Chk2 Mediates Stabilization of the FoxM1 Transcription Factor To Stimulate Expression of DNA Repair Genes

ABSTRACT The forkhead box M1 (FoxM1) transcription factor regulates expression of cell cycle genes essential for DNA replication and mitosis during organ repair and cancer progression. Here, we demonstrate that FoxM1-deficient (−/−) mouse embryonic fibroblasts and osteosarcoma U2OS cells depleted in FoxM1 levels by small interfering RNA transfection display increased DNA breaks, as evidenced by immunofluorescence focus staining for phosphospecific histone H2AX. FoxM1-deficient cells also exhibit stimulation of p53 transcriptional activity, as evidenced by increased expression of the p21cip1 gene. FoxM1-deficient cells display reduced expression of the base excision repair factor X-ray cross-complementing group 1 (XRCC1) and breast cancer-associated gene 2 (BRCA2), the latter of which is involved in homologous recombination repair of DNA double-strand breaks. Furthermore, FoxM1 protein is phosphorylated by checkpoint kinase 2 (Chk2) in response to DNA damage. This phosphorylation of FoxM1 on serine residue 361 caused increased stability of the FoxM1 protein with corresponding increased transcription of XRCC1 and BRCA2 genes, both of which are required for repair of DNA damage. These results identify a novel role for FoxM1 in the transcriptional response during DNA damage/checkpoint signaling and show a novel mechanism by which Chk2 protein regulates expression of DNA repair enzymes.

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DNA Excision Repair and DNA Damage-Induced Apoptosis Are Linked to Poly(ADP-Ribosyl)ation but Have Different Requirements for p53

Molecular and Cellular Biology ◽

10.1128/mcb.20.18.6695-6703.2000 ◽

2000 ◽

Vol 20 (18) ◽

pp. 6695-6703 ◽

Cited By ~ 39

Author(s):

Ralph Beneke ◽

Christoph Geisen ◽

Branko Zevnik ◽

Thomas Bauch ◽

Wolfgang-Ulrich Müller ◽

...

Keyword(s):

Dna Damage ◽

Cell Death ◽

Dna Repair ◽

Zinc Finger Protein ◽

Excision Repair ◽

Dna Breaks ◽

In Vivo Models ◽

P53 Expression ◽

Dna Excision Repair ◽

Parp Activity

ABSTRACT Poly(ADP-ribose) polymerase (PARP) is a DNA binding zinc finger protein that catalyzes the transfer of ADP-ribose residues from NAD+ to itself and different chromatin constituents, forming branched ADP-ribose polymers. The enzymatic activity of PARP is induced upon DNA damage and the PARP protein is cleaved during apoptosis, which suggested a role of PARP in DNA repair and DNA damage-induced cell death. We have generated transgenic mice that lack PARP activity in thymocytes owing to the targeted expression of a dominant negative form of PARP. In the presence of single-strand DNA breaks, the absence of PARP activity correlated with a strongly increased rate of apoptosis compared to cells with intact PARP activity. We found that blockage of PARP activity leads to a drastic increase of p53 expression and activity after DNA damage and correlates with an accelerated onset of Bax expression. DNA repair is almost completely blocked in PARP-deficient thymocytes regardless of p53 status. We found the same increased susceptibility to apoptosis in PARP null mice, a similar inhibition of DNA repair kinetics, and the same upregulation of p53 in response to DNA damage. Thus, based on two different experimental in vivo models, we identify a direct, p53-independent, functional connection between poly(ADP-ribosyl)ation and the DNA excision repair machinery. Furthermore, we propose a p53-dependent link between PARP activity and DNA damage-induced cell death.

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XRCC1 Is Specifically Associated with Poly(ADP-Ribose) Polymerase and Negatively Regulates Its Activity following DNA Damage

Molecular and Cellular Biology ◽

10.1128/mcb.18.6.3563 ◽

1998 ◽

Vol 18 (6) ◽

pp. 3563-3571 ◽

Cited By ~ 606

Author(s):

Murielle Masson ◽

Claude Niedergang ◽

Valérie Schreiber ◽

Sylviane Muller ◽

Josiane Menissier-de Murcia ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Mammalian Cells ◽

