Figure 1.

EGF signaling causes FOXO4 down regulation with concurrent elevation of CSN6. A) HCT116 (left) and DLD1 (right) cells were treated with 100 ng mL⁻¹ EGF for the indicated minutes. Cell lysates were analyzed by immunoblotting with indicated antibodies. B) HCT116 cells were transfected with His‐ubi plasmid and treated with 100 ng mL⁻¹ EGF for the indicated times. Cells were treated with proteasome inhibitor MG132 before collecting lysate. Cells were lysed in denaturing buffer (6 m guanidine‐HCl). The cell lysates were then pull down (PD) with nickel beads and immunoblotted with indicated antibodies. C) HCT116 cells were treated with 100 ng mL⁻¹ EGF for the indicated minutes with or without MG132. Lysates were immunoblotted with the indicated antibodies. D) HCT116 cells infected with either CSN6‐shRNA or control shRNA were treated with 100 ng mL⁻¹ EGF for the indicated minutes. Cell lysates were immunoblotted with the indicated antibodies. E) DLD1 (left) or 293T cells were cotransfected with the indicated CSN6, FOXO4 (middle), or FOXO4A3 (right) expression vectors. Cell lysates were immunoblotted with indicated antibodies. *Nonspecific. F) Cell lysates of HCT116 cells infected with CSN6 shRNA were immunoblotted with indicated antibodies. G) DLD1 (left) or 293T (middle and right) cells were cotransfected with the indicated plasmids. Cell lysates were immunoblotted with indicated antibodies. *Nonspecific. H) Cell lysates of HCT116 cells infected with COP1 shRNA were immunoblotted with indicated antibodies.