A rescue of cell death by concurrent introduction of CTCFL would show that the t

Polypeptides associated with GST vIRF fusion proteins after pull down assays were immunoblotted with an anti p53 antibody. This confirmed that GST vIRF fusion proteins containing the possible DNA binding site of vIRF might bind to p53 in vitro, In contrast, GST and GST vIRF fusion proteins containing the amino terminal pro line loaded region or the carboxyl service region did not bind to p53 Bromosporine Epigenetic Reader Domain under the same circumstances, The interaction between vIRF and p53 was further evaluated by in vivo coimmunoprecipitation assay. Four deletion muta tions were developed the following. 1 vector and expressed in COS 1 cells. After transfection, whole cell lysates were used for immunoblotting having an anti Flag antibody. Wild-Type and mutant vIRF were indicated at relatively variable but still comparable levels in COS 1 cells, Precisely the same cell lysates were used for immunoprecipitation with an anti,Flag Immune system antibody, followed closely by immunoblotting with an anti p53 antibody to detect the clear presence of p53 in anti Flag immune processes, The results demonstrated that vIRFmt2, vIRFmt4, and vIRFmt5 interacted with p53, whereas vIRFmt3 did not. Several parts of p53 interact with vIRF. To further de lineate an interaction between p53 and vIRF, we attempted to dene the regions of p53 needed for this interaction. The p53 protein contains PF-04620110 Transferase inhibitor ve distinct areas. A series of GFP p53 fusion protein containing individual areas of p53,were produced as described in Fig. 4A.