms of Adaptive Amplification

描述（由申请人提供）：应激细胞经历许多生理变化，使它们能够忍受条件。现在越来越清楚的是，它们也经历了基因变化，而且基因变化的速度作为应激反应的一部分而加快。这种由压力引起的基因变化过程被称为适应性突变。细菌的适应性点突变过程已经得到了很好的研究，并表明它不同于生长依赖性突变。在另一个适应性过程中，即适应性基因扩增，细胞会复制一定长度的DNA，这种DNA携带的基因在高水平表达时是有利的。该项目研究了大肠杆菌适应性扩增的分子机制，验证了这样一个假设：扩增的起始事件是复制分叉停止，当它们重新开始时，复制错误的模板，从而复制了一段基因组长度。遗传和分子操作用于探索这一假设，并了解在停滞复制分叉处发生的事件。只有当细胞表达控制细胞一般应激反应的因子时，适应性扩增才会发生。通过研究哪些应激反应的成分需要扩增，这使得研究细胞在应激反应中诱导这些遗传变化的事件成为可能。这项研究的结果将与许多在压力下发生遗传变化的情况有关。这些包括致病性的进化，病原体和肿瘤细胞中耐药性的发展，以及肿瘤的发展。复制叉的停滞也与重复不稳定性相关遗传疾病的基础过程有关。细菌和人类之间高度保守的多重DNA修复过程的发现，以及在不易处理的人体系统中缺乏机制信息，强调了这项工作与人类健康的许多方面的相关性。

DESCRIPTION (provided by applicant): Stressed cells undergo many physiological changes that allow them to tolerate the conditions. It is now becoming clear that they also undergo genetic change, and that the rate of genetic change is accelerated as a part of the stress-response. This process of stress-induced genetic change is called adaptive mutation. The processes of adaptive point mutation in bacteria have been well studied and shown to be distinct from growth-dependent mutation. In another adaptive process, adaptive gene amplification, the cell makes many copies of a length of DNA carrying a gene that is advantageous when expressed at a high level. This project investigates the molecular mechanism of adaptive amplificiation in Escherichia coli, testing the hypothesis that the initiating event of amplification is that replication forks stall and that, when they restart, the wrong template is copied, thus duplicating a length of the genome. Genetic and molecular manipulation is used to explore this hypothesis and to learn about the events occurring at stalled replication forks. Adaptive amplification only occurs if the cells are expressing the factors that control the cell's general stress response. This allows investigation of the events by which a cell induces these genetic changes in response to stress by studying which components of stress responses are needed for amplification. The results of this study will be relevant to many situations in which genetic change occurs under stress. These include the evolution of pathogenicity, the development of drug resistance in pathogens and in tumor cells, and the development of tumors. Replication fork stalling is also relevant to the processes that underlie repeat instability-related genetic diseases. The discoveries of multiple DNA repair process that are highly conserved between bacteria and humans, and the paucity of mechanistic information in the less tractable human system, underscore the relevance of this work to may aspects of human health.