Bt Research 2025, Vol.16, No.5, 194-203 http://microbescipublisher.com/index.php/bt 195 2 Basic Framework of Bt Gene Regulation Network 2.1 Recognition of transcription factors and regulatory elements In the Bt gene regulation network, transcription factors are the core components, which determine when and where a specific gene is expressed. Studies have shown that Bt shares many global regulatory factors with relative species such as Bacillus cereus, which initiate transcription of downstream genes in stages during the bud formation stage. Among them, Spo0A is the main regulatory protein that initiates spore formation. Studies have found that activation of Spo0A not only triggers spore-related gene expression (Hou et al., 2022), but also inhibits the early transcription of certain toxin genes to ensure the reasonable allocation of metabolic resources. In addition, the unique regulatory proteins in Bt have been continuously identified. In recent years, through comparative genomic analysis and high-throughput DNA binding site screening, researchers have identified multiple new Bt transcription factors and their binding elements. The promoter region of the Bt rocE gene (encoding an arginine transporter) was analyzed, and the transcription unit of its operon was determined, and the β-galactosidase reporter system and electrophoretic mobility analysis were used to reveal the regulatory effect of a GntR family transcription factor on rocE transcription (Lereclus et al., 2000). 2.2 The role of non-coding RNA in regulation In addition to traditional protein transcription factors, non-coding regulatory elements such as small RNA (sRNA) are also present in the Bt gene regulatory network, which fine-tune gene expression by affecting mRNA stability or translation efficiency. Researchers have identified a variety of sRNA molecules in Bt and its relatives. For example, Yu Zhaoqing et al. (2022) from Central Agricultural University found through transcriptomic comparative analysis that there are three sets of hfq genes (coding for RNA chaperone protein Hfq) on the genome of the Bt BMB171 strain. After knocking out all three sets of hfq, the global transcription of Bt changed significantly, with differences in the transcription levels of about 32.5% of genes, with many gene expressions related to bud formation and virulence being significantly altered (Yu et al., 2022b). This suggests that Hfq-mediated sRNA regulation plays a key role in Bt (Figure 1). Hfq is an RNA chaperone protein that binds sRNA and target mRNA and promotes complementary pairing between the two, thereby affecting the expression of target genes (Peng et al., 2017). In Bt, Hfq may coordinate the regulation of phenotypes such as movement, biofilm, spore formation and toxin synthesis by stabilizing certain stress-related sRNAs or blocking the translation of specific mRNAs. 2.3 Epigenetic modification and gene expression regulation In addition to the DNA sequence itself, epigenetic markers such as DNA methylation will also affect the expression pattern of the Bt gene. Some studies have pointed out that bacterial DNA methylation is involved in the regulation of environmental responses and biofilm formation. For Bt, its genome carries multiple methylases encoded by type I and type II restriction-modification systems. It is speculated that these enzymes may also play a role in gene expression regulation in addition to defending against phages (Dordet-Frisoni et al., 2021). In addition, nucleosome-like proteins resemble eukaryotic histones in Bacillus, and may also affect transcriptional efficiency by changing DNA conformation. Population-level genomic analysis found that in the evolution of Bacillus cereal complexes, the CRISPR-Cas system genes were selectively inactivated, allowing strains to more easily obtain plasmids and virulence genes through horizontal gene transfer. Although this discovery focuses on population evolution, it also reveals that in order to adapt to diverse environments, Bt strains can strengthen their own genetic diversity and regulatory capabilities by "turning off" certain defense systems without changing the base sequence encoding information (Jogam et al., 2022). 3 Application of Omics Technology in Bt Research 3.1 Genomics: genome sequencing and comparison of Bt strains Since the whole genome sequencing of the first Bt strain was completed, thousands of Bt and their relative strains have been sequenced worldwide. It is reported that as of 2024, more than 10 000 Bacillus genomic sequences have been published in the NCBI database, of which Bt accounts for a considerable proportion, and more than 1 000 genomes have been fully assembled and annotated. Genomic comparative analysis shows that Bt has a set of
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