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. 2007 Aug 16;26(17):3910–3922. doi: 10.1038/sj.emboj.7601823

Figure 8.

Figure 8

TAX1BP1 interacts with RIP1 and negatively regulates RIP1 ubiquitination. (A) 293T cells were transfected with Flag-RIP1 (1 μg), Flag-TRAF6 (1 μg) and GFP-TAX1BP1 (1 μg). After 36 h, cells were lysed and immunoprecipitated with anti-GFP followed by immunoblotting with anti-Flag. Lysates were examined for TAX1BP1 expression by immunoblotting with anti-GFP. RIP1 and TRAF6 were detected by immunoblotting with anti-Flag. (B) TAX1BP1m/m and TAX1BP1+/m MEFs were treated with TNF-α (20 ng/ml) as indicated. Cells were lysed and RIP1 was immunoprecipitated with anti-RIP1 followed by immunoblotting with anti-ubiquitin or anti-RIP1. (C) 293T cells were transfected with Flag-A20 (1 μg) and HA-RIP1 (1 μg) together with pCAGGS-TAX1BP1 (0, 0.25, 0.5 and 1 μg). After 36 h, cells were lysed and RIP1 was immunoprecipitated with anti-HA followed by immunoblotting with anti-Flag. Lysates were examined for A20, RIP1 and TAX1BP1 expression by immunoblotting with anti-Flag, anti-HA and anti-TAX1BP1, respectively. (D) 293T cells were transfected with Flag-TAX1BP1 together with either control or TAX1BP1 siRNA. (E) 293T cells were transfected with control or TAX1BP1 siRNA. Cells were either untreated or treated with TNF-α (20 ng/ml) for the indicated times. Immunoprecipitations were performed with either anti-control Ig or anti-RIP1 followed by immunoblotting with anti-A20.