FIGURE 6.

SIAH1 binds and ubiquitinates MyD88. (A) HeLa cells were treated with MG132 for 6 h and then transfected with plasmids expressing SIAH1-FLAG and MyD88-myc in the presence of MG132. Twenty-four hours post-transfection, total cell lysates were collected, immunoprecipitated with α-FLAG or an isotype control antibody, and analyzed by western blot (TCL, total cell lysate). (B) HeLa cells were transfected with or without SIAH1 siRNA as well as with plasmids expressing MyD88-FLAG and Ub-myc. Twenty-four hours post-transfection, cells were treated with thapsigargin to induce SIAH1 expression, and 2 days post-transfection with MG132 to inhibit the proteasome. Total cell lysates were immunoprecipitated with α-myc and analyzed by western blot.