The mechanisms organisms used to adapt to osmotic stress are not understood very extensively. The experiments in this grant proposal are aimed at the elucidation of two aspects of osmotic adaptation in the bacterium Salmonella typhimurium. Two compounds, proline and glycinebetaine, can alleviate the inhibitory effects of osmotic stress in S. typhimurium. These two compounds are taken up by a transport system that is encoded in the proU operon, which is subject to osmotic regulation such that the exposure of the cells to osmotic stress results in a 200-fold induction of its transcription. We shall carry out experiments to elucidate the mechanism of the transcriptional control of the proU operon. The mode of action of proline and glycinebetaine in alleviating osmotic inhibition is not known. We isolated S. typhimurium mutants which are not stimulated by glycinebetaine in media of high osmolarity. We shall carry out a genetic and biochemical analysis of these mutants in order to discover the mechanism whereby glycinebetaine can overcome the inhibitory effects of osmotic stress in the wild type strain. Because there are close similarities in the cellular responses of plants and bacteria to osmotic stress, the insights gained into the osmotic adaptation in bacteria will aid the understanding of this process in plants. Our analysis of the transcriptional control of the proU operon also has potential applications for biotechnology. The promoter of this operon could be fused to desired genes of industrial utility. Over- production of the proteins encoded by these genes could be triggered by the addition of high concentrations of nutrients, which would be required at the same time for the production of large quantities of cells.
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