DNA adducts of 10-decarbamoyl mitomycin C activate p53-dependent and p53-independent cell death
Item
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Title
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DNA adducts of 10-decarbamoyl mitomycin C activate p53-dependent and p53-independent cell death
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Identifier
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d_2009_2013:e1d160b85f31:10148
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identifier
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10326
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Creator
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Boamah, Ernest Kojo,
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Contributor
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Jill Bargonetti
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Date
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2009
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Language
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English
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Publisher
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City University of New York.
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Subject
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Molecular biology | Cell Death | decarbamoyl Mitomycin C | DNA Adducts | Mitomycin C | p53-dependent | p53-independent
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Abstract
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Mitomycin C (MC), a natural antibiotic and DNA cross-linking agent, has cytotoxic activity and is known to activate the tumor suppressor p53 protein. 10-decarbamoyl mitomycin C (DMC), a derivative of MC, has increased cytotoxicity compared to MC. Both MC and DMC induce cellular cytotoxicity in cells with wild-type p53, while only DMC shows significant cell death activity in the absence of wild-type p53. We investigated the difference in MC and DMC cytotoxicity by comparing DNA adduct composition and the cellular regulation of molecular targets in human cancer cell lines with or without wild-type p53. Compared to MC, DMC produced substantially more mitosene-1-beta mono and 1-beta cross-link adducts in DNA and resulted in abnormal nuclear morphology in human cancer cells with or without p53. Significantly, greater poly(ADP-ribose)polymerase (PARP) activity was observed after DMC treatment in both the presence and absence of wild-type p53. Both MC and DMC induced double strand breaks as indicated by gamma-H2AX foci formation irrespective of the p53 status, suggesting that double strand breaks cannot account for DMC's increased cytotoxicity. In cell lines expressing wild-type p53, both MC and DMC signaled for p53 stability and apoptosis induction resulting in cleavage of procaspase-3 and -8. Despite the DMC induced cellular cytotoxicity observed in cell lines lacking wild-type p53, cleavage of procaspase-3 or -8 was not observed in these cells. However, we observed an increase in caspase activity. Caspase-2 activation has been suggested as a pathway for p53-independent cell death in the absence of Chk1. Interestingly, Chk1 was depleted following DMC, but not MC treatment in cells with or without wild-type p53. This Chk1 depletion was achieved through the ubiquitin proteasome pathway since chemical inhibition of the proteasome protected against Chk1 depletion. Additionally, gene silencing of Chk1 by siRNA increased the cytotoxicity of MC but not of DMC. DMC treatment also caused a decrease in the level of total ubiquitinated proteins without increasing proteasome activity. This suggests that DMC-mediated DNA adducts facilitate signal transduction to a pathway targeting proteins for proteolysis. In conclusion, we have found that DMC generates significantly more mitosene-1-beta stereoisomeric DNA adducts than MC and causes rapid down-regulation of multiple cellular targets. These studies suggest increased mitosene-1-beta stereoisomeric DNA adducts more effectively signal for a mode of cell death which does not require a functional p53 protein.
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Type
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dissertation
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Source
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2009_2013.csv
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degree
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Ph.D.
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Program
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Biology
