The objective of this proposal is to gain an understanding of the mechanism whereby molecular oxygen regulates procaryotic gene expression. It is well established that changes in the environment can influence gene expression. In procaryotic organisms, the presence of molecular oxygen inhibits expression of genes involved in carbon fixation, nitrogen fixation, electron transport, and photosynthesis. Despite well documented cases of oxygen repression of gene expression, very little is known about the molecular mechanism of this form of gene regulation. In this proposal, a study of oxygen regulation of gene expression in the phototrophic bacterium Rhodobacter capsulatus will be undertaken using genetic and molecular biology techniques. Specifically, regions of DNA involved in oxygen regulation will be identified by comparing different oxygen regulated promoters for DNA sequence similarity. Regions of the promoters that are responsible for oxygen regulation will subsequently be characterized by in vivo and in vitro mutagenesis techniques. Besides characterizing cis-dominant features of DNA responsible for oxygen regulation, trans-acting protein components involved in oxygen regulation will be identified and characterized by several independent means. Specifically, trans-acting mutations that exhibit pleiotropic repression or derepression of oxygen regulated genes will be isolated using established selection procedures. These trans-acting mutants will subsequently be used to isolate and sequence trans-acting regulatory genes responsible for oxygen regulation. Finally, study of DNA-protein interactions will be undertaken to determine which trans-acting regulatory proteins interact with specific DNA regulatory sequences present in oxygen regulated promoters. It should be noted that a variety of diseases are genetic in origin, and thus, one goal of genetic research is to gain an understanding of the molecular events with which genes are regul- ated. Consequently, it is believed that basic research on procaryotic gene regulation is of such a fundamental nature that it can contribute significantly to our basic understanding of the mechanism of aberrant gene expression, which in many cases, is thought to result in genetic diseases.
