The objective is the characterization of accessory factors that influence transcription One major goal is to extend the characterization of the functions and interactions influencing regulation of gene expression b the N protein of coliphage lambda. This interaction, which results in the generation of a transcription complex that overrides transcription termination signals (transcription antitermination), requires in addition to the lambda-encoded N protein, host Nus proteins, and an RNA site, NUT. Genetic, physiological, and biochemical studies are directed toward understanding how the participants interact to form a termination- resistant transcription complex. In addition to continuing studies on the roles of functions previously identified with N-mediated antitermination, we propose to study the roles of other factors directly implicated in modulating N action. These include a putative inhibitor of N action and the alpha subunit of RNA polymerase. A newly developed technology based on surface plasmon resonance that allows measurements of molecular interactions in real time will be employed to study interactions between participants in N-mediated antitermination. A second major goal is to extend studies on the E. coli 10Sa RNA. Our work suggests a previously unknown role for this RNA, binding of DNA- binding proteins. We propose to determine its structure of 10Sa RNA and characterize the nature of its interactions with various DNA-binding proteins, characterize the regulation its synthesis. This combination of genetic, biochemical and physiological approaches should continue to provide new information about transcription antitermination. The lambda N system will undoubtedly remain a useful model for studies on eucaryotic gene expression systems thought to be regulated by antitermination, such as the TAT regulation of HIV expression and c-myc transcription. 10Sa RNA is highly conserved and thus may play an important physiological role in nearly all, if not all, bacteria. It could play an important role in maximizing bacterial growth and thus could be a important auxiliary factor fostering pathogenesis.
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