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. 2024 Jan 19;10(3):eadi4162. doi: 10.1126/sciadv.adi4162

Fig. 2. Tat enhances NF-κB activation through direct interaction with TRAF6.

Fig. 2.

(A) Top: Domain representation of TRAF6. Bottom: Flag-tagged wild-type TRAF6 and truncation mutants (TRAF6 N or TRAF6 C) were cotransfected with Tat-Strep (Tat-S) into HEK293T cells. Tat was immunoprecipitated using an anti-Strep antibody to assess the interaction with TRAF6. (B) Luciferase reporter assays to measure NF-κB activation in HEK293T cells transfected with empty vector or plasmids encoding TRAF6, TRAF2, or TRAF3 together with an empty vector or Tat for 24 hours. (C) Co-IP analysis (with anti-Flag) and Western blot analysis (with anti-Flag or anti-Strep) of HEK293T cells transfected with plasmids encoding Tat-Strep and Flag-tagged TRAF6, TRAF2, TRAF3, or the empty vector for 48 hours. The graphs in (B) show the means ± SD of three biological replicates and were normalized by cotransfected NLuc activities. ns P > 0.05, ****P < 0.0001, and statistical significance was assessed by a two-tailed unpaired Student’s t test. GAPDH, glyceraldehyde-3-phosphate dehydrogenase.