Analysis of HIV-1 envelope mutants and pseudotyping of replication-defective HIV-1 vectors by genetic complementation.
Infectious HIV-1 particles containing replication-defective vectors that express the hygromycin B phosphotransferase gene were generated by transient complementation in COS-1 cells. A defective vector dependent only on trans-complementation with an env gene and a small vector containing a deletion of almost all of the trans region were used to examine pseudotyping of HIV-1 by an amphotropic murine retrovirus. Although pseudotyping by the heterologous envelope glycoprotein occurred with efficiency, no pseudotyping at the RNA level was observed. Genetic complementation was used to rapidly analyze the effect of env mutations in the V3, proteolytic processing site, fusion domain, and cytoplasmic tail on viral infectivity. Mutations decreasing syncytium formation usually also lowered infectivity. However, a mutation in the cytoplasmic tail and a separate mutation adjacent to the fusion domain dramatically decreased viral particle infectivity but did not appreciably decrease envelope glycoprotein-mediated cell-to-cell fusion. These results may indicate that these regions of the transmembrane peptide are necessary for acquisition of envelope glycoprotein by budding virus particles or for virus entry.
Delwart
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McArdle Laboratory for Cancer Research, University of Wisconsin, Madison 53706.
Buchschacher
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Freed
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Panganiban
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D000595:Amino Acid Sequence; D002460:Cell Line; D015686:Gene Products, env / Q000235:genetics*; D005816:Genetic Complementation Test; D005822:Genetic Vectors; D015726:Giant Cells / Q000382:microbiology; D015497:HIV-1 / Q000235:genetics* / Q000472:pathogenicity / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids; D014779:Virus Replication / Q000235:genetics
D015686:Gene Products, env