FIG. 6.
Iron-dependent degradation of IRP1S138E is blocked by proteasomal inhibitors. (A and B) HEK293 cells expressing IRP1S138A or IRP1S138E were treated with the indicated concentrations of hemin or Fe-SIH for 16 h. The expression levels of IRP1S138A (A) and IRP1S138E (B) were analyzed by Western blotting with the FLAG antibody (top). (C and D) HEK293 cells expressing IRP1S138E were treated for the indicated time intervals with 100 μM hemin in the presence or absence of 100 μM MG132 (C) or 10 μM lactacystin (D). The expression of IRP1S138E was analyzed by Western blotting with the FLAG antibody (top). All blots shown were also hybridized with an antibody against β-actin (bottom). The cells were viable under all experimental conditions.