Fig. 2
From: TRAF3 can interact with GMEB1 and modulate its anti-apoptotic function
Co-immunoprecipitation of TRAF3 and GMEB1 in mammalian cells. HEK293 cells were cotransfected with vectors expressing a His-tagged GMEB1 protein (GMEB1-His) (1 μg) together with (a) a FLAG-tagged TRAF3 protein (TRAF3 wt-flag) (0.5 μg) or a FLAG-tagged mutant TRAF3 M7 (TRAF3 M7-flag) (0.5 μg), defective in the RING domain. b a truncated TRAF3 (TRAF3ΔC-flag) (0.5 μg), which is lacking the TRAF-C domain. 24 h post transfection, the cell extracts were subjected to immunoprecipitation using the Ni–NTA beads (Qiagen), which bind to GMEB1-His. The immunoprecipitated complex (IP) was analyzed by SDS-PAGE on a 7.5% gel and subjected to immunoblot analysis with an anti-His rabbit polyclonal antibody recognizing GMEB1 (His probe) (80 kDa) or with an anti-flag mouse monoclonal antibody recognizing both TRAF3 wt (58 kDa) and TRAF3 M7 (58 kDa)