Klebsiella pneumoniae type VI secretion system-mediated microbial competition is PhoPQ controlled and reactive oxygen species dependent
Klebsiella pneumoniae type VI secretion system-mediated microbial competition is PhoPQ controlled and reactive oxygen species dependent
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DOI：
10.1101/698415
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发表时间：
2019-07
期刊：
影响因子：
6.7
通讯作者：
Danielle Storey;A. McNally;M. Åstrand;Joana Sá-Pessoa Graca Santos;I. Rodríguez-Escudero;Bronagh Elmore;Leyre Palacios;H. Marshall;L. Hobley;M. Martin;V. J. Cid;T. Salminen;J. Bengoechea
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作者：
Danielle Storey;A. McNally;M. Åstrand;Joana Sá-Pessoa Graca Santos;I. Rodríguez-Escudero;Bronagh Elmore;Leyre Palacios;H. Marshall;L. Hobley;M. Martin;V. J. Cid;T. Salminen;J. Bengoechea
由于多重耐药菌株的日益分离，肺炎克雷伯菌被认为是对人类健康的紧迫威胁。高毒力菌株是一个主要问题，因为它们能够在健康宿主中引起危及生命的感染。 VI 型分泌系统（T6SS）广泛参与微生物拮抗作用，并且在某些情况下介导与宿主真核细胞的相互作用。在 700 个肺炎克雷伯菌基因组中对与 T6SS 组成基因和 T6SS 效应基因同源的基因进行计算机搜索，结果显示肺炎克雷伯菌物种中 T6SS 基因具有广泛的多样性。温度、氧张力、pH、渗透压、铁水平和 NaCl 调节高毒力肺炎克雷伯菌菌株编码的 T6SS 的表达。多粘菌素和人防御素 3 也会增加 T6SS 的活性。对 T6SS 调控因子的筛选揭示了 T6SS 转录与杀死大肠杆菌猎物的能力之间的相关性。 H-NS 抑制 T6SS，而 PhoPQ、PmrAB、Hfq、Fur、RpoS 和 RpoN 正向调节 T6SS。肺炎克雷伯菌 T6SS 介导种内和种间细菌竞争。这种对抗只有当猎物拥有活跃的 T6SS 时才会明显。 PhoPQ 二元系统控制肺炎克雷伯菌 T6SS 在细菌竞争中的激活。从机制上讲，PhoQ 周质结构域和其中的酸性斑块对于激活肺炎克雷伯菌 T6SS 至关重要。克雷伯菌 T6SS 还介导抗真菌竞争。我们描述了每个 VgrG 在微生物竞争中的贡献，并将 VgrG4 确定为 T6SS 效应子。从结构上讲，VgrG4 的结构域 DUF2345 足以使细菌和酵母中毒。 ROS 的产生介导 VgrG4 的抗菌作用，抗毒素 Sel1E 可以防止 VgrG4 的毒性活性。我们的研究结果使人们更好地了解 T6SS 在细菌竞争中的调节，并将 ROS 视为微生物竞争的早期事件。作者摘要 肺炎克雷伯菌由于具有极高的耐药性菌株而被列为“对人类健康的紧急威胁”。许多研究调查了肺炎克雷伯菌抗生素耐药性的分子机制，而其他研究则剖析了这种病原体的毒力策略。然而，对于克雷伯氏菌在环境中的适应性，特别是克雷伯氏菌与其他物种的竞争，人们的了解仍然有限。在这里，我们证明了克雷伯氏菌利用 VI 型分泌系统 (T6SS) 纳米武器来杀死细菌竞争对手和真菌。肺炎克雷伯菌感知到细菌竞争对手的 T6SS 攻击，从而对攻击性细胞进行报复。对攻击的感知涉及传感器 PhoPQ 并导致 T6SS 上调。我们鉴定了 T6SS 用来对抗其他微生物的一种毒素，并揭示了克雷伯氏菌如何保护自己免受这种毒素的侵害。我们的研究结果使人们更好地了解 T6SS 在微生物竞争中的作用，并揭示了细菌如何调节 T6SS 介导的微生物拮抗作用的新方面。
Klebsiella pneumoniae is recognized as an urgent threat to human health due to the increasing isolation of multidrug resistant strains. Hypervirulent strains are a major concern due to their ability to cause life-threating infections in healthy hosts. The type VI secretion system (T6SS) is widely implicated in microbial antagonism, and it mediates interactions with host eukaryotic cells in some cases. In silico search for genes orthologous to T6SS component genes and T6SS effector genes across 700 K. pneumoniae genomes shows extensive diversity in T6SS genes across the K. pneumoniae species. Temperature, oxygen tension, pH, osmolarity, iron levels, and NaCl regulate the expression of the T6SS encoded by a hypervirulent K. pneumoniae strain. Polymyxins and human defensin 3 also increase the activity of the T6SS. A screen for regulators governing T6SS uncover the correlation between the transcription of the T6SS and the ability to kill E. coli prey. Whereas H-NS represses the T6SS, PhoPQ, PmrAB, Hfq, Fur, RpoS and RpoN positively regulate the T6SS. K. pneumoniae T6SS mediates intra and inter species bacterial competition. This antagonism is only evident when the prey possess an active T6SS. The PhoPQ two component system governs the activation of K. pneumoniae T6SS in bacterial competitions. Mechanistically, PhoQ periplasmic domain, and the acid patch within, is essential to activate K. pneumoniae T6SS. Klebsiella T6SS also mediates anti-fungal competition. We have delineated the contribution of each of the individual VgrGs in microbial competition, and identified VgrG4 as a T6SS effector. Structurally, domain DUF2345 of VgrG4 is sufficient to intoxicate bacteria and yeast. ROS generation mediates the antibacterial effects of VgrG4, and the antitoxin Sel1E protects against the toxic activity of VgrG4. Our findings provide a better understanding of the regulation of the T6SS in bacterial competitions, and place ROS as an early event in microbial competition. AUTHOR SUMMARY Klebsiella pneumoniae has been singled out as an “urgent threat to human health” due to extremely drug resistant strains. Numerous studies investigate the molecular mechanisms underlying antibiotic resistance in K. pneumoniae, while others dissect the virulence strategies of this pathogen. However, there is still limited knowledge on the fitness of Klebsiella in the environment, and particularly the competition of Klebsiella with other species. Here, we demonstrated that Klebsiella exploits the type VI secretion system (T6SS) nanoweapon to kill bacterial competitors and fungi. K. pneumoniae perceives T6SS attacks from bacterial competitors, resulting in retaliation against the aggressive cell. The perception of the attack involved the sensor PhoPQ and led to the up-regulation of the T6SS. We identified one of the toxins deployed by the T6SS to antagonize other microbes, and revealed how Klebsiella protects itself from this toxin. Our findings provide a better understanding of the T6SS role in microbial competition and uncover new aspects on how bacteria regulate T6SS-mediated microbial antagonism.