HIV-1 Vpr modulates macrophage metabolic pathways: a SILAC-based quantitative analysis.
HIV-1 Vpr modulates macrophage metabolic pathways: a SILAC-based quantitative analysis.
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DOI：
10.1371/journal.pone.0068376
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Merali S
中科院分区：
文献类型：
--
作者：
Barrero CA;Datta PK;Sen S;Deshmane S;Amini S;Khalili K;Merali S
人类免疫缺陷病毒1型编码的病毒蛋白Vpr是HIV-1感染巨噬细胞所必需的。此外，这些巨噬细胞对细胞死亡具有抵抗力，是病毒库。然而，VPR对巨噬细胞蛋白质组的影响尚不清楚。本研究的目的是使用细胞培养中氨基酸稳定同位素标记(SILAC)结合基于质谱学的蛋白质组学方法来表征巨噬细胞中的VPR反应。将培养的人单核细胞U937分化为巨噬细胞，并用携带VPR基因的腺病毒载体进行转导。SILAC结合LC-MS/MS方法对600多个蛋白质进行了定量，其中136个蛋白质在巨噬细胞中过表达后发生了显著变化。利用独创性计算路径分析，通过生物功能、路径和网络分析来选择和聚类量化的蛋白质。蛋白质组数据显示糖酵解途径(磷酸戊糖和丙酮酸代谢)中酶的丰度增加，并通过蛋白质印迹分析进一步验证。此外，蛋白质组学数据显示一些关键的线粒体酶如谷氨酸脱氢酶2(GLUD2)、腺苷酸激酶2(AK2)和转酮醇酶(TKT)表达下调。基于这些观察，我们推测HIV-1通过VPR-低氧诱导因子1α(HIF-1α)轴劫持巨噬细胞的葡萄糖代谢途径，诱导己糖激酶(HK)、葡萄糖-6-磷酸脱氢酶(G6PD)和丙酮酸激酶2型(PKM2)的表达，从而促进病毒的复制和生物发生，并促进巨噬细胞的长期生存。此外，在神经艾滋病的背景下，巨噬细胞内线粒体谷氨酸代谢的失调可通过神经兴奋毒性机制促进神经变性。
Human immunodeficiency virus type 1 encoded viral protein Vpr is essential for infection of macrophages by HIV-1. Furthermore, these macrophages are resistant to cell death and are viral reservoir. However, the impact of Vpr on the macrophage proteome is yet to be comprehended. The goal of the present study was to use a stable-isotope labeling by amino acids in cell culture (SILAC) coupled with mass spectrometry-based proteomics approach to characterize the Vpr response in macrophages. Cultured human monocytic cells, U937, were differentiated into macrophages and transduced with adenovirus construct harboring the Vpr gene. More than 600 proteins were quantified in SILAC coupled with LC-MS/MS approach, among which 136 were significantly altered upon Vpr overexpression in macrophages. Quantified proteins were selected and clustered by biological functions, pathway and network analysis using Ingenuity computational pathway analysis. The proteomic data illustrating increase in abundance of enzymes in the glycolytic pathway (pentose phosphate and pyruvate metabolism) was further validated by western blot analysis. In addition, the proteomic data demonstrate down regulation of some key mitochondrial enzymes such as glutamate dehydrogenase 2 (GLUD2), adenylate kinase 2 (AK2) and transketolase (TKT). Based on these observations we postulate that HIV-1 hijacks the macrophage glucose metabolism pathway via the Vpr-hypoxia inducible factor 1 alpha (HIF-1 alpha) axis to induce expression of hexokinase (HK), glucose-6-phosphate dehyrogenase (G6PD) and pyruvate kinase muscle type 2 (PKM2) that facilitates viral replication and biogenesis, and long-term survival of macrophages. Furthermore, dysregulation of mitochondrial glutamate metabolism in macrophages can contribute to neurodegeneration via neuroexcitotoxic mechanisms in the context of NeuroAIDS.
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