Mismatch Repair Proteins Are Activators of Toxic Responses to Chromium-DNA Damage

Peterson-Roth, Elizabeth; Reynolds, Mindy; Quievryn, George; Zhitkovich, Anatoly

doi:10.1128/mcb.25.9.3596-3607.2005

Cited by 114 publications

(120 citation statements)

References 63 publications

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“…Cells were either treated with PARP inhibitors or transfected with siRNA or protein expression vector for the specified length of time, and harvested and lysed as previously described (31). Western blotting was then performed as previously described (32).…”

Section: Resultsmentioning

confidence: 99%

Inhibition of poly(ADP-ribose) polymerase down-regulates BRCA1 and RAD51 in a pathway mediated by E2F4 and p130

Hegan¹,

Lü²,

Stachelek³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

186

111

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Inhibitors of poly(ADP-ribose) polymerase (PARP) are in clinical trials for cancer therapy, on the basis of the role of PARP in recruitment of base excision repair (BER) factors to sites of DNA damage. Here we show that PARP inhibition to block BER is toxic to hypoxic cancer cells, in which homology-dependent repair (HDR) is known to be down-regulated. However, we also report the unexpected finding that disruption of PARP, itself, either via chemical PARP inhibitors or siRNAs targeted to PARP-1, can inhibit HDR by suppressing expression of BRCA1 and RAD51, key factors in HDR of DNA breaks. Mechanistically, PARP inhibition was found to cause increased occupancy of the BRCA1 and RAD51 promoters by repressive E2F4/p130 complexes, a pathway prevented by expression of HPV E7, which disrupts p130 activity, or by siRNAs to knock down p130 expression. Functionally, disruption of p130 by E7 expression or by siRNA knockdown also reverses the cytotoxicity and radiosensitivity associated with PARP inhibition, suggesting that the down-regulation of BRCA1 and RAD51 is central to these effects. Direct measurement of HDR using a GFP-based assay demonstrates reduced HDR in cells treated with PARP inhibitors. This work identifies a mechanism by which PARP regulates DNA repair and suggests new strategies for combination cancer therapies.DNA repair | hypoxia P oly(ADP-ribose) polymerases (PARPs) comprise a family of enzymes that catalyze ADP ribosylation of a variety of cellular factors (1-4). PARP-1 is thought to play a key role in DNA repair, primarily by modifying chromatin factors at sites of DNA damage and thereby recruiting repair factors. Inhibitors of PARP have attracted interest for cancer therapy because cancer cells deficient in BRCA1 or BRCA2 due to inactivating mutations are sensitive to PARP inhibition (5-8). This has been attributed to the role of PARP in recruiting base excision repair (BER) factors that remove damaged bases and fix single-strand breaks (SSBs) (1). SSBs persisting into S-phase produce replication fork collapse, requiring BRCA1-and BRCA2-mediated homology-dependent repair (HDR) for resolution (5, 9, 10).In prior work, we found that hypoxia suppresses HDR in human cells via transcriptional down-regulation of BRCA1 and RAD51 (11-15). Hence, we hypothesized that cancer cells in hypoxia, with acquired deficiency in HDR, might have increased sensitivity to PARP inhibition. Work presented here confirms this hypothesis, showing that PARP inhibitors are more cytotoxic to hypoxic than to normoxic cells. Because hypoxia causes BRCA1 and RAD51 down-regulation by stimulating E2F4/p130 occupancy of the BRCA1 and RAD51 promoters, we asked whether disruption of p130 function via expression of human papillomavirus (HPV) E7 would reverse the sensitivity of hypoxic cells to PARP inhibition. We found that E7 expression, as predicted, does confer resistance to PARP inhibitors on hypoxic cells, but surprisingly, it also blocks the toxicity of PARP inhibition in normoxic cells.As a basis for this effect, we present eviden...

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Section: Resultsmentioning

confidence: 99%

Inhibition of poly(ADP-ribose) polymerase down-regulates BRCA1 and RAD51 in a pathway mediated by E2F4 and p130

Hegan¹,

Lü²,

Stachelek³

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

186

111

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“…It is known that MMR proteins are required for both cell cycle arrest and induction of apoptosis by DNA damage (59,60). Although the specific mechanisms have yet to be identified, one hypothesis is that MMR proteins recognize misincorporated nucleotides or damaged base sites as mismatches and during attempts to repair them DNA breaks and gaps might be continuously produced and consequently cell cycle arrest and apoptosis are activated.…”

Section: Discussionmentioning

confidence: 99%

Postreplicative Mismatch Repair Factors Are Recruited to Epstein-Barr Virus Replication Compartments

