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Figure 3 | Retrovirology

Figure 3

From: Human Polycomb groupEED protein negatively affects HIV-1 assembly and release

Figure 3

Influence of EED3/4 expression on virus yields. (a), Experimental protocol. (b), Virus infectivity. Virions produced by 293T cells transfected with pTracer-EED (filled symbols) or control pTracer-Emp plasmid (open symbols) were used to infect recipient 293T cells, and luciferase expression monitored at different times pi, as indicated. (c, d), Vector titer was determined by CAp24 immunoassay (c) or genomic RNA level (d). (e-g), SDS-PAGE polypeptide pattern of virus particles released from cells transfected with pTracer-EED (+EED) or control pTracer-Emp plasmid (-EED). Blots were reacted with anti-Gag (e; f, bottom panel ; g, bottom panel), anti-VSV-G (f, top panel) and anti-Nef (g, top panel) antibodies. Virus production was significantly reduced in the presence of EED, ranging from 20- to 25-fold at 24–48 h posttransfection, based on CAp24 immunoassay (c), to 10- to 80-fold, based on genomic RNA levels (d). The Gag protein composition (e), the VSV-G-to-CAp24 (f) and the Nef-to-CAp24 (g) ratios did not differ significantly between particles produced in the presence or absence of EED. Note that the load of virus samples produced in the presence of EED (e ; +EED) was 5-fold higher than control samples (-EED), and that coexpression of Nef restored the virus yields, as shown by CAp24 immunoblotting (g ; bottom panel).

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