RNA-based thermoregulation of a Campylobacter jejuni zinc resistance determinant.
RNA-based thermoregulation of a Campylobacter jejuni zinc resistance determinant.
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DOI：
10.1371/journal.ppat.1009008
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发表时间：
2020-10
期刊：
影响因子：
6.7
通讯作者：
Linton D
中科院分区：
文献类型：
--
作者：
Barnawi H;Masri N;Hussain N;Al-Lawati B;Mayasari E;Gulbicka A;Jervis AJ;Huang MH;Cavet JS;Linton D
RNA温度计（RNAT）触发细菌毒力因子的表达，以响应进入温血宿主的温度变化。在较低的温度下，这些二级结构隔离核糖体结合位点（RBS），以防止翻译起始，而在升高的温度下，它们“融化”，允许翻译。空肠弯曲菌是引起人类胃肠炎的主要致病菌，但其与宿主的相互作用机制，包括宿主诱导的基因调控机制尚不清楚。在这里，我们证明了RNAT调节C。空肠基因，Cj 1163 c或czcD，编码阳离子扩散促进剂家族的成员。czcD上游非翻译区在mRNA内含有预测的茎环，其在低于37°C的温度下螯合RBS以抑制翻译。破坏或增强预测的二级结构的突变对温度调节具有显著和可预测的影响。我们还表明，在一个RNAT独立的方式，CzcD的表达是由锌（II）诱导。缺乏czcD的突变体对Zn（II）过敏，并且相对于野生型也过度积累Zn（II），所有这些都与CzcD作为Zn（II）输出者的功能一致。重要的是，我们证明了C。空肠在32°C（RNAT限制CzcD产生的温度）下的Zn（II）耐受性通过RNAT破坏而增加。最后，我们表明czcD失活减弱了Galleria感染模型中的幼虫杀伤，并且在32°C下破坏RNAT二级结构以允许CzcD产生可以增强杀伤。我们假设金属和温度对CzcD的调节为C.空肠克服先天免疫系统介导的Zn（II）毒性在温血动物宿主。通过所谓的RNA温度计中的结构变化来感测升高的温血宿主体温是细菌病原体中公认的毒力策略，并且允许在宿主环境中适当地产生毒力因子。我们证明，这种调节机制是由空肠弯曲杆菌，一个全球性的人类胃肠炎的重要原因，以控制生产的CzcD蛋白，我们显示作为一个锌（II）出口商提供耐受性高水平的锌（II）。从已发表的数据，锌（II）出口商是已知的，使细菌病原体克服主机诱导的锌（II）中毒，一个新发现的先天免疫反应感染。事实上，使用简单的幼虫感染模型，我们证明了CzcD增强了幼虫杀伤，并且破坏RNA温度计以允许CzcD产生也增强了在较低温度下的杀伤。这些发现表明CzcD温度调节使C.空肠克服宿主诱导的Zn（II）毒性在动物宿主包括人的定殖期间。我们的研究结果开辟了一个新的领域的宿主-病原体界面的调查，在这个鲜为人知的人类病原体。也有可能利用这些发现来开发基于金属的干预策略，以减少食物链中弯曲杆菌的水平和随之而来的人畜共患感染。
RNA thermometers (RNATs) trigger bacterial virulence factor expression in response to the temperature shift on entering a warm-blooded host. At lower temperatures these secondary structures sequester ribosome-binding sites (RBSs) to prevent translation initiation, whereas at elevated temperatures they “melt” allowing translation. Campylobacter jejuni is the leading bacterial cause of human gastroenteritis worldwide yet little is known about how it interacts with the host including host induced gene regulation. Here we demonstrate that an RNAT regulates a C. jejuni gene, Cj1163c or czcD, encoding a member of the Cation Diffusion Facilitator family. The czcD upstream untranslated region contains a predicted stem loop within the mRNA that sequesters the RBS to inhibit translation at temperatures below 37°C. Mutations that disrupt or enhance predicted secondary structure have significant and predictable effects on temperature regulation. We also show that in an RNAT independent manner, CzcD expression is induced by Zn(II). Mutants lacking czcD are hypersensitive to Zn(II) and also over-accumulate Zn(II) relative to wild-type, all consistent with CzcD functioning as a Zn(II) exporter. Importantly, we demonstrate that C. jejuni Zn(II)-tolerance at 32°C, a temperature at which the RNAT limits CzcD production, is increased by RNAT disruption. Finally we show that czcD inactivation attenuates larval killing in a Galleria infection model and that at 32°C disrupting RNAT secondary structure to allow CzcD production can enhance killing. We hypothesise that CzcD regulation by metals and temperature provides a mechanism for C. jejuni to overcome innate immune system-mediated Zn(II) toxicity in warm-blooded animal hosts. Sensing the elevated warm-blooded host body temperature through structural changes in so-called RNA thermometers is a recognised virulence strategy in bacterial pathogens and allows appropriate production of virulence factors in the host environment. We demonstrated that this regulatory mechanism is used by Campylobacter jejuni, a globally significant cause of human gastroenteritis, to control production of the CzcD protein that we show acts as a Zn(II) exporter providing tolerance to high levels of Zn(II). From published data, Zn(II) exporters are known to enable bacterial pathogens to overcome host-induced Zn(II) poisoning—a newly discovered innate immune response to infection. Indeed using a simple larval infection model we demonstrated that CzcD enhances larval killing and disrupting the RNA thermometer to allow CzcD production also enhanced killing at lower temperatures. These findings indicate that CzcD temperature regulation enables C. jejuni to overcome host induced Zn(II) toxicity during colonisation of animal hosts including humans. Our results open up a novel area of the host-pathogen interface for investigation in this poorly understood human pathogen. It may also be possible to exploit these findings to develop metal-based intervention strategies to reduce both levels of Campylobacter in the food chain and consequent zoonotic infections.
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