The understanding of how gene expression is controlled is fundamental in elucidating the mechanisms involved in normal and abnormal cellular growth and differentiation. Recent developments indicate that a number of different regulatory circuits are operative in controlling individual bacterial operons and in integrating the products of these pathways into the general metabolism of the cell. These data suggest that these biosynthetic systems may be valuable models for discovering how gene expression is regulated in complex as well as simple organisms. With this in mind we wish to continue out study of the mechanisms involved in control of the ilvB operon in E. coli K-12. A variety of regulatory elements are operative in this system including multi- valent endproduct attenuation by multiple species of tRNA, guanosine 3', 5'-bispyrophosphate, cyclic AMP and integration host factor. Studies on the mechanism of the individual control systems as well as how these systems interact will be investigated both in vivo and in vitro. The sequence of the promoter-regulatory region has been determined, and cloned fragments will be used in transcription studies to confirm the sites and nature of the promoter and regulatory region. In vivo investigators will focus on hybridization experiments with radioactively labelled RNA and ilvB DNA fragment probes which have defined regions of the regulatory-promoter region. In addition, the mechanism and interactions of the various regulatory elements will be investigated in a coupled in vitro transcription-translation system.
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