Journal: FEBS letters

Article Title: Agonist antibody activates death receptor 6 downstream signaling involving TRADD recruitment.

doi: 10.1016/j.febslet.2013.12.010

Figure Lengend Snippet: Fig. 5. TRADD is involved in DR6-induced IL-6 activation. (A and B) HEK293T cells were transiently transfected with IL-6 reporter plasmid, Renilla and increasing amounts of expression vectors for TRADD or TRADD-DN (A) or together with DR6-FL (B). 24 h later, promoter activity was measured by luciferase assay. (C) HEK293T cells were co- transfeced with IL-6-Luc, Renilla, DR6-FL and TRADD as described in (B), and then stimulated with DQM3. 16 h later, luciferase activity was measured. (D) HEK293T cells were co-transfected with TRADD siRNA (si-TRADD) or negative control siRNA (si-NC) and expression plasmid for TRADD. Western blot analysis was performed to verify the efficiency of siRNA. (E) TRADD siRNA-treated cells were transfected with IL-6 reporter plasmid and DR6-FL. 24 h after transfection, cells were subjected to DQM3 stimulation for 16 h, and then luciferase activity was monitored as described above. Results are shown as the mean ± S.E.M. ⁄P < 0.05; ⁄⁄P < 0.01.

Article Snippet: The rabbit polyclonal anti-TRADD antibody was purchased from Abcam.

Techniques: Activation Assay, Transfection, Plasmid Preparation, Expressing, Activity Assay, Luciferase, Negative Control, Western Blot

Journal: FEBS letters

Article Title: Agonist antibody activates death receptor 6 downstream signaling involving TRADD recruitment.

doi: 10.1016/j.febslet.2013.12.010

Figure Lengend Snippet: Fig. 6. DR6 interacts with TRADD in co-immunoprecipitation assays. (A and B) HEK293T cells in 60-mm dishes co-transfected with DR6-FL/EX and Flag-tagged TRADD or TRADD-DN were harvested 48 h after transfection. Total cell lysates were immunoprecipitated with anti-DR6 (DQM3) and immunoblotted with anti-TRADD (A) or anti-Flag (B). Western blot analysis of DR6, TRADD and Flag was performed on whole cell lysates as a control (Input). (C) HEK293T cells were co-transfected with expression vectors for DR6-FL and Flag-tagged TRADD, and then stimulated or not with DQM3 for the indicated times. The interaction of TRADD with DR6 was assessed after DQM3 stimulation by co-immunoprecipitation. The immunoprecipitates and cell extracts were analyzed by immunoblot. Results representative of at least three independent experiments are shown. b-Actin was used as loading control for each fraction. ⁄, Heavy chain (DQM3); IP, immunoprecipitates; IB, immunoblot.

Article Snippet: The rabbit polyclonal anti-TRADD antibody was purchased from Abcam.

Techniques: Immunoprecipitation, Transfection, Western Blot, Control, Expressing