Construction of a Promoter-probe Vector for Bacillus thuringiensis: the Identification of cis-acting Elements of the chiA Locus

Xie, Chichu; Luo, Yuanchan; Chen, Yuehua; Cai, Jin

doi:10.1007/s00284-012-0100-0

Cited by 11 publications

(11 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In our previous work, we also found another chitinase gene (chiA) in Bti75 (47). We found that there is also a 16-bp sequence (ATACATCTAGACAACT) (dre chiA ), which is similar to dre Bacillus downstream of the core promoter region of chiA.…”

Section: Discussionmentioning

confidence: 67%

YvoA and CcpA Repress the Expression of chiB in Bacillus thuringiensis

Jiang

Pan

et al. 2015

Appl Environ Microbiol

Self Cite

View full text Add to dashboard Cite

c Bacillus thuringiensis produces chitinases, which are involved in its antifungal activity and facilitate its insecticidal activity. In our recent work, we found that a 16-bp sequence, dre chiB (AGACTTCGTGATGTCT), downstream of the minimal promoter region of the chitinase B gene (chiB) was a critical site for the inducible expression of chiB in B. thuringiensis Bti75. In this work, we show that a GntR family transcriptional regulator (named YvoA Bt ), which is homologous to YvoA of Bacillus subtilis, can specifically bind to the dre chiB oligonucleotide sequences in vitro by using electrophoretic mobility shift assays (EMSAs) and isothermal titration calorimetry (ITC) assays. The results of quantitative real-time reverse transcription-PCR (qRT-PCR) and Western blotting indicated that deletion of yvoA caused an ϳ7.5-fold increase in the expression level of chiB. Furthermore, binding of purified YvoA Bt to its target DNA could be abolished by glucosamine-6-phosphate (GlcN-6-P). We also confirmed, in the presence of the phosphoprotein Hpr-Ser 45 -P, that purified CcpA Bt bound specifically to the promoter of chiB, which contains the "cre chiB " sequence (ATAAAGCGTTTACA). According to the results of qRT-PCR and Western blotting, deletion of ccpA resulted in a 39-fold increase in the chiB expression level, and glucose no longer influenced the expression of chiB. We confirm that chiB is negatively controlled by both CcpA Bt and YvoA Bt in Bti75.

show abstract

Section: Discussionmentioning

confidence: 67%

YvoA and CcpA Repress the Expression of chiB in Bacillus thuringiensis

Jiang

Pan

et al. 2015

Appl Environ Microbiol

Self Cite

View full text Add to dashboard Cite

show abstract

“…Interestingly, a cis-active element of ∼16 bp, designed dre (DasR responsive elements), is present downstream of the core promoter. The dre element plays a role in gene expression as deletion of this sequence resulted in constitutive expression of the chitinase gene (Xie et al, 2012). The dre sequence has also been reported in other bacteria, including Streptomyces and other Bacillus species (Colson et al, 2007;Bertram et al, 2011).…”

Section: Phylogenetic Analysis and Chitinase Expressionmentioning

confidence: 90%

“…Deletion of the CcpA regulator increases chitinase gene expression and enzyme activity by ∼39-fold (Jiang et al, 2015). Based on previously reports (Deutscher et al, 2006;Xie et al, 2012;Jiang et al, 2015;Cao et al, 2018), we have depicted a model for the regulation mediated by cre/CcpA and dre/NagR B in Figure 4.…”

Section: Phylogenetic Analysis and Chitinase Expressionmentioning

confidence: 93%

Chitinases of Bacillus thuringiensis: Phylogeny, Modular Structure, and Applied Potentials

et al. 2020

View full text Add to dashboard Cite

The most important bioinsecticide used worldwide is Bacillus thuringiensis and its hallmark is a rich variety of insecticidal Cry protein, many of which have been genetically engineered for expression in transgenic crops. Over the past 20 years, the discovery of other insecticidal proteins and metabolites synthesized by B. thuringiensis, including chitinases, antimicrobial peptides, vegetative insecticidal proteins (VIP), and siderophores, has expanded the applied value of this bacterium for use as an antibacterial, fungicidal, and nematicidal resource. These properties allow us to view B. thuringiensis not only as an entity for the production of a particular metabolite, but also as a multifaceted microbial factory. In particular, chitinases of B. thuringiensis are secreted enzymes that hydrolyze chitin, an abundant molecule in the biosphere, second only to cellulose. The observation that chitinases increase the insecticidal activity of Cry proteins has stimulated further study of these enzymes produced by B. thuringiensis. Here, we provide a review of a subset of our knowledge of B. thuringiensis chitinases as it relates to their phylogenetic relationships, regulation of expression, biotechnological potential for controlling entomopathogens, fungi, and nematodes, and their use in generating chitin-derived oligosaccharides (ChOGs) that possess antibacterial activities against a number of clinically significant bacterial pathogens. Recent advances in the structural organization of these enzymes are also discussed, as are our perspective for future studies.

show abstract

“…Promoter/ dre position swapping experiments were carried out using a promoter-probe vector pCB containing the reporter gene β-galactosidase ( bgaB ) (Xie et al, 2012 ). This vector was used to determine the effect of the changes in the relative position between the promoter and the dre site on the expression of the reporter gene.…”

Section: Methodsmentioning

confidence: 99%

NagRBt Is a Pleiotropic and Dual Transcriptional Regulator in Bacillus thuringiensis

et al. 2018

Self Cite

View full text Add to dashboard Cite

NagR, belonging to the GntR/HutC family, is a negative regulator that directly represses the nagP and nagAB genes, which are involved in GlcNAc transport and utilization in Bacillus subtilis. Our previous work confirmed that the chitinase B gene (chiB) of Bacillus thuringiensis strain Bti75 is also negatively controlled by YvoABt, the ortholog of NagR from B. subtilis. In this work, we investigated its regulatory network in Bti75 and found that YvoABt is an N-acetylglucosamine utilization regulator primarily involved in GlcNAc catabolism; therefore YvoABt is renamed as NagRBt. The RNA-seq data revealed that 27 genes were upregulated and 14 genes were downregulated in the ΔnagR mutant compared with the wild-type strain. The regulon (exponential phase) was characterized by RNA-seq, bioinformatics software, electrophoretic mobility shift assays, and quantitative real-time reverse transcription PCR. In the Bti75 genome, 19 genes that were directly regulated and 30 genes that were indirectly regulated by NagRBt were identified. We compiled in silico, in vitro, and in vivo evidence that NagRBt behaves as a repressor and activator to directly or indirectly influence major biological processes involved in amino sugar metabolism, nucleotide metabolism, fatty acid metabolism, phosphotransferase system, and the Embden–Meyerhof–Parnas pathway.

show abstract

Construction of a Promoter-probe Vector for Bacillus thuringiensis: the Identification of cis-acting Elements of the chiA Locus

Cited by 11 publications

References 34 publications

YvoA and CcpA Repress the Expression of chiB in Bacillus thuringiensis

YvoA and CcpA Repress the Expression of chiB in Bacillus thuringiensis

Chitinases of Bacillus thuringiensis: Phylogeny, Modular Structure, and Applied Potentials

NagRBt Is a Pleiotropic and Dual Transcriptional Regulator in Bacillus thuringiensis

Contact Info

Product

Resources

About