The virion-associated Gag-Pol is decreased in chimeric Moloney murine leukemia viruses in which the readthrough region is replaced by the frameshift region of the human immunodeficiency virus type 1.
The human immunodeficiency virus type 1 (HIV-1) requires a programmed -1 translational frameshift event to synthesize the precursor of its enzymes, Gag-Pol, when ribosomes from the infected cells translate the full-length viral messenger RNA. Translation of the same RNA according to conventional translational rules produces Gag, the precursor of the structural proteins of the virus. The efficiency of the frameshift controls the ratio of Gag-Pol to Gag, which is critical for viral infectivity. The Moloney murine leukemia virus (MoMuLV) uses a different strategy, the programmed readthrough of a stop codon, to synthesize Gag-Pol. In this study, we investigated whether different forms of the HIV-1 frameshift region can functionally replace the readthrough signal in MoMuLV. Chimeric proviral DNAs were obtained by inserting into the MoMuLV genome the HIV-1 frameshift region encompassing the slippery sequence where the frameshift occurs, followed by the frameshift stimulatory signal. The inserted signal was either a simple stem-loop, previously considered as the stimulatory signal, or a longer bulged helix, now shown to be the complete stimulatory signal, or a mutated version of the complete signal with a three-nucleotide deletion. Although the three chimeric viruses can propagate essentially as the wild-type virus in NIH 3T3 cells, single-round infectivity assays revealed that the infectivity of the chimeric virions is about three to fivefold lower than that of the wild-type virions, depending upon the nature of the frameshift signal. It was also observed that the Gag-Pol to Gag ratio was decreased about two to threefold in chimeric virions. Comparison of the readthrough efficiency of MoMuLV to the HIV-1 frameshift efficiency, by monitoring the expression of a luciferase reporter in cultured cells, revealed that the frameshift efficiencies were only 30-60% of the readthrough efficiency. Altogether, these observations indicate that replacement of the readthrough region of MoMuLV with the frameshift region of HIV-1 results in virions that are replication competent, although less infectious than wild-type MoMuLV. This type of chimera could provide an interesting tool for in vivo studies of novel drugs targeted against the HIV-1 frameshift event.
Gendron
Karine
K
Département de Biochimie, Université de Montréal, 2900, Boulevard Edouard-Montpetit, D-353, Montréal, Québec, Canada H3T 1J4.
Dulude
Dominic
D
Lemay
Guy
G
Ferbeyre
Gerardo
G
Brakier-Gingras
Léa
L
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D018965:Frameshifting, Ribosomal; D015685:Fusion Proteins, gag-pol / Q000378:metabolism*; D015967:Gene Expression Regulation, Viral; D015750:Genes, gag; D015751:Genes, pol; D015497:HIV-1 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D008979:Moloney murine leukemia virus / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D041681:NIH 3T3 Cells; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D014771:Virion / Q000378:metabolism* / Q000472:pathogenicity; D014779:Virus Replication
D015685:Fusion Proteins, gag-pol; D011993:Recombinant Fusion Proteins