The level of DNA supercoiling, the intracellular concentration of ppGpp, and the temperature and constituents of growth media all affect gene expression in E. coli and S. typhimurium, in particular the ribosomal RNA operons (rrn) and histidine biosynthetic operon (his). These controls are termed universal controls because the majority of the genes in the bacterial chromosome respond to changes in these conditions. DNA sequence features have been proposed as being involved in the mediation of universal controls. Although over 25% of the E. coli genome has been sequenced, this information has not been organized in a cohesive fashion. A large scale computerized mapping system and associated database has been developed by us in order to be able to access E. coli DNA sequence information in a systematic fashion and to produce a better genetic map of the E. coli chromosome than has been previously available. Major findings this reporting period include: (1) Treatment of E. coli and S. typhimurium with DNA gyrase inhibiting antibiotics slows growth and derepresses the operon in a fashion that mimics the effects of graded DNA gyrase mutants. This effect can be detected with a simple plate test that should be useful in screening for new DNA gyrase antibiotics. (2) A series of mutations that reverse the growth rate defect caused by a mutation in the spoT gene all involve insertions of IS10 or DNA inversions in a small region in the center of the spoT gene, possible uncoupling two domains of the SpoT protein, one involved in ppGpp degradation and one involved in ppGpp synthesis. (3) A non-overlapping database of 1.3 million base pairs of the E. coli genome has been assembled from published and unpublished DNA sequences. These sequences and the genes they contain have been aligned to the genomic restriction map of E. coli, producing a high resolution, integrated genomic map as a valuable resource for the entire E. coli genetic community.
