Proteomic analysis of growth phase-dependent expression of Legionella pneumophila proteins which involves regulation of bacterial virulence traits.
Proteomic analysis of growth phase-dependent expression of Legionella pneumophila proteins which involves regulation of bacterial virulence traits.
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DOI：
10.1371/journal.pone.0011718
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发表时间：
2010-07-22
期刊：
影响因子：
3.7
通讯作者：
Miyake M
中科院分区：
文献类型：
--
作者：
Hayashi T;Nakamichi M;Naitou H;Ohashi N;Imai Y;Miyake M
嗜肺军团菌是一种引起军团病的病原体，它对生长条件的反应表现出毒力特征。特别是，已知在体外培养中，它在指数后阶段变得有毒。在这项研究中，我们使用二维荧光差示凝胶电泳(2D-DGE)和基质辅助激光解吸/电离-质谱仪(MALDI-TOF-MS)相结合的方法，对指数期和指数期后表达的差异进行了蛋白质组学分析，以确定与嗜肺乳杆菌毒力相关的候选基因。在两个生长阶段中发现的68个显著差异表达的蛋白质中，在指数后阶段上调。上调的蛋白质包括糖酵解、酮体生物合成和聚羟基丁酸酯(PHB)生物合成相关的酶，这表明当氨基酸变得稀缺时，嗜肺乳杆菌可能利用糖和脂肪作为能量来源。与运动相关的蛋白质(鞭毛成分和抽动运动相关蛋白)也上调，预示着它们在一定条件下增强了宿主细胞中细菌的感染力。此外，还发现了9个功能未知的上调蛋白。其中两个被鉴定为与绵羊红细胞(SRBC)溶血相关的新细菌因子。另有2个通过ICM/Dot IV型分泌器转运到巨噬细胞中，作为百日咳杆菌腺苷环化酶报告试验的候选效应物。本研究将有助于从依赖生长阶段的生理或代谢调控的角度分析嗜肺乳杆菌的毒力。
Legionella pneumophila, which is a causative pathogen of Legionnaires' disease, expresses its virulent traits in response to growth conditions. In particular, it is known to become virulent at a post-exponential phase in vitro culture. In this study, we performed a proteomic analysis of differences in expression between the exponential phase and post-exponential phase to identify candidates associated with L. pneumophila virulence using 2-Dimentional Fluorescence Difference Gel Electrophoresis (2D-DIGE) combined with Matrix-Assisted Laser Desorption/Ionization–Mass Spectrometry (MALDI-TOF-MS). Of 68 identified proteins that significantly differed in expression between the two growth phases, 64 were up-regulated at a post-exponential phase. The up-regulated proteins included enzymes related to glycolysis, ketone body biogenesis and poly-3-hydroxybutyrate (PHB) biogenesis, suggesting that L. pneumophila may utilize sugars and lipids as energy sources, when amino acids become scarce. Proteins related to motility (flagella components and twitching motility-associated proteins) were also up-regulated, predicting that they enhance infectivity of the bacteria in host cells under certain conditions. Furthermore, 9 up-regulated proteins of unknown function were found. Two of them were identified as novel bacterial factors associated with hemolysis of sheep red blood cells (SRBCs). Another 2 were found to be translocated into macrophages via the Icm/Dot type IV secretion apparatus as effector candidates in a reporter assay with Bordetella pertussis adenylate cyclase. The study will be helpful for virulent analysis of L. pneumophila from the viewpoint of physiological or metabolic modulation dependent on growth phase.
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