Figure 7

HIV-1 produced in A549 cells overexpressing Myc-PRMT6 are competent for infectivity. (A) A549 cells were transfected with a HIV-1 proviral plasmid in which the EGFP gene had been inserted into the nef reading frame, along with a plasmid expressing the VSV-G envelope glycoprotein to enable pseudotyping. Cells were simultaneously cotransfected with either empty vector (-), the wild type Myc-PRMT6 plasmid (WT) or a methyltransferase-inactive mutant Myc-PRMT6 plasmid (Mut.). Western blotting was performed on producer cell lysates using an anti-PRMT6 antibody. Loading of cell lysates was normalized for equal amounts of co expressed Renilla luciferase in each sample. (B) HeLa and A549 target cells were infected with the pseudotyped viruses collected from the A549 producer cells in A after normalizing for capsid levels. EGFP expression in the target cells, which indicates successful infection, was then quantified by flow cytometry. A representative dot plot of target cells infected with virus from the empty vector-cotransfected producer cells is shown, in which GFP-positive, phycoerythrin (PE)-negative cells are gated (green). (C) The proportions of GFP-positive HeLa (black columns) and A549 (white columns) target cells were quantified for each infection sample and are shown arranged by the Myc-PRMT6 plasmid cotransfected in the virus producer cells. Data are expressed relative to the respective empty vector sample (“None”), and columns represent the means and standard deviations of two independent infections using independent virus stocks.