Use of a green fluorescent protein as a marker for human immunodeficiency virus type 1 infection.
We constructed a recombinant human immunodeficiency virus type 1 (HIV-1) provirus called R7-GFP that expresses a modified form of a green fluorescent protein (GFP) from the jellyfish Aequorea victoria by substituting GFP-coding sequences for Nef-coding sequences. Alanine was substituted for serine at amino acid position 65 in the modified GFP, resulting in markedly increased fluorescence at an excitation wavelength of 488 nm as compared to wild-type GFP. The replication kinetics of R7-GFP were identical to that measured with an isogenic, nef-negative strain lacking GFP. Expression of GFP by replication-competent HIV-1 allowed simultaneous quantitation of viral infection and cell surface CD4 levels, revealing rapid and nearly complete CD4 downregulation on R7-GFP-infected PBMCs.
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Gladstone Institute of Virology and Immunology and the Department of Microbiology and Immunology, University of California, San Francisco 94141, USA.
Liegler
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Feinberg
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D015704:CD4 Antigens / Q000096:biosynthesis*; D002478:Cells, Cultured; D015536:Down-Regulation; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins; D015497:HIV-1 / Q000235:genetics / Q000254:growth & development*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000821:virology*; D008164:Luminescent Proteins / Q000235:genetics / Q000302:isolation & purification; D011533:Proviruses / Q000235:genetics / Q000254:growth & development; D014776:Virus Cultivation / Q000379:methods
D015704:CD4 Antigens; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins