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

IF 3.3 4区 生物学 Q2 MICROBIOLOGY
Journal of Microbiology Pub Date : 2024-11-01 Epub Date: 2024-11-11 DOI:10.1007/s12275-024-00182-5
Kyeongmin Kim, Md Maidul Islam, Seunghyeok Bang, Jeongah Kim, Chung-Young Lee, Je Chul Lee, Minsang Shin
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引用次数: 0

Abstract

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 19606T 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 AbH-NS using size exclusion chromatography and cross-linking experiments and demonstrated that the Cas3 promoter can be regulated by AbH-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 19606T and demonstrate that AbH-NS is a transcriptional repressor of CRISPR-Cas-related genes in A. baumannii.

H-NS 是鲍曼不动杆菌 ATCC 19606 中 CRISPR-Cas 系统的转录抑制因子。
鲍曼不动杆菌是一种耐多药的机会性病原体,主要与医院获得性感染有关。该细菌可通过多种机制获得多重耐药性,包括水平基因转移。包括多个 Cas 基因的 CRISPR-Cas 系统可以限制水平基因转移。然而,CRISPR-Cas转录调控的分子机制仍不清楚。根据序列分析,我们在鲍曼不动杆菌 ATCC 19606T 标准菌株中发现了 I-F 型 CRISPR-Cas 系统。我们重点研究了CRISPR-Cas系统的关键蛋白Cas3的转录调控。我们采用 DNA 亲和层析-下拉实验来鉴定 Cas3 启动子的转录调控因子。我们发现了几种推定的转录因子,如H-NS、整合宿主因子和HU,它们可以与Cas3的启动子区域结合。我们利用尺寸排阻色谱法和交联实验对 AbH-NS 进行了表征,并通过体外转录实验证明了 Cas3 启动子能以浓度依赖性的方式受到 AbH-NS 的调控。通过 qPCR 和 β-半乳糖苷酶检测法检测了野生型和 hns 突变株在静止早期的 CRISPR-Cas 表达水平。我们发现,H-NS 可作为 Cas3 的抑制因子。我们的转化效率结果表明,hns突变降低了转化效率,而Cas3突变则提高了转化效率。我们报告了鲍曼不动杆菌 19606T 中 CRISPR-Cas 系统的存在和特征,并证明 AbH-NS 是鲍曼不动杆菌中 CRISPR-Cas 相关基因的转录抑制因子。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
Journal of Microbiology
Journal of Microbiology 生物-微生物学
CiteScore
5.70
自引率
3.30%
发文量
0
审稿时长
3 months
期刊介绍: Publishes papers that deal with research on microorganisms, including archaea, bacteria, yeasts, fungi, microalgae, protozoa, and simple eukaryotic microorganisms. Topics considered for publication include Microbial Systematics, Evolutionary Microbiology, Microbial Ecology, Environmental Microbiology, Microbial Genetics, Genomics, Molecular Biology, Microbial Physiology, Biochemistry, Microbial Pathogenesis, Host-Microbe Interaction, Systems Microbiology, Synthetic Microbiology, Bioinformatics and Virology. Manuscripts dealing with simple identification of microorganism(s), cloning of a known gene and its expression in a microbial host, and clinical statistics will not be considered for publication by JM.
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