Rao Prahlad K, Singh Christoher R, Jagannath Chinnaswamy, Li Qingbo
Int J Clin Exp Med. 2009 Sep 30;2(3):233-47.
Systems biology and proteomics have recently contributed significantly to the insight into the biogenesis and immunity-related functions of the phagosome. To gain insight into the modulation of the phagosomal proteome by the wild-type Mycobacterium tuberculosis H37Rv reference strain, an attenuated mutant of the H37Rv strain, and the BCG Pasteur vaccine strain, we employed the nano-liquid chromatography/LTQ-FTMS based proteomics approach and a systems biology approach to analyze the bacillus-containing phagosomes purified from the bone-marrow-derived BMA3.A3 macrophages infected with the three different mycobacterial strains. We identified 322 proteins at a false-discovery rate of 2%. These proteins were quantified with a label-free proteomics method. All but one of these proteins is mouse proteins. The gene ontology analysis of these mouse proteins suggests that lysosomal proteins represented <3% of the detected proteins, supporting the observation that these mycobacterial strains inhibit or limit the phagosome maturation process. The results also indicate that the endoplasmic reticulum (ER) proteins do not constitute a major part of the phagosome proteome, supporting the phagosome maturation model of the role of ER in phagosome biogenesis. This phagosome maturation model is in contrast to the phagocytosis model which predicts that half of the phagosome membrane is derived from ER. This pilot study demonstrates that a combination of proteomics, multivariate analysis, and systems biology promises to bring forward new insights into the mycobacterial pathogenesis and the interconnected phagosome biology.
系统生物学和蛋白质组学最近在深入了解吞噬体的生物发生和免疫相关功能方面做出了重大贡献。为了深入了解野生型结核分枝杆菌H37Rv参考菌株、H37Rv菌株的减毒突变体以及卡介苗巴斯德疫苗菌株对吞噬体蛋白质组的调节作用，我们采用了基于纳升液相色谱/LTQ-FTMS的蛋白质组学方法和系统生物学方法，来分析从感染了这三种不同分枝杆菌菌株的骨髓来源BMA3.A3巨噬细胞中纯化得到的含杆菌吞噬体。我们以2%的错误发现率鉴定出了322种蛋白质。这些蛋白质采用无标记蛋白质组学方法进行定量。这些蛋白质中除一种外均为小鼠蛋白。对这些小鼠蛋白的基因本体分析表明，溶酶体蛋白占检测到的蛋白不到3%，这支持了这些分枝杆菌菌株抑制或限制吞噬体成熟过程的观察结果。结果还表明，内质网（ER）蛋白并不构成吞噬体蛋白质组的主要部分，这支持了内质网在吞噬体生物发生中作用的吞噬体成熟模型。这种吞噬体成熟模型与预测吞噬体膜一半来源于内质网的吞噬作用模型形成对比。这项初步研究表明，蛋白质组学、多变量分析和系统生物学的结合有望为分枝杆菌发病机制和相互关联的吞噬体生物学带来新的见解。
