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MLH1 mediates PARP-dependent cell death in response to the methylating agent N-methyl-N-nitrosourea

Biomedical Sciences Research Institute Computer Science Research Institute Environmental Sciences Research Institute Nanotechnology & Advanced Materials Research Institute

McDaid, J. R., Loughery, J., Dunne, P., Boyer, J. C., Downes, Stephen, Farber, R. A. and Walsh, Colum (2009) MLH1 mediates PARP-dependent cell death in response to the methylating agent N-methyl-N-nitrosourea. BRITISH JOURNAL OF CANCER, 101 (3). pp. 441-451. [Journal article]

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DOI: 10.1038/sj.bjc.6605186

Abstract

BACKGROUND: Methylating agents such as N-methyl-N-nitrosourea (MNU) can cause cell cycle arrest and death either via caspase-dependent apoptosis or via a poly(ADP-ribose) polymerase (PARP)-dependent form of apoptosis. We wished to investigate the possible role of MLH1 in signalling cell death through PARP. METHODS: Fibroblasts are particularly dependent on a PARP-mediated cell death response to methylating agents. We used hTERT-immortalised normal human fibroblasts (WT) to generate isogenic MLH1-depleted cells, confirmed by quantitative PCR and western blotting. Drug resistance was measured by clonogenic and cell viability assays and effects on the cell cycle by cell sorting. Damage signalling was additionally investigated using immunostaining. RESULTS: MLH1-depleted cells were more resistant to MNU, as expected. Despite having an intact G(2)/M checkpoint, the WT cells did not initially undergo cell cycle arrest but instead triggered cell death directly by PARP overactivation and nuclear translocation of apoptosis-inducing factor (AIF). The MLH1-depleted cells showed defects in this pathway, with decreased staining for phosphorylated H2AX, altered PARP activity and reduced AIF translocation. Inhibitors of PARP, but not of caspases, blocked AIF translocation and greatly decreased short-term cell death in both WT and MLH1-depleted cells. This MLH1-dependent response to MNU was not blocked by inhibitors of ATM/ATR or p53. CONCLUSION: These novel data indicate an important role for MLH1 in signalling PARP-dependent cell death in response to the methylating agent MNU. British Journal of Cancer (2009) 101, 441-451. doi:10.1038/sj.bjc.6605186 www.bjcancer.com Published online 28 July 2009 (C) 2009 Cancer Research UK

Item Type:Journal article
Faculties and Schools:Faculty of Life and Health Sciences
Faculty of Life and Health Sciences > School of Biomedical Sciences
Research Institutes and Groups:Biomedical Sciences Research Institute
Biomedical Sciences Research Institute > Molecular Medicine
Biomedical Sciences Research Institute > Molecular Medicine > Nano Systems Biology
Biomedical Sciences Research Institute > Molecular Medicine > Transcriptional Regulation & Epigenetics
ID Code:3374
Deposited By:Professor Stephen Downes
Deposited On:15 Dec 2009 11:43
Last Modified:11 Dec 2012 14:19

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