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. 2007 Feb 28;104(10):4130–4135. doi: 10.1073/pnas.0610167104

Fig. 2.

Fig. 2.

Vpr interacts with DDB1-Cul4A E3 ubiquitin ligase complexes. (A Left) HA-Vpr and Myc-Cul4 were expressed in cotransfected 293T cells, and HA-Vpr was immunoprecipitated with anti-HA mAb. Coimmunoprecipitated DDB1, myc-Cul4, and Roc1 were detected on an immunoblot. (Right) Expression of the proteins was confirmed by immunoblot analysis of the cell lysates. (B Left) DDB1 was knocked down by siRNA transfection. To detect the association of Vpr with DDB1 Cul4A and Roc1, Vpr was immunoprecipitated, and coimmunoprecipitated proteins were detected on an immunoblot. (Right) The efficiency of DDB1 knockdown was determined by immunoblot analysis of the cell lysates. (C) HA-Vpr point mutants were expressed in transfected 293T cells. HA-Vpr was immunoprecipitated, and the coimmunoprecipitated DDB1 was detected on an immunoblot (Left). Stability of the mutant Vpr was determined by immunoblot analysis of the cell lysates (second panel from the Left). The W54R Vpr mutant was tested in a separate experiment (Right two panels).