A quasi-lentiviral green fluorescent protein reporter exhibits nuclear export features of late human immunodeficiency virus type 1 transcripts.
We have previously shown that Rev-dependent expression of HIV-1 Gag from CMV immediate early promoter critically depends on the AU-rich codon bias of the gag gene. Here, we demonstrate that adaptation of the green fluorescent protein (GFP) reporter gene to HIV codon bias is sufficient to turn this hivGFP RNA into a quasi-lentiviral message following the rules of late lentiviral gene expression. Accordingly, GFP expression was significantly decreased in transfected cells strictly correlating with reduced RNA levels. In the presence of the HIV 5' major splice donor, the hivGFP RNAs were stabilized in the nucleus and efficiently exported to the cytoplasm following fusion of the 3' Rev-responsive element (RRE) and coexpression of HIV-1 Rev. This Rev-dependent translocation was specifically inhibited by leptomycin B suggesting export via the CRM1-dependent pathway used by late lentiviral transcripts. In conclusion, this quasi-lentiviral reporter system may provide a new platform for developing sensitive Rev screening assays.
Graf
Marcus
M
Institute of Medical Microbiology and Hygiene, Molecular Microbiology and Gene Therapy, University of Regensburg, 93053 Regensburg, Germany.
Ludwig
Christine
C
Kehlenbeck
Sylvia
S
Jungert
Kerstin
K
Wagner
Ralf
R
D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D003062:Codon / Q000235:genetics; D017930:Genes, Reporter; D005814:Genes, Viral; D049452:Green Fluorescent Proteins / Q000235:genetics* / Q000378:metabolism*; D015497:HIV-1 / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic
D003062:Codon; D012367:RNA, Viral; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins