Regulation Of Exosporium Genes By Transcription Factor Sigmak And GerE

Bacillus thuringiensis is a gram-positive and spore-forming bacterium, belonging to theBacillus cereus group. Bacillus cereus groups respond to nutrient limitation by forming an endospore.Spore from Bacillus cereus groups and Clostridium is encased in exosporium. It consists of aparacrystalline basal layer and an exernal hair-like nap. At present, the studies about exosporium focuson its composition, structure and function. Few studies about its transcription were reported. In thisstudy, we study on the transcription of exosporium genes by SigmaK and GerE, which are two globetranscriptional factors in mother cells during the late stage of sporulation.According to protein sequences of the exosporium in B. cereus and B. anthracis, We found10exosporium proteins in HD73strain with high similarity to those in B. anthracis and B. cereus,including hairy nap proteins BclA and BclB, basal layer proteins ExsY, ExsB, BxpB, CotY, CotB-1andBxpA, enzymes IUNH and Sod. We constructed expression vectors of10promoter-lacZ fusions andtransformed them into HD73, HDΔgerE and HDΔsigK, respectively.Beta-galactosidase assays showed that deletion of sigK gene decreased the activities of bxpB, exsB,exsY, bxpA, cotB-1, bclA, bclB and iunh promoters. It suggests that transcription of these genes iscontrolled by SigmaK during the late stage of sporultion. Deletion of gerE gene decreased the activitiesof bxpB, exsB, cotB-1, bclB and iunh promoters and increased the activaty of bxpA promoter. It meansthat transcription of bclB, exsB, bxpB, cotB-1and iunh is positively regulated and transcription of bxpAis negatively regulated by GerE. In order to identify these promoters directly regulated by GerE or not,GerE protein was expressed and purified. Electrophoretic mobility shift assays showed that GerE canbind to promoters of bxpB, exsB, cotB-1, bxpA, bclB and iunh. It indicates that these genes are directlyregulated by GerE.Compared the bxpB, exsB, bxpA, cotB-1, bclA and bclB promotors recognized by SigmaK insome strains of the B. cereus group, we found the promoters of bxpB, exsB, bxpA, cotB-1, bclA andbclB recognized by SigmaK are very conserved. It suggests these genes in these strains maybetranscriptionally regulated by SigmaK.