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. ? ? ?
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