Reverse transcription-PCR (RT-PCR) analysis revealed that this locus forms two operons:yodT (yodT-yodS-yodR-yodQ-yodPkamR) and kamA (kamA-yokU-yozE). The transcriptional start sites (TSSs) of the kamA gene were determined using 5= rapid amplification of cDNA ends (RACE). A typical ؊12/؊24 54 binding site was identified in the promoter P kamA , which is located upstream of the kamA gene TSS. A -galactosidase assay showed that P kamA , which directs the transcription of the kamA operon, is controlled by the 54 factor and is activated through the 54 -dependent transcriptional regulator KamR. The kamA operon is also controlled by K and regulated by the GerE protein in the late stage of sporulation. kamR and kamA mutants were prepared by homologous recombination to examine the role of the kam locus. The results showed that the sporulation rate in B. thuringiensis HD(⌬kamR) was slightly decreased compared to that in HD73, whereas that in HD(⌬kamA) was similar to that in HD73. This means that other genes regulated by KamR are important for sporulation.
Bacteria synthesize a number of different sigma factors that allow the coordinated expression of groups of genes due to the ability of sigma factors to confer promoter-specific transcriptional initiation of RNA polymerase (RNAP) (1). Most of these sigma factors belong to a single family of proteins that appear to be related structurally and functionally to the major Escherichia coli sigma factor, 70 . A second class of these factors is represented solely by the alternative factor 54 , which is widely present in prokaryotes (2, 3). Studies of 54 have showed that this sigma factor is quite distinct both structurally and functionally from the 70 family: (i) unlike the 70 factor, which recognizes Ϫ10/Ϫ35 regions (4), 54 directs its RNAP to promoters characterized by conserved sequences located at Ϫ24 and Ϫ12 bp upstream from the transcriptional start site (TSS) (5), and (ii) the activator ATPase approaches the leading edge of the closed RNAP-promoter complex, and a regulatory protein is required for 54 to stimulate the isomerization of the closed complexes to the corresponding open complexes (1, 3). 54 participates in the regulation of many metabolic pathways in bacteria, such as the arginine degradation pathway, the isoleucine and valine degradation pathway, and the acetoin catabolic pathway in Bacillus subtilis (6-8). We previously found that the gab gene cluster involved in the GABA shunt for the utilization of GABA is controlled by 54 in Bacillus thuringiensis (9), a process that is not 54 dependent in B. subtilis (10). However, other metabolic pathways controlled by the 54 factor in B. thuringiensis remain unknown.L-Lysine is first converted to L--lysine by a lysine-2,3-aminomutase (KAM; EC 5.4.3.2) in the lysine degradation pathway (11), and this intermediate is then acetylated to N ε -acetyl--lysine by the action of an acetyltransferase (12). Genes potentially encoding lysine-2,3-aminomutase (ablA) and -lysine acetyltransferase (ablB) have been identified on th...