The objective of this proposal is to elucidate the organization and expression of tRNA genes from Bacillus subtilis. B. subtilis is chosen for study because it exhibits interesting differences from Escherichia coli, the current prototype for bacterial systems. Of special interest is the transcriptional regulation during development within the two rRNA-tRNA operons of B. subtilis, which contain large clusters of tRNA genes. Their function as promoter elements will be validated using the promoter probe plasmid system pDT. Promoter elements will be synthesized or amplified with the polymerase chain technique. The activity of these promoters will then be tested during development in B subtilis. Promoter activity will be evaluated by measuring levels of lacZ mRNA. If these promoters show temporal regulation, the protein factors affecting transcriptional regulation and flanking regions to which they bind will be investigated. These studies will greatly aid our understanding of tRNA gene expression, one of the key steps in the regulation of the growth of bacteria and other cells.
