Abstract
Protein kinase C (PKC) is an important constituent of the signaling pathways involved in apoptosis. We report here that like staurosporine, withaferin A is a potent inhibitor of PKC. In Leishmania donovani, the inhibition of PKC by withaferin A causes depolarization of DeltaPsim and generates ROS inside cells. Loss of DeltaPsim leads to the release of cytochrome c into the cytosol and subsequently activates caspase-like proteases and oligonucleosomal DNA cleavage. Moreover, in treated cells, oxidative DNA lesions facilitate the stabilization of topoisomerase I-mediated cleavable complexes, which also contribute to DNA fragmentation. However, withaferin A and staurosporine cannot induce cleavable complex formation in vitro with recombinant topoisomerase I nor with nuclear extracts from control cells. Taken together, our results indicate that inhibition of PKC by withaferin A is a central event for the induction of apoptosis and that the stabilization of topoisomerase I-DNA complex is necessary to amplify apoptotic process.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Apoptosis / drug effects*
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Caspases / metabolism
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Catalysis / drug effects
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Cytochromes c / metabolism
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Cytosol / drug effects
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DNA Cleavage / drug effects
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DNA Fragmentation / drug effects
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DNA Topoisomerases, Type I / metabolism*
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DNA, Protozoan / metabolism*
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Enzyme Activation / drug effects
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Ergosterol / analogs & derivatives*
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Ergosterol / chemistry
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Ergosterol / pharmacology
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Glutathione / metabolism
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Leishmania donovani / cytology*
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Leishmania donovani / drug effects*
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Leishmania donovani / enzymology
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Membrane Potential, Mitochondrial / drug effects
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Oxidative Stress / drug effects
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Protein Kinase C / antagonists & inhibitors
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Protein Kinase Inhibitors / chemistry
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Protein Kinase Inhibitors / pharmacology*
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Reactive Oxygen Species / metabolism
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Staurosporine / chemistry
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Staurosporine / pharmacology
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Withanolides
Substances
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DNA, Protozoan
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Protein Kinase Inhibitors
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Reactive Oxygen Species
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Withanolides
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Cytochromes c
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Protein Kinase C
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Caspases
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DNA Topoisomerases, Type I
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Glutathione
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Staurosporine
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withaferin A
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Ergosterol