FIGURE 4.

Effect of P₃-25 on TRAF2, TANK, and MEKK1 interaction. HuT-78 cells were transfected with vector, TRAF2, or TRAF2-DN constructs. Cells were cultured for 12 h and then treated with P₃-25 (100 nm) for 12 h. The whole cell extracts were prepared and incubated with Sepharose CL 4B beads coupled with anti-TRAF2 (A) or anti-TANK (B) antibody for 4 h. Beads were packed in a 2-ml column, washed with 0.2 m NaCl solution, and then eluted with 0.5 m NaCl solution. The fractions were run in 12% SDS-PAGE, and TRAF2, MEKK1, and TANK were detected by Western blot. IP, immunoprecipitation; WB, Western blot.