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 protein kinase C alpha (PKC alpha) isoform (80 KDa). Treatment of LLC-PK1 cells with iodoacetamide (0.5-15 mM) for 30-90 minutes at room temperature allowed by western blotting on total cell homogenate, revealed the appearance of an 50 KDa band that was still recognized with the antibody. However, 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 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 activities in iodoacetamide-treated cells were also inhibited. In conclusion, the degradation of PKC alpha 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 degrees C and appears to involve interactions with unknown cellular intermediates.
Dibas
A
A
Department of Pharmacology, University of North Texas Health Science Center at Fort Worth, 76107, USA. adibas@hsc.unt.edu
Wood
J
J
Mia
A J
AJ
Yorio
T
T
D000255:Adenosine Triphosphate / Q000037:antagonists & inhibitors*; D000818:Animals; D006868:Hydrolysis; D007460:Iodoacetamide / Q000494:pharmacology*; D007527:Isoenzymes / Q000378:metabolism*; D007668:Kidney / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D018374:LLC-PK1 Cells; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D013552:Swine
D007527:Isoenzymes; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D007460:Iodoacetamide