Daikoku

Kudoh

Sugaya

et al. 2006

Journal of Biological Chemistry

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The mismatch repair (MMR) system, highly conserved throughout evolution, corrects nucleotide mispairing that arise during cellular DNA replication. We report here that proliferating cell nuclear antigen (PCNA), the clamp loader complex (RF-C), and a series of MMR proteins like MSH-2, MSH-6, MLH1, and hPSM2 can be assembled to Epstein-Barr virus replication compartments, the sites of viral DNA synthesis. Levels of the DNA-bound form of PCNA increased with progression of viral productive replication. Bromodeoxyuridine-labeled chromatin immunodepletion analyses confirmed that PCNA is loaded onto newly synthesized viral DNA as well as BALF2 and BMRF1 viral proteins during lytic replication. Furthermore, the anti-PCNA, -MSH2, -MSH3, or -MSH6 antibodies could immunoprecipitate BMRF1 replication protein probably via the viral DNA genome. PCNA loading might trigger transfer of a series of host MMR proteins to the sites of viral DNA synthesis. The MMR factors might function for the repair of mismatches that arise during viral replication or act to inhibit recombination between moderately divergent (homologous) sequences. Mismatch repair (MMR)2 systems play a primary role in mutation avoidance by removing base-base and small insertion-deletion mismatches that arise during DNA replication (1). Prokaryotes and eukaryotes have evolved similar systems for repair (2) and in Escherichia coli MMR is initiated when MutS binds to mismatched DNA, possibly through its interaction with the ␤-clamp accessory protein that is required for processive DNA replication (3-6). MutL binds to MutS to form MutS-MutL-DNA complexes that stimulate MutH binding and cleavage of unmethylated DNA strands at GATC sequences, either 5Ј or 3Ј of recognized mismatches. Exonucleases then chew away at the DNA beyond the mismatch site so that highly accurate DNA polymerase III can correctly re-synthesize the strand.In eukaryotes, mismatch recognition is accomplished by MSH2 (MutS homolog 2) forming a heterodimer with either MSH3 or MSH6 to bind to distinct but overlapping spectra of mismatches (7). In both the yeast Saccharomyces cerevisiae and humans, the repair of base-base mismatches appears to be solely dependent on MSH2-MSH6, whereas both MSH2-MSH6 and MSH2-MSH3 can participate in the repair of small (1 to 12-nucleotide) loop insertions. Currently, it is thought that MSH heterodimer binding to a mismatch triggers ATP-dependent steps that allow interactions with MLH (MutL homolog) heterodimers composed of MLH1-Pms1 or MLH1-MLH3 (7,8). No MutH homolog, however, has been identified in eukaryotes, and the exact details of strand discrimination and error removal are not known, although in both yeast and humans a number of other proteins have been implicated in MMR, including proliferating cell nuclear antigen (PCNA), replication factor C (RF-C), and DNA polymerases ␦ and ⑀ (9 -16).PCNA was originally characterized as a DNA sliding clamp for replicative DNA polymerases, but subsequent studies have revealed its striking ability to interact with multiple partner...

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“…A recent paper demonstrated that this response requires a functional MMR system. 52 Cells lacking hMLH1 or hMSH6 show improved survival compared to wild type cells after exposure to Cr(VI). This is apparently due to DNA double strand breaks that are caused by functional MMR and trigger apoptosis.…”

Section: Regulation Of Mmrmentioning

confidence: 99%

Structure and function of the components of the human DNA mismatch repair system

Jascur

Boland

2006

Intl Journal of Cancer

103

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DNA mismatch repair (MMR) is one of the several enzyme systems involved in DNA homeostasis. DNA MMR is involved in the repair of specific types of errors that occur during new DNA synthesis; loss of this system leads to an accelerated accumulation of potential mutations, and predisposes to certain types of cancers. Germline mutations in some of the DNA MMR genes cause the hereditary cancer predisposition, Lynch syndrome. This review addresses advances in the biochemistry of DNA MMR and its relationship to carcinogenesis. ' 2006 Wiley-Liss, Inc.Key words: DNA mismatch repair; microsatellite instability; lynch syndrome; HNPCC; colorectal cancer; hMSH2; hMLH1; hMSH6; hPMS2; exonuclease IThe DNA mismatch repair (MMR) system was at one time in the exclusive domain of the microbial biochemist, but has been thrust into the mainstream of the cancer biologist because of its importance in carcinogenesis. Approximately 15% of colorectal cancers develop through mechanisms whereby this form of DNA maintenance is lost, leading to 100-to 1,000-fold increases in error rates during replication. This review of recent progress in DNA MMR research is intended for the tumor biologist who is interested in structural and functional details of this system. There have been several reviews of the clinical aspects of DNA MMR that emphasize the diagnostic uses of testing for microsatellite instability and the use of immunohistochemistry of the DNA MMR proteins in colorectal cancer. [1][2][3][4][5][6][7] This review will serve to complement those reviews, and will highlight some of the biochemical issues that underlie DNA MMR. Components of the DNA MMR systemThe DNA MMR system corrects DNA base pairing errors in newly replicated DNA. The primary DNA polymerase in eukaryotic S phase DNA replication, polymerase d, has 3 0 > 5 0 proofreading activity that corrects 99% of replication errors. Nevertheless, mispaired nucleotides are occasionally left behind, as are small insertion/deletion mutations that are prone to occur at repetitive sequences. Microsatellites are multiple tandem repeats in which the repetitive element consists of a short number of nucleotides (perhaps 6 or fewer, and for functional studies, usually 1-4), and these are particularly prone to slippage and inefficient proofreading by DNA polymerase. The MMR system is critical to correct these problems, and if inactivated, the resulting shortening of microsatellites is a telltale sign of the ensuing ''microsatellite instability'' (MSI) phenotype. MSI testing is most often performed on mononucleotide or dinucleotide repeats.A casual description of the MMR system would state that the MMR system is chiefly a sensory system that scans DNA, and when a nucleotide mispair is detected, removes the error and summons DNA polymerase to repeat the synthesis, and this time corrects the error. This works because even in repetitive sequences, the error rate of DNA polymerase is low, and so simply having a second chance usually fixes the problem.More formally, the MMR system is an excision/resynt...

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Mismatch Repair Proteins Are Activators of Toxic Responses to Chromium-DNA Damage

Cited by 114 publications

References 63 publications

Inhibition of poly(ADP-ribose) polymerase down-regulates BRCA1 and RAD51 in a pathway mediated by E2F4 and p130

Inhibition of poly(ADP-ribose) polymerase down-regulates BRCA1 and RAD51 in a pathway mediated by E2F4 and p130

Postreplicative Mismatch Repair Factors Are Recruited to Epstein-Barr Virus Replication Compartments

Structure and function of the components of the human DNA mismatch repair system

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