FIG. 6.

Activation of NF-κB in response to ER stress involves release but not proteolysis of IκB. S/S (lanes 1 to 7) and A/A (lanes 8 to 14) MEF cells were treated with thapsigargin for up to 6 h, with TNF-α for 30 min, or with no stress (0 h). Equal amounts of whole-cell lysates were separated by SDS-PAGE; phosphorylated eIF2α (A), total eIF2α (B), IκBα phosphorylated at Ser-32 (C), total levels of IκBα and IκBβ (D), and p65 (E) were measured by immunoblotting using specific antibodies. IκBα was immunoprecipitated and the levels of p65 (F) or IκBα (G) in the immunocomplexes were measured by immunoblot analysis. The IgG present in the immunocomplex migrated near p65 in the SDS-PAGE as indicated in panel F. Each panel is derived from one immunoblotting experiment, and the dotted line between lanes 7 and 8 is shown only for alignment purposes.