Figure 1

Characterization of the HIV-1 ASP mutant proviral clone. (A) Schematic representation of the HIV-1 proviral genome. The viral ORFs are presented based on the nature of their encoding transcripts, i.e. multiply-spliced, mono-spliced, and unspliced sense transcripts (red, blue, and white). The antisense strand-encoded asp ORF (green) is also indicated. The reported asp coding region is further indicated below showing the two cysteine triplets, the SH3 binding motif (PxxPxxP), and potential transmembrane regions (TM). The numbers shown indicate amino acid positions. (B) Reduction of extracellular p24 Gag levels from 293T cells transfected with ASP-deficient HIV-1 proviral DNA. 293T cells were cotransfected with pNL4.3WT or pNL4.3ASPmut12 and pRcActin-LacZ. Forty-eight hours after transfection, supernatants were harvested and quantified through a p24 ELISA assay. Results are presented as the average p24 value +/- S.D. of β-galactosidase-normalized values from three independently transfected cell samples. Cell lysates were prepared from non-transfected 293T cells (NT) or cells transfected with pNL4.3WT or pNL4.3ASPmut12. Western blot analyses (under the histogram) were conducted on these preparations using anti-p24 and HRP-conjugated goat anti-rabbit IgG antibodies (two independent transfections are presented per condition). (C) Analysis of WT and ASPmut12 virion morphology. Virus particles produced from 293T cells transfected with pNL4.3WT or pNL4.3ASPmut12 were analyzed in thin-layer electron microscopy. The black bars correspond to a scale of 100 nm.