Guangdong Laboratory for Lingnan Modern Agriculture, South China Agricultural Universitygrid.20561.30, Guangzhou, China.
Guangdong Province Key Laboratory of Microbial Signals and Disease Control, Integrative Microbiology Research Center, South China Agricultural Universitygrid.20561.30, Guangzhou, China.
Appl Environ Microbiol. 2022 Jan 25;88(2):e0165521. doi: 10.1128/AEM.01655-21. Epub 2021 Nov 3.
The type VI secretion system (T6SS) is an important translocation apparatus that is widely employed by Gram-negative bacteria to deliver toxic effectors into eukaryotic and prokaryotic target cells, causing host damage and providing competitive advantages in polymicrobial environments. The genome of Pseudomonas aeruginosa harbors three T6SS clusters (H1-T6SS, H2-T6SS, H3-T6SS). Activities of these systems are tightly regulated by a complicated signaling network which remains largely elusive. In this study, we focused on a previously characterized two-component system FleS/FleR, and performed comparative transcriptome analysis between the PAO1 wild-type strain and its isogenic Δ mutant, which revealed the important role of FleS/FleR in regulating multiple physiological pathways including T6SS. Gene expression and bacterial killing assays showed that the expression and activity of H1-T6SS are repressed in the wild-type strain owing to the high intracellular c-di-GMP content. Further explorations demonstrated that c-di-GMP relies on the transcription factor FleQ to repress H1-T6SS and its synthesis is controlled by a global regulator AmrZ which is induced by the active FleS/FleR. Interestingly, repression of H1-T6SS by FleS/FleR in PAO1 is independent of RetS which is known to regulate H1-T6SS by controlling the central post-transcriptional factor RsmA. Together, our results identified a novel regulator of H1-T6SS and provided detailed mechanisms of this signaling pathway in PAO1. Pseudomonas aeruginosa is an opportunistic human pathogen distributed widely in the environment. The genome of this pathogen contains three T6SS clusters which contribute significantly to its virulence. Understanding the complex regulatory network that controls the activity of T6SS is essential for the development of effective therapeutic treatments for P. aeruginosa infections. In this study, transcriptome analysis led to the identification of a novel regulator FleS/FleR which inversely regulates H1-T6SS and H2-T6SS in P. aeruginosa PAO1. We further revealed a detailed FleS/FleR-mediated regulatory pathway of H1-T6SS in PAO1 which involves two additional transcriptional regulators AmrZ and FleQ and the second messenger c-di-GMP, providing important implications to develop novel anti-infective strategies and antimicrobial drugs.
VI 型分泌系统（T6SS）是一种重要的转运装置，广泛存在于革兰氏阴性菌中，用于将毒性效应器输送到真核和原核靶细胞中，导致宿主损伤，并在多微生物环境中提供竞争优势。铜绿假单胞菌的基因组包含三个 T6SS 簇（H1-T6SS、H2-T6SS、H3-T6SS）。这些系统的活性受到一个复杂的信号网络的严格调控，但这个信号网络在很大程度上仍然难以捉摸。在这项研究中，我们专注于一个以前被表征的双组分系统 FleS/FleR，并在 PAO1 野生型菌株及其同源缺失突变体之间进行了比较转录组分析，结果表明 FleS/FleR 在调节包括 T6SS 在内的多种生理途径中起着重要作用。基因表达和细菌杀伤实验表明，由于细胞内 c-di-GMP 含量高，野生型菌株中 H1-T6SS 的表达和活性受到抑制。进一步的探索表明，c-di-GMP 依赖转录因子 FleQ 来抑制 H1-T6SS，其合成受到全局调控因子 AmrZ 的控制，AmrZ 被活性 FleS/FleR 诱导。有趣的是，FleS/FleR 在 PAO1 中对 H1-T6SS 的抑制作用不依赖于 RetS，RetS 已知通过控制中央转录后因子 RsmA 来调节 H1-T6SS。总之，我们的研究结果确定了 H1-T6SS 的一个新调控因子，并提供了 PAO1 中该信号通路的详细机制。铜绿假单胞菌是一种机会性人类病原体，广泛分布于环境中。该病原体的基因组包含三个 T6SS 簇，对其毒力有重要贡献。了解控制 T6SS 活性的复杂调控网络对于开发治疗铜绿假单胞菌感染的有效治疗方法至关重要。在这项研究中，转录组分析导致鉴定出一个新的调控因子 FleS/FleR，它在 PAO1 中反向调节 H1-T6SS 和 H2-T6SS。我们进一步揭示了 PAO1 中 FleS/FleR 介导的 H1-T6SS 详细调控途径，该途径涉及两个额外的转录调控因子 AmrZ 和 FleQ 以及第二信使 c-di-GMP，为开发新的抗感染策略和抗菌药物提供了重要启示。
