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Figure 4 | Retrovirology

Figure 4

From: Vpx rescue of HIV-1 from the antiviral state in mature dendritic cells is independent of the intracellular deoxynucleotide concentration

Figure 4

Vpx increases HIV-1 2-LTR circles in LPS-treated MDDCs. 2 × 106 MDDCs were stimulated with 100 ng/ml LPS for 18 hrs, treated for 2 hrs with either ΔVpx-VLPs (ΔVpx) or Vpx-VLPs alone or combined with 2.5 mM deoxynucleosides (dN) and infected with NL4-3 GFP or NL4-3 GFP lacking VSV G protein (ΔVSVg) as negative control. 24 hrs later low molecular weight DNA was isolated and real-time PCR for full length HIV-1 cDNA (LRT) (A) or 2-LTR circles (B) was performed and the ratio of fold change of 2-LTR circles over LTR was displayed (C). LRT and 2-LTR circles were normalized to mitochondrial DNA and compared to ΔVpx levels. GFP positve cells were assessed for GFP expression by flow cytometry after 72 hrs (D). MDDC from two donors were stimualted with 100 ng/ml LPS, treated with either ΔVpx-VLPs, Vpx-VLPs or 2.5 mM deoxynucleosides and infected 3 hrs later. Low molecular DNA from single samples was isolated 6 hrs p.i. and real-time PCR for full length HIV-1 cDNA was prefromed (E). MDDC were treated with either ΔVpx-VLPs, Vpx-VLPs or 2.5 mM deoxynucleosides and infected 3 hrs later. Low molecular DNA from single samples was isolated 12 hrs p.i. for real-time PCR for full length HIV-1 cDNA and 24 hrs p.i for 2-LTR cirlce real-time PCR (F).

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