TRAF2 and RIP1 interact in the context of CD40 signaling. (A) Ectopic expression of CD40 enhances the interaction of RIP1 with endogenous TRAF2. Two million 293 cells were transfected with 0.5 µg FLAG-tagged RIP1 in the presence of increasing amounts of a CD40 expression vector (0, 1, or 2.5 µg). Lysates were immunoprecipitated (IP) with anti-FLAG and immunoblotted (IB) with an anti-TRAF2 or RIP1 antibody as indicated. (B and C) CD40 ligation induces the association of endogenous RIP1 with TRAF2 but not CD40. HeLa/CD40mT6 cells were stimulated with CD154, then lysates were sequentially immunoprecipitated with anti-CD40 (B) and anti-RIP1 (C) and immunoblotted with TRAF2, RIP1, or CD40 antibodies.