Structure and Regulation of thegabGene Cluster, Involved in the γ-Aminobutyric Acid Shunt, Are Controlled by a σ54Factor inBacillus thuringiensis

Abstract

ABSTRACTThe structure and regulation of thegabgene cluster, involved in γ-aminobutyric acid (GABA) shunt, were studied by characterizinggabTandgabDgenes cloned fromBacillus thuringiensis. Deletions of thegabTandgabDgenes inB. thuringiensisstrain HD-73 did not affect the growth of mutant strains in rich culture media, but the growth of agabTdeletion mutant strain was reduced in basic media (containing 0.2% GABA). Genome analysis indicates that the structure of thegabgene cluster inB. thuringiensisHD-73 is different from that inEscherichia coliandBacillus subtilisbut is common in strains of theBacillus cereusgroup. This suggests that the gene cluster involved in GABA shunt is specific to theB. cereusgroup. Based on reverse transcription-PCR and transcriptional fusion analysis, we confirmed that thegabTandgabDgenes belong to different transcriptional units, while thegabDandgabRgenes form an operon. We also demonstrated that thegabRgene plays a positive regulatory role ingabDandgabTexpression. The GabR protein may be a σ54-dependent transcriptional activator, according to a conserved domain search in the NCBI database, and it is highly conserved in theB. cereusgroup. The −24/−12 consensus sequence of a promoter upstream fromgabTsuggests that the promoter can be recognized by a σ54factor. Further analysis of the genetic complementation studies also suggests that the expression of thegabTgene is controlled by a σ54factor. Thus, the expression of thegabcluster is regulated by a σ54factor by way of the transcription activator GabR.