Excision Repair ◽

Cell Cycle Checkpoint ◽

Dna Breaks ◽

Protective Function ◽

C Terminus ◽

Base Excision

ABSTRACT Poly(ADP-ribose) polymerase (PARP; EC 2.4.2.30 ) is a zinc-finger DNA-binding protein that detects and signals DNA strand breaks generated directly or indirectly by genotoxic agents. In response to these breaks, the immediate poly(ADP-ribosyl)ation of nuclear proteins involved in chromatin architecture and DNA metabolism converts DNA damage into intracellular signals that can activate DNA repair programs or cell death options. To have greater insight into the physiological function of this enzyme, we have used the two-hybrid system to find genes encoding proteins putatively interacting with PARP. We have identified a physical association between PARP and the base excision repair (BER) protein XRCC1 (X-ray repair cross-complementing 1) in theSaccharomyces cerevisiae system, which was further confirmed to exist in mammalian cells. XRCC1 interacts with PARP by its central region (amino acids 301 to 402), which contains a BRCT (BRCA1 C terminus) module, a widespread motif in DNA repair and DNA damage-responsive cell cycle checkpoint proteins. Overexpression of XRCC1 in Cos-7 or HeLa cells dramatically decreases PARP activity in vivo, reinforcing the potential protective function of PARP at DNA breaks. Given that XRCC1 is also associated with DNA ligase III via a second BRCT module and with DNA polymerase β, our results provide strong evidence that PARP is a member of a BER multiprotein complex involved in the detection of DNA interruptions and possibly in the recruitment of XRCC1 and its partners for efficient processing of these breaks in a coordinated manner. The modular organizations of these interactors, associated with small conserved domains, may contribute to increasing the efficiency of the overall pathway.

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Quantitative description of early transcriptional response of plant cells to radiation-induced DNA damage using a Poisson initiation event model

Faktori eksperimental'noi evolucii organizmiv ◽

10.7124/feeo.v26.1256 ◽

2020 ◽

Vol 26 ◽

pp. 139-143

Author(s):

S. V. Litvinov ◽

N. M. Rashydov

Keyword(s):

Mathematical Model ◽

Dna Damage ◽

Arabidopsis Thaliana ◽

Dna Repair ◽

Ionizing Radiation ◽

Plant Cells ◽

Transcriptional Response ◽

Dna Breaks ◽

Radiation Induced ◽

Early Transcriptional Response

Aim. One of the problems that have not lost their relevance is the study of the mechanisms of adaptation of higher plants to the effects of radiation associated with the modification of the DNA repair system in response to radiation. This paper presents a Poisson mathematical model of the radiation-induced early transcriptional response of genes of key enzymes, which catalyze recovery of double-stranded DNA breaks in active plant cells. Methods. We used total X-ray irradiation of a model object – 35-day-old Arabidopsis thaliana (L.) Heynh plants at sublethal doses of 3-21 Gy, total RNA extraction, reverse transcription with random hexanucleotide primers, PCR amplification of the obtained cDNA with primers to target genes, fluorescence gel densitometry of amplified products. Results. A mathematical model of transcriptional response to the genotoxic action of ionizing radiation in a subpopulation of active plant cells based on Poisson distribution, which satisfactorily describes the experimental data obtained, is proposed. Conclusions. To initiate a maximal transcriptional response to DNA damage, one two-strand lesion per chromosome, detected by DNA repair systems, is sufficient, while the absence of double-stranded lesions, or the appearance of more than one double-stranded lesion per chromosome inhibits early transcriptional response of the cell on the action of ionizing radiation. The Poisson model of the initiating event makes it possible to predict the response of subpopulations of active cells of angiosperms to the action of genotoxic factors. Keywords: ionizing radiation, DNA damage response (DDR), Arabidopsis thaliana, DNA repair, gene transcriptional activity.

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HPF1 remodels the active site of PARP1 to enable the serine ADP-ribosylation of histones

Nature Communications ◽

10.1038/s41467-021-21302-4 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Fa-Hui Sun ◽

Peng Zhao ◽

Nan Zhang ◽

Lu-Lu Kong ◽

Catherine C. L. Wong ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Active Site ◽

Negative Charge ◽

Structural Data ◽

Dna Breaks ◽

Adp Ribosylation ◽

Local Conformation ◽

Key Residues ◽

Structural Insights

AbstractUpon binding to DNA breaks, poly(ADP-ribose) polymerase 1 (PARP1) ADP-ribosylates itself and other factors to initiate DNA repair. Serine is the major residue for ADP-ribosylation upon DNA damage, which strictly depends on HPF1. Here, we report the crystal structures of human HPF1/PARP1-CAT ΔHD complex at 1.98 Å resolution, and mouse and human HPF1 at 1.71 Å and 1.57 Å resolution, respectively. Our structures and mutagenesis data confirm that the structural insights obtained in a recent HPF1/PARP2 study by Suskiewicz et al. apply to PARP1. Moreover, we quantitatively characterize the key residues necessary for HPF1/PARP1 binding. Our data show that through salt-bridging to Glu284/Asp286, Arg239 positions Glu284 to catalyze serine ADP-ribosylation, maintains the local conformation of HPF1 to limit PARP1 automodification, and facilitates HPF1/PARP1 binding by neutralizing the negative charge of Glu284. These findings, along with the high-resolution structural data, may facilitate drug discovery targeting PARP1.

