Sapra R, Gaucher S P, Lachmann J S, Buffleben G M, Chirica G S, Comer J E, Peterson J W, Chopra A K, Singh A K
Sandia National Laboratories, 7011 East Avenue, Livermore, CA 94551, USA.
Microb Pathog. 2006 Oct-Nov;41(4-5):157-67. doi: 10.1016/j.micpath.2006.07.002. Epub 2006 Sep 1.
Bacillus anthracis is the etiological agent of anthrax and the bacterium produces a tripartite anthrax toxin composed of protective antigen (PA), lethal factor (LF) and edema factor (EF). PA represents the binding domain of the toxin and acts in concert with either LF, a metalloprotease, or EF, an adenylate cyclase, to form lethal toxin (LeTx) or edema toxin (EdTx), respectively. We analyzed the proteomics response of two murine macrophage cell lines (J774.1A and RAW264.7) following B. anthracis LeTx treatment to detect unique host proteins involved in anthrax infection using difference in-gel electrophoresis (DIGE) followed by nanoLC-MS for identification of the proteins. The comparative proteomics approach identified a set of proteins in each cell line that was consistently upregulated when the two macrophage cell lines were treated with LeTx. The upregulated proteins include those involved in energy metabolism, cytoskeleton structure and stress response. A subset of five proteins (ATP synthase beta subunit, beta-actin, Hsp70, vimentin, and Hsp60 homolog) was identified that were commonly upregulated in both cell lines. The proteomic data suggest the involvement of reactive oxygen species (ROS) in cell lysis as seen by the upregulation of proteins that lead to the production of ROS in both the cell lines used in our study. However, proteins that afford protection against ROS may play an important role in the survival of the macrophage to LeTx infection as shown by the differences in proteomic responses of the two cell lines to the action of LeTx. These identified proteins may have the potential to be used as biomarkers for diagnostics and therapeutics.
炭疽芽孢杆菌是炭疽病的病原体，该细菌产生一种由保护性抗原（PA）、致死因子（LF）和水肿因子（EF）组成的三联体炭疽毒素。PA代表毒素的结合结构域，分别与金属蛋白酶LF或腺苷酸环化酶EF协同作用，形成致死毒素（LeTx）或水肿毒素（EdTx）。我们分析了两种小鼠巨噬细胞系（J774.1A和RAW264.7）在炭疽芽孢杆菌LeTx处理后的蛋白质组学反应，使用差异凝胶电泳（DIGE）检测参与炭疽感染的独特宿主蛋白，随后通过纳升液相色谱-质谱联用技术鉴定这些蛋白。比较蛋白质组学方法在每个细胞系中鉴定出一组蛋白质，当这两种巨噬细胞系用LeTx处理时，这些蛋白质持续上调。上调的蛋白质包括参与能量代谢、细胞骨架结构和应激反应的蛋白质。鉴定出一个由五种蛋白质（ATP合酶β亚基、β-肌动蛋白、热休克蛋白70、波形蛋白和热休克蛋白60同源物）组成的子集，它们在两种细胞系中均普遍上调。蛋白质组学数据表明，活性氧（ROS）参与细胞裂解，这在我们研究中使用的两种细胞系中均表现为导致ROS产生的蛋白质上调。然而，正如两种细胞系对LeTx作用的蛋白质组学反应差异所示，具有抗ROS保护作用的蛋白质可能在巨噬细胞对LeTx感染的存活中起重要作用。这些鉴定出的蛋白质可能具有用作诊断和治疗生物标志物的潜力。
