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Parp1 activation in mouse embryonic fibroblasts promotes Pol beta-dependent cellular hypersensitivity to alkylation damage.

ABSTRACT: Alkylating agents induce cell death in wild-type (WT) mouse embryonic fibroblasts (MEFs) by multiple mechanisms, including apoptosis, autophagy and necrosis. DNA polymerase beta (Pol beta) knockout (KO) MEFs are hypersensitive to the cytotoxic effect of alkylating agents, as compared to WT MEFs. To test the hypothesis that Parp1 is preferentially activated by methyl methanesulfonate (MMS) exposure of Pol beta KO MEFs, we have examined the relationship between Pol beta expression, Parp1 activation and cell survival following MMS exposure in a series of WT and Pol beta deficient MEF cell lines. Consistent with our hypothesis, we observed elevated Parp1 activation in Pol beta KO MEFs as compared to matched WT MEFs. Both the MMS-induced activation of Parp1 and the MMS-induced cytotoxicity of Pol beta KO MEFs are attenuated by pre-treatment with the Parp1/Parp2 inhibitor PJ34. Further, elevated Parp1 activation is observed following knockdown (KD) of endogenous Pol beta, as compared to WT cells. Pol beta KD MEFs are hypersensitive to MMS and both the MMS-induced hypersensitivity and Parp1 activation is prevented by pre-treatment with PJ34. In addition, the MMS-induced cellular sensitivity of Pol beta KO MEFs is reversed when Parp1 is also deleted (Pol beta/Parp1 double KO MEFs) and we observe no MMS sensitivity differential between Pol beta/Parp1 double KO MEFs and those that express recombinant mouse Pol beta. These studies suggest that Parp1 may function as a sensor of BER to initiate cell death when BER is aborted or fails. Parp1 may therefore function in BER as a tumor suppressor by initiating cell death and preventing the accumulation of cells with chromosomal damage due to a BER defect.

SUBMITTER: Jelezcova E

PROVIDER: S-EPMC2834876 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Parp1 activation in mouse embryonic fibroblasts promotes Pol beta-dependent cellular hypersensitivity to alkylation damage.

Jelezcova Elena E Trivedi Ram N RN Wang Xiao-Hong XH Tang Jiang-Bo JB Brown Ashley R AR Goellner Eva M EM Schamus Sandy S Fornsaglio Jamie L JL Sobol Robert W RW

Mutation research 20100122 1-2

Alkylating agents induce cell death in wild-type (WT) mouse embryonic fibroblasts (MEFs) by multiple mechanisms, including apoptosis, autophagy and necrosis. DNA polymerase beta (Pol beta) knockout (KO) MEFs are hypersensitive to the cytotoxic effect of alkylating agents, as compared to WT MEFs. To test the hypothesis that Parp1 is preferentially activated by methyl methanesulfonate (MMS) exposure of Pol beta KO MEFs, we have examined the relationship between Pol beta expression, Parp1 activatio ...[more]

PMID: 20096707

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Project description:ATP synthase inhibitory factor subunit 1 (IF1) is an inhibitory subunit of mitochondrial ATP synthase, playing a crucial role in regulating mitochondrial respiration and energetics. It is well-established that IF1 interacts with the F1 sector of ATP synthase to inhibit the reversal rotation and, thus, ATP hydrolysis. Recent evidence supports that IF1 also inhibits forward rotation or the ATP synthesis activity. Adding to the complexity, IF1 may also facilitate mitophagy and cristae formation. The implications of these complex actions of IF1 for cellular function remain obscure. In the present study, we found that IF1 expression was markedly upregulated in hypoxic MEFs relative to normoxic MEFs. We investigate how IF1 affects cellular growth and function in cultured mouse embryonic fibroblasts derived from mouse lines with systemic IF1 overexpression and knockout under normoxia and hypoxia. Cell survival and proliferation analyses revealed that IF1 overexpression exerted limited effects on cellular viability but substantially increased proliferation under normoxia, whereas it facilitated both cellular viability and proliferation under hypoxia. The absence of IF1 may have a pro-survival effect but not a proliferative one in both normoxia and hypoxia. Cellular bioenergetic analyses revealed that IF1 suppressed cellular respiration when subjected to normoxia and was even more pronounced when subjected to hypoxia with increased mitochondrial ATP production. In contrast, IF1 knockout MEFs showed markedly increased cellular respiration under both normoxia and hypoxia with little change in mitochondrial ATP. Glycolytic stress assay revealed that IF1 overexpression modestly increased glycolysis in normoxia and hypoxia. Interestingly, the absence of IF1 in MEFs led to substantial increases in glycolysis. Therefore, we conclude that IF1 mainly inhibits cellular respiration and enhances cellular glycolysis to preserve mitochondrial ATP. On the other hand, IF1 deletion can significantly facilitate cellular respiration and glycolysis without leading to mitochondrial ATP deficit.

Dataset Information

Parp1 activation in mouse embryonic fibroblasts promotes Pol beta-dependent cellular hypersensitivity to alkylation damage.

Publications

Parp1 activation in mouse embryonic fibroblasts promotes Pol beta-dependent cellular hypersensitivity to alkylation damage.

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