The ATP-depleting reagent iodoacetamide induces the degradation of protein kinase C alpha (PKCα) in LLC-PK1 pig kidney cells

The alkylating reagent iodoacetamide, a potent inhibitor of sulfhydryl proteases, was found to stimulate the selective degradation of protein kinase C alpha (PKCα) isoform (80 KDa). Treatment of LLC-PK, cells with iodoacetamide (0.5-15 mM) for 30-90 minutes at room temperature followed by western blotting on total cell homogenate, revealed the appearance of an 50 KDa band that was still recognized with the antibody. iodoacetamide (15 mM) resulted in the total disappearance of the 80 KDa protein. Serine protease inhibitors, metalloprotease inhibitors and leupeptin failed to prevent the degradation of PKCα. The degradation persisted at 4°C and in the absence of Ca²⁺. Iodoacetamide had no direct effect on purified PKCa. PKC activities in iodoacetamide-treated cells were also inhibited. In conclusion, the degradation of PKCα is a novel phenomenon. The degradation process could not be prevented by known protease inhibitors or in the absence of Ca²⁺ or by incubation at 4°C and appears to involve interactions with unknown cellular intermediates.