Caspase-mediated protein kinase C-delta cleavage is necessary for apoptosis of vascular smooth muscle cells.
Apoptotic death of vascular smooth muscle cells (SMCs) is a prominent feature of blood vessel remodeling and various vascular diseases. We have previously shown that protein kinase C-delta (PKC-delta) plays a critical role in SMC apoptosis. In this study, we tested the importance of PKC-delta proteolytic cleavage and tyrosine phosphorylation within the apoptosis pathway. Using hydrogen peroxide as a paradigm for oxidative stress, we showed that proteolytic cleavage of PKC-delta occurred in SMCs that underwent apoptosis, while tyrosine phosphorylation was detected only in necrotic cells. Furthermore, using a peptide (z-DIPD-fmk) that mimics the caspase-3 binding motif within the linker region of PKC-delta, we were able to prevent the cleavage of PKC-delta, as well as apoptosis. Inhibition of PKC-delta with rottlerin or small-interfering RNA diminished caspase-3 cleavage, caspase-3 activity, cleavage of poly (ADP-ribose) polymerase, cleavage of PKC-delta, and DNA fragmentation, confirming the previously reported role of PKC-delta in initiation of apoptosis. In contrast, z-DIPD-fmk markedly diminished caspase-3 activity, cleavage of PKC-delta, and DNA fragmentation without affecting cleavage of caspase-3 and poly (ADP-ribose) polymerase. Taken together, our data suggest that caspase-3-mediated PKC-delta cleavage underlies SMC apoptosis induced by oxidative stress, and that PKC-delta acts both upstream and downstream of caspase-3.
Kato
Kaori
K
Department of Surgery, University of Wisconsin, Madison, Wisconsin 53705, USA.
Yamanouchi
Dai
D
Esbona
Karla
K
Kamiya
Kentaro
K
Zhang
Fan
F
Kent
K Craig
KC
Liu
Bo
B
D000098:Acetophenones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001578:Benzopyrans / Q000494:pharmacology; D053148:Caspase 3 / Q000378:metabolism*; D002460:Cell Line; D053938:DNA Fragmentation; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D006861:Hydrogen Peroxide / Q000633:toxicity; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology* / Q000473:pathology; D009842:Oligopeptides / Q000494:pharmacology; D016877:Oxidants / Q000633:toxicity; D018384:Oxidative Stress; D010766:Phosphorylation; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D051745:Protein Kinase C-delta / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D034622:RNA Interference; D051381:Rats; D013997:Time Factors; D014443:Tyrosine
D000098:Acetophenones; D001578:Benzopyrans; D009842:Oligopeptides; D016877:Oxidants; D047428:Protein Kinase Inhibitors; C516668:benzyloxycarbonyl-aspartyl(OMe)-isoleucyl-prolyl-aspartyl(OMe) fluoromethylketone; D014443:Tyrosine; D006861:Hydrogen Peroxide; C085746:rottlerin; D011065:Poly(ADP-ribose) Polymerases; C497260:Prkcd protein, rat; D051745:Protein Kinase C-delta; C505410:Casp3 protein, rat; D053148:Caspase 3