HLA-A2 down-regulation on primary human macrophages infected with an M-tropic EGFP-tagged HIV-1 reporter virus
HLA-A2 down-regulation on primary human macrophages infected with an M-tropic EGFP-tagged HIV-1 reporter virus
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DOI：
10.1189/jlb.0505237
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发表时间：
2005-09-01
影响因子：
5.5
通讯作者：
Cheng-Mayer, C
中科院分区：
文献类型：
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作者：
Brown, A;Gartner, S;Cheng-Mayer, C
人类免疫缺陷病毒1型（HIV-1）使用多种机制干扰宿主细胞免疫效应子功能。27-kD Nef蛋白已显示下调感染的原代T细胞表面上的主要组织相容性复合物I类（I）（MHC-I）的特定基因，促进它们逃避溶细胞性T淋巴细胞的裂解。巨噬细胞作为病毒靶向的另一种主要免疫细胞类型，也有助于HIV-1的传播、持续和发病。然而，Nef是否调节HIV感染的原代巨噬细胞上MHC-I的表达仍不清楚。目前可用的表达报告基因的感染性HIV-1分子克隆仅感染T细胞和/或不表达Nef。为了克服这些局限性，我们产生了嗜巨噬细胞的绿色荧光蛋白（GFP）标记的HIV-1病毒，其表达完整的病毒基因组，并使用这些来评估人类白细胞抗原（HLA）-A2在表面的表达有效感染的巨噬细胞。报告病毒基因组是复制能力和稳定的，因为Nef，p24抗原和GFP表达可以通过感染的单核细胞衍生的巨噬细胞（MDM）在感染后2个月以上的免疫染色来检测。感染的巨噬细胞和T细胞的趋化激活细胞分选仪分析显示，尽管野生型报告病毒感染诱导表面BLA-A2密度的统计学显著降低，但在用编码Nef中脯氨酸P-74和P-80处的移码或单点突变的报告病毒感染的细胞中未观察到HLA-A2的下调。Nef对MDM中HLA-A2表面表达的影响也通过共聚焦显微镜证实。这些结果表明，HLA-A2下调的机制在原代T细胞和巨噬细胞中是相似的。
Multiple mechanisms are used by the human immunodeficiency virus type 1 (HIV-1) to interfere with host-cell immune effector functions. The 27-kD Nef protein has been shown to down-modulate specific genes of the major histocompatibility complex class I (I)MHC-I) on the surface of infected primary T cells, facilitating their escape from lysis by cytolytic T lymphocytes. Macrophages, as the other major immune cell type targeted by the virus, also contribute to the transmission, persistence, and pathogenesis of HIV-1. Yet, whether Nef modulates MHC-I expression on HIV-infected primary macrophages remains unclear. Currently available infectious HIV-1 molecular clones, which express a reporter gene, only infect T cells and/or do not express Nef. To overcome these limitations, we generated macrophage-tropic green fluorescent protein (GFP)-tagged HIV-1 viruses, which express the complete viral genome, and used these to assess the expression of human leukocyte antigen (HLA)-A2 on the surface of productively infected macrophages. The reporter viral genomes were replication-competent and stable, as Nef, p24 antigen, and GFP expression could he detected by immunostaining of infected, monocyte-derived macrophages (MDM) after more than 2 months postinfection. Fluorescence-activated cell sorter analyses of infected macrophages and T cells revealed that although wild-type reporter virus infection induced a statistically significant decrease in the density of surface BLA-A2, down-regulation of HLA-A2 was not seen m cells infected with reporter viruses encoding a frame-shift or a single point mutation in Nef at prolines P-74 and P-80. The impact of Nef on HLA-A2 surface expression in MDM was also confirmed by confocal microscopy. These results suggest that the mechanisms of HLA-A2 down-modulation are similar in primary T cells and macrophages.