Essential Role for the Response Regulator PmrA in Coxiella burnetii Type 4B Secretion and Colonization of Mammalian Host Cells
Essential Role for the Response Regulator PmrA in Coxiella burnetii Type 4B Secretion and Colonization of Mammalian Host Cells
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DOI：
10.1128/jb.01532-14
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发表时间：
2014-06-01
影响因子：
3.2
通讯作者：
Heinzen, Robert A.
中科院分区：
文献类型：
--
作者：
Beare, Paul A.;Sandoz, Kelsi M.;Heinzen, Robert A.
Q热病病原体伯氏柯克斯体成功的宿主细胞定植需要通过Dot/ICM4B型分泌系统(T4BSS)将效应蛋白转位到宿主胞浆中。在嗜肺军团菌中，双组分系统(TCS)PmrAB调控Dot/ICM T4BSS和其他几个与发病相关的生理过程。由于PmrA共识调控元件与一些Dot/ICM和底物基因相关，因此PmrA在调节伯氏梭菌T4BSS中具有类似的作用。在这里，我们构建了一个伯氏梭菌PmrA缺失突变体，以直接检测PmrA介导的基因调控。与野生型细菌相比，Burnetii Delta pmrA表现出严重的细胞内生长缺陷，这与效应蛋白的分泌失败相吻合。荧光素酶基因报告分析显示，7个Dot/ICM操纵子中有5个依赖于PmrA的表达，11个效应编码基因中有9个具有预测的上游PmrA调控元件。突变分析验证了PmrA定向转录所需的一致序列核苷酸。野生型伯氏卡氏杆菌和Delta pmrA突变体的RNA测序和全细菌质谱分析发现了PmrA调节子的新成分，包括几个编码Dot/ICM底物的PmrA基序缺失的基因。综上所述，我们的结果表明，PmrAB TCS是调节伯氏梭菌Dot/ICM分泌的关键毒力因子。缺乏PmrA调控元件的PmrA反应基因的存在也表明PmrAB TCS控制与参与宿主细胞寄生的额外伯氏杆菌蛋白的产生相关的调控系统的表达。
Successful host cell colonization by the Q fever pathogen, Coxiella burnetii, requires translocation of effector proteins into the host cytosol by a Dot/Icm type 4B secretion system (T4BSS). In Legionella pneumophila, the two-component system ( TCS) PmrAB regulates the Dot/Icm T4BSS and several additional physiological processes associated with pathogenesis. Because PmrA consensus regulatory elements are associated with some dot/icm and substrate genes, a similar role for PmrA in regulation of the C. burnetii T4BSS has been proposed. Here, we constructed a C. burnetii pmrA deletion mutant to directly probe PmrA-mediated gene regulation. Compared to wild-type bacteria, C. burnetii Delta pmrA exhibited severe intracellular growth defects that coincided with failed secretion of effector proteins. Luciferase gene reporter assays demonstrated PmrA-dependent expression of 5 of 7 dot/icm operons and 9 of 11 effector-encoding genes with a predicted upstream PmrA regulatory element. Mutational analysis verified consensus sequence nucleotides required for PmrA-directed transcription. RNA sequencing and whole bacterial cell mass spectrometry of wild-type C. burnetii and the Delta pmrA mutant uncovered new components of the PmrA regulon, including several genes lacking PmrA motifs that encoded Dot/Icm substrates. Collectively, our results indicate that the PmrAB TCS is a critical virulence factor that regulates C. burnetii Dot/Icm secretion. The presence of PmrA-responsive genes lacking PmrA regulatory elements also suggests that the PmrAB TCS controls expression of regulatory systems associated with the production of additional C. burnetii proteins involved in host cell parasitism.