H-NS is a Transcriptional Repressor of the CRISPR-Cas System in Acinetobacter baumannii ATCC 19606

• Published in: The journal of microbiology 2024, 62(11), , pp.999-1012
• Main Authors: Kim, Kyeongmin, Islam, Md. Maidul, Bang, Seunghyeok, Kim, Jeongah, Lee, Chung-Young, Lee, Je Chul, Shin, Minsang
• Format: Article
• Language: English
• Subjects: Acinetobacter baumannii; Acinetobacter baumannii - genetics; Acinetobacter baumannii - metabolism; Affinity chromatography; Amino acid sequence; Assaying; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Biomedical and Life Sciences; Chromatography; CRISPR; CRISPR-Associated Proteins - genetics; CRISPR-Associated Proteins - metabolism; CRISPR-Cas Systems; Crosslinking; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Galactosidase; Gene Expression Regulation, Bacterial; Gene regulation; Gene transfer; Genes; Genetic transformation; Genomics and Molecular Biology (Prokaryote); Horizontal transfer; Integration host factor; Life Sciences; Microbial Genetics; Microbiology; Molecular modelling; Multidrug resistance; Mutation; Nosocomial infection; Nucleotide sequence; Opportunist infection; Promoter Regions, Genetic; Promoters; Repressor Proteins - genetics; Repressor Proteins - metabolism; Sequence analysis; Size exclusion chromatography; Stationary phase; Strain analysis; Transcription factors; β-Galactosidase; 생물학
• ISSN: 1225-8873; 1976-3794; 1976-3794
• DOI: 10.1007/s12275-024-00182-5

Acinetobacter baumannii is a multidrug-resistant opportunistic pathogen primarily associated with hospital-acquired infections. The bacterium can gain multidrug resistance through several mechanisms, including horizontal gene transfer. A CRISPR-Cas system including several Cas genes could restrict the horizontal gene transfer. However, the molecular mechanism of CRISPR- Cas transcriptional regulation remains unclear. We identified a type I-F CRISPR-Cas system in A. baumannii ATCC 19606 T standard strain based on sequence analysis. We focused on the transcriptional regulation of Cas3, a key protein of the CRISPR-Cas system. We performed a DNA affinity chromatography-pulldown assay to identify transcriptional regulators of the Cas3 promoter. We identified several putative transcriptional factors, such as H-NS, integration host factor, and HU, that can bind to the promoter region of Cas3 . We characterized Ab H-NS using size exclusion chromatography and cross-linking experiments and demonstrated that the Cas3 promoter can be regulated by Ab H-NS in a concentration-dependent manner via an in vitro transcription assay. CRISPR-Cas expression levels in wild-type and hns mutant strains in the early stationary phase were examined by qPCR and β-galactosidase assay. We found that H-NS can act as a repressor of Cas3. Our transformation efficiency results indicated that the hns mutation decreased the transformation efficiency, while the Cas3 mutation increased it. We report the existence and characterization of the CRISPR-Cas system in A. baumannii 19606 T and demonstrate that Ab H-NS is a transcriptional repressor of CRISPR-Cas -related genes in A. baumannii .