Transcription and regulation of rocE in Bacillus thuringiensis

Abstract:

[Objective] rocE encodes arginine permease in arginine degradation pathway. To determine the mechanism of transcriptional regulation of rocE, we analyzed the transcriptional activity of rocE in Bacillus thuringiensis (Bt). [Methods] Transcriptional units of rocE gene cluster were analyzed by RT-PCR. Transcriptional activity of rocE promoter (ProcE) was analyzed by β-galactosidase assay. rocE mutant of Bt HD73 strain was constructed by homologous recombination. The HTH domain of RocR with His fusion protein was purified by HiTrap chelating column. The binding of rocE promoter with RocR-HTH protein was verified by electrophoresis mobility shift assays. [Results] Transcriptional activity of ProcE was induced by arginine in M9 medium. The transcriptional activity of ProcE was sharply decreased in sigL (encodes Sigma 54) and rocR mutants compared with that in HD73 wild-type in Schaeffer's sporulation medium (SSM) and arginine induced medium. ProcE could bind to RocR-HTH protein. Mutation of rocE had no significant differences on growth and Cry1Ac protein production. However, the sporulation efficiency of the rocE mutant was 65.5%, and that of the HD73 strain was 85.7%. The results of significance analysis show that the difference was significant (P<0.05). [Conclusion] Transcriptional activation of rocE is controlled by Sigma 54, and positive regulated by RocR. rocE gene is related to sporulation efficiency.

Key words:Bacillus thuringiensis;arginine metabolism;rocE;transcriptional regulation