THE ATP-DEPLETING REAGENT IODOACETAMIDE INDUCES THE DEGRADATION OF PROTEIN-KINASE-C-ALPHA (PKC-ALPHA) IN LLC-PK1 PIG-KIDNEY CELLS

The alkylating reagent iodoacetamide, a potent inhibitor of sulfhydryl proteases, was found to stimulate the selective degradation of protei n kinase C alpha (PKC alpha) isoform (80 KDa). Treatment of LLC-PK1 ce lls with iodoacetamide (0.5-15 mM) for 30-90 minutes at room temperatu re followed by western blotting on total cell homogenate, revealed the appearance of an 50 KDa band that was still recognized with the antib ody. However, iodoacetamide (15 mM) resulted in the total disappearanc e of the 80 KDa protein. Serine protease inhibitors, metalloprotease i nhibitors and leupeptin failed to prevent the degradation of PKC alpha . The degradation persisted at 4 degrees C and in the absence of Ca2+. Iodoacetamide had no direct effect on purified PKC alpha. PKC activit ies in iodoacetamide-treated cells were also inhibited. In conclusion, the degradation of PKC alpha is a novel phenomenon. The degradation p rocess could not be prevented by known protease inhibitors or in the a bsence of Ca2+ or by incubation at 4 degrees C and appears to involve interactions with unknown cellular intermediates.