食源性病原体鼠伤寒沙门氏菌亚种血清型鼠伤寒沙门氏菌实验性启动子的近单碱基对分辨率鉴定。

Experimental promoter identification of a foodborne pathogen subsp. serovar Typhimurium with near single base-pair resolution.

作者信息

Lee Sang-Mok, Le Hoa Thi, Taizhanova Assiya, Nong Linh Khanh, Park Joon Young, Lee Eun-Jin, Palsson Bernhard O, Kim Donghyuk

机构信息

School of Energy and Chemical Engineering, Ulsan National Institute of Science and Technology (UNIST), Ulsan, Republic of Korea.

Department of Genetic Engineering and Graduate School of Biotechnology, College of Life Sciences, Kyung Hee University, Yongin, Republic of Korea.

出版信息

Front Microbiol. 2024 Jan 4;14:1271121. doi: 10.3389/fmicb.2023.1271121. eCollection 2023.

Abstract

serovar Typhimurium ( Typhimurium) is a common foodborne pathogen which is frequently used as the reference strain for . Investigating the sigma factor network and protomers is crucial to understand the genomic and transcriptomic properties of the bacterium. Its promoters were identified using various methods such as dRNA-seq, ChIP-chip, or ChIP-Seq. However, validation using ChIP-exo, which exhibits higher-resolution performance compared to conventional ChIP, has not been conducted to date. In this study, using the representative strain Typhimurium LT2 (LT2), the ChIP-exo experiment was conducted to accurately determine the binding sites of catalytic RNA polymerase subunit RpoB and major sigma factors (RpoD, RpoN, RpoS, and RpoE) during exponential phase. Integrated with the results of RNA-Seq, promoters and sigmulons for the sigma factors and their association with RpoB have been discovered. Notably, the overlapping regions among binding sites of each alternative sigma factor were found. Furthermore, comparative analysis with str. K-12 substr. MG1655 (MG1655) revealed conserved binding sites of RpoD and RpoN across different species. In the case of small RNAs (sRNAs), 50 sRNAs observed their expression during the exponential growth of LT2. Collectively, the integration of ChIP-exo and RNA-Seq enables genome-scale promoter mapping with high resolution and facilitates the characterization of binding events of alternative sigma factors, enabling a comprehensive understanding of the bacterial sigma factor network and condition-specific active promoters.

摘要

鼠伤寒血清型沙门氏菌(鼠伤寒沙门氏菌)是一种常见的食源性病原体,常被用作……的参考菌株。研究西格玛因子网络和启动子对于理解该细菌的基因组和转录组特性至关重要。其启动子通过多种方法鉴定,如dRNA-seq、ChIP-chip或ChIP-Seq。然而,与传统ChIP相比具有更高分辨率性能的ChIP-exo验证迄今尚未进行。在本研究中,使用代表性菌株鼠伤寒沙门氏菌LT2(LT2)进行ChIP-exo实验,以准确确定指数生长期催化RNA聚合酶亚基RpoB和主要西格玛因子(RpoD、RpoN、RpoS和RpoE)的结合位点。结合RNA-Seq的结果,发现了西格玛因子的启动子和西格玛操纵子及其与RpoB的关联。值得注意的是,发现了每个替代西格玛因子结合位点之间的重叠区域。此外,与大肠杆菌K-12菌株MG1655(MG1655)的比较分析揭示了不同物种间RpoD和RpoN的保守结合位点。在小RNA(sRNA)方面,50种sRNA在LT2指数生长期间观察到其表达。总体而言,ChIP-exo和RNA-Seq的整合能够实现高分辨率的全基因组规模启动子图谱绘制,并有助于表征替代西格玛因子的结合事件,从而全面了解细菌西格玛因子网络和条件特异性活性启动子。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/cea8/10794520/5d6e33886da6/fmicb-14-1271121-g001.jpg

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