The BvgAS Regulon of Bordetella pertussis.
The BvgAS Regulon of Bordetella pertussis.
复制标题
DOI：
10.1128/mbio.01526-17
复制
发表时间：
2017-10-10
期刊：
影响因子：
6.4
通讯作者：
Hinton DM
中科院分区：
文献类型：
--
作者：
Moon K;Bonocora RP;Kim DD;Chen Q;Wade JT;Stibitz S;Hinton DM
几乎所有的百日咳博德特氏菌的毒力因子激活的主双组分系统，BvgAS，传感器激酶BvgS和反应调节BvgA组成。当BvgS有活性时，BvgA被磷酸化（BvgA~P），并表达毒力激活基因（vags）[Bvg（+）模式]。当BvgS失活且BvgA未被磷酸化时，诱导了毒性抑制基因（vrgs）[Bvg（-）模式]。在此，我们使用转录组测序（RNA-seq）和逆转录-定量PCR（RT-qPCR）来确定B. pertussis Tohama I的BvgAS依赖性调节子。我们的分析揭示了超过550个BvgA调控基因，其中353个是新发现的。BvgA激活的基因包括编码双组分系统（如kdpED）、多种其他转录调节因子和3型分泌系统（T3 SS）表达所需的细胞质外功能（ECF）σ因子brpL的那些基因，进一步确立了BvgA~P作为促进毒力的转录网络的顶端调节因子的重要性。使用体外转录，我们证明了brpL的启动子直接被BvgA~ P激活。BvgA-FeBABE切割反应鉴定了bprL中位于−41.5和−63.5位置的BvgA~P结合位点。最重要的是，我们第一次证明了在B. pertussis Bvg（−）模式中，多种不同代谢途径的基因显著上调。这些基因包括脂肪酸和脂质代谢、糖和氨基酸转运蛋白、丙酮酸脱氢酶、苯乙酸降解和乙醇酸/乙醛酸利用途径的基因。我们的研究结果表明，Bvg（-）模式的代谢变化可能参与细菌的存活，传播和/或持久性，并确定了200多个新的vrg可以测试功能。在过去20年中，尽管疫苗接种覆盖率很高，但由百日咳杆菌引起的百日咳爆发已导致呼吸道疾病和婴儿死亡。这至少部分是由于20世纪90年代引入了一种效果较差的无细胞疫苗。因此，了解B.百日咳生长和感染的分子基础是至关重要的。双组分系统BvgA（应答调节子）/BvgS（组氨酸激酶）是B.百日咳毒力基因的主要调节子。我们在这里报告了B. pertussis中BvgAS调节子的第一个RNA-seq分析，揭示了超过550个基因受BvgAS调节。我们发现，多个和不同的代谢途径的基因在Bvg（−）模式（缺乏BvgA磷酸化）中受到高度调节。我们的研究结果表明，Bvg（-）模式的代谢变化可能参与细菌的存活，传播和/或持久性。
Nearly all virulence factors in Bordetella pertussis are activated by a master two-component system, BvgAS, composed of the sensor kinase BvgS and the response regulator BvgA. When BvgS is active, BvgA is phosphorylated (BvgA~P), and virulence-activated genes (vags) are expressed [Bvg(+) mode]. When BvgS is inactive and BvgA is not phosphorylated, virulence-repressed genes (vrgs) are induced [Bvg(−) mode]. Here, we have used transcriptome sequencing (RNA-seq) and reverse transcription-quantitative PCR (RT-qPCR) to define the BvgAS-dependent regulon of B. pertussis Tohama I. Our analyses reveal more than 550 BvgA-regulated genes, of which 353 are newly identified. BvgA-activated genes include those encoding two-component systems (such as kdpED), multiple other transcriptional regulators, and the extracytoplasmic function (ECF) sigma factor brpL, which is needed for type 3 secretion system (T3SS) expression, further establishing the importance of BvgA~P as an apex regulator of transcriptional networks promoting virulence. Using in vitro transcription, we demonstrate that the promoter for brpL is directly activated by BvgA~P. BvgA-FeBABE cleavage reactions identify BvgA~P binding sites centered at positions −41.5 and −63.5 in bprL. Most importantly, we show for the first time that genes for multiple and varied metabolic pathways are significantly upregulated in the B. pertussis Bvg(−) mode. These include genes for fatty acid and lipid metabolism, sugar and amino acid transporters, pyruvate dehydrogenase, phenylacetic acid degradation, and the glycolate/glyoxylate utilization pathway. Our results suggest that metabolic changes in the Bvg(−) mode may be participating in bacterial survival, transmission, and/or persistence and identify over 200 new vrgs that can be tested for function. Within the past 20 years, outbreaks of whooping cough, caused by Bordetella pertussis, have led to respiratory disease and infant mortalities, despite good vaccination coverage. This is due, at least in part, to the introduction of a less effective acellular vaccine in the 1990s. It is crucial, then, to understand the molecular basis of B. pertussis growth and infection. The two-component system BvgA (response regulator)/BvgS (histidine kinase) is the master regulator of B. pertussis virulence genes. We report here the first RNA-seq analysis of the BvgAS regulon in B. pertussis, revealing that more than 550 genes are regulated by BvgAS. We show that genes for multiple and varied metabolic pathways are highly regulated in the Bvg(−) mode (absence of BvgA phosphorylation). Our results suggest that metabolic changes in the Bvg(−) mode may be participating in bacterial survival, transmission, and/or persistence.