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Figure 3 | Virology Journal

Figure 3

From: Interactions with DCAF1 and DDB1 in the CRL4 E3 ubiquitin ligase are required for Vpr-mediated G2 arrest

Figure 3

The species-specificity of SIV agm Vpr is correlated with an inability to associate with DDB1 in human cells. (A) 293 T cells were transfected with SIVmac Vpr or SIVagm Vpr, Flag-DCAF1, and EGFP expression vectors. After two days, the cells were fixed, stained with propidium iodide and analyzed by flow cytometry. The G2:G1 ratio was calculated after gating for the GFP+ cells. The results are representative of three independent experiments. (B) COS cells were transfected and the cell cycle profiles were analyzed as in (A). (C) 293 T cells were cotransfected with myc-Vpr of SIVmac or SIVagm and Flag-DCAF1 expression vectors. Myc-Vpr was immunoprecipitated with anti-myc MAb and the immunoprecipitated complexes were analyzed on an immunoblot probed with anti-Flag MAb, anti-DDB1 antibody, and anti-myc MAb. (D) COS cells were transfected with myc-Vpr of SIVmac or SIVagm. Myc-Vpr was immunoprecipitated with anti-myc MAb and coimmunoprecipitated endogenous AGM DCAF1 and DDB1 were detected with anti-DCAF1 and anti-DDB1 antibodies.

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