HLA-A2 down-regulation on primary human macrophages infected with an M-tropic EGFP-tagged HIV-1 reporter virus.
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 (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 be 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 HLA-A2, down-regulation of HLA-A2 was not seen in cells infected with reporter viruses encoding a frameshift or a single point mutation in Nef at prolines 74P and P80. 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.
Brown
Amanda
A
Department of Neurology, Meyer 6-181, 600 North Wolfe Street, Baltimore, MD 21287, USA. abrown76@jhmi.edu
Gartner
Suzanne
S
Kawano
Thomas
T
Benoit
Nicole
N
Cheng-Mayer
Cecilia
C
D015704:CD4 Antigens / Q000276:immunology; D002460:Cell Line; D015536:Down-Regulation / Q000276:immunology*; D005786:Gene Expression Regulation / Q000276:immunology; D015693:Gene Products, nef / Q000235:genetics / Q000276:immunology; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D015497:HIV-1 / Q000472:pathogenicity / Q000502:physiology*; D015789:HLA-A2 Antigen / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000821:virology*; D008264:Macrophages / Q000276:immunology* / Q000821:virology; D013601:T-Lymphocytes / Q000276:immunology / Q000821:virology; D013997:Time Factors; D014779:Virus Replication; D054311:nef Gene Products, Human Immunodeficiency Virus
D015704:CD4 Antigens; D015693:Gene Products, nef; D015789:HLA-A2 Antigen; C485184:enhanced green fluorescent protein; D054311:nef Gene Products, Human Immunodeficiency Virus; D049452:Green Fluorescent Proteins