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Mutational signatures are jointly shaped by DNA damage and repair

10.1101/686295 ◽

2019 ◽

Cited By ~ 2

Author(s):

Nadezda V Volkova ◽

Bettina Meier ◽

Víctor González-Huici ◽

Simone Bertolini ◽

Santiago Gonzalez ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Excision Repair ◽

Dna Lesions ◽

Single Nucleotide Variants ◽

Mutational Signatures ◽

Experimental Conditions ◽

Repair Deficiency ◽

Dna Repair Deficiency ◽

Mutation Spectra

AbstractMutations arise when DNA lesions escape DNA repair. To delineate the contributions of DNA damage and DNA repair deficiency to mutagenesis we sequenced 2,717 genomes of wild-type and 53 DNA repair defective C. elegans strains propagated through several generations or exposed to 11 genotoxins at multiple doses. Combining genotoxin exposure and DNA repair deficiency alters mutation rates or leads to unexpected mutation spectra in nearly 40% of all experimental conditions involving 9/11 of genotoxins tested and 32/53 genotypes. For 8/11 genotoxins, signatures change in response to more than one DNA repair deficiency, indicating that multiple genes and pathways are involved in repairing DNA lesions induced by one genotoxin. For many genotoxins, the majority of observed single nucleotide variants results from error-prone translesion synthesis, rather than primary mutagenicity of altered nucleotides. Nucleotide excision repair mends the vast majority of genotoxic lesions, preventing up to 99% of mutations. Analogous mutagenic DNA damage-repair interactions can also be found in cancers, but, except for rare cases, effects are weak owing to the unknown histories of genotoxic exposures and DNA repair status. Overall, our data underscore that mutation spectra are joint products of DNA damage and DNA repair and imply that mutational signatures computationally derived from cancer genomes are more variable than currently anticipated.

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DNA damage shifts circadian clock time via Hausp-dependent Cry1 stabilization

eLife ◽

10.7554/elife.04883 ◽

2015 ◽

Vol 4 ◽

Cited By ~ 45

Author(s):

Stephanie J Papp ◽

Anne-Laure Huber ◽

Sabine D Jordan ◽

Anna Kriebs ◽

Madelena Nguyen ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Circadian Clock ◽

Transcriptional Response ◽

Genotoxic Stress ◽

Genomic Integrity ◽

Clock Time ◽

Dna Repair Enzymes ◽

Repair Enzymes ◽

Specific Protease

The circadian transcriptional repressors cryptochrome 1 (Cry1) and 2 (Cry2) evolved from photolyases, bacterial light-activated DNA repair enzymes. In this study, we report that while they have lost DNA repair activity, Cry1/2 adapted to protect genomic integrity by responding to DNA damage through posttranslational modification and coordinating the downstream transcriptional response. We demonstrate that genotoxic stress stimulates Cry1 phosphorylation and its deubiquitination by Herpes virus associated ubiquitin-specific protease (Hausp, a.k.a Usp7), stabilizing Cry1 and shifting circadian clock time. DNA damage also increases Cry2 interaction with Fbxl3, destabilizing Cry2. Thus, genotoxic stress increases the Cry1/Cry2 ratio, suggesting distinct functions for Cry1 and Cry2 following DNA damage. Indeed, the transcriptional response to genotoxic stress is enhanced in Cry1−/− and blunted in Cry2−/− cells. Furthermore, Cry2−/− cells accumulate damaged DNA. These results suggest that Cry1 and Cry2, which evolved from DNA repair enzymes, protect genomic integrity via coordinated transcriptional regulation.

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Structural basis of TFIIH activation for nucleotide excision repair

10.1101/628032 ◽

2019 ◽

Author(s):

Goran Kokic ◽

Aleksandar Chernev ◽

Dimitry Tegunov ◽

Christian Dienemann ◽

Henning Urlaub ◽

...

Keyword(s):

Dna Repair ◽

Transcription Factor ◽

Nucleotide Excision Repair ◽

Excision Repair ◽

Dna Lesions ◽

Structural Basis ◽

Disease Mutations ◽

Mechanistic Analysis ◽

Nucleotide Excision ◽

Dna Complex

AbstractGenomes are constantly threatened by DNA damage, but cells can remove a large variety of DNA lesions by nucleotide excision repair (NER)1. Mutations in NER factors compromise cellular fitness and cause human diseases such as Xeroderma pigmentosum (XP), Cockayne syndrome and trichothiodystrophy2,3. The NER machinery is built around the multisubunit transcription factor IIH (TFIIH), which opens the DNA repair bubble, scans for the lesion, and coordinates excision of the damaged DNA single strand fragment1,4. TFIIH consists of a kinase module and a core module that contains the ATPases XPB and XPD5. Here we prepare recombinant human TFIIH and show that XPB and XPD are stimulated by the additional NER factors XPA and XPG, respectively. We then determine the cryo-electron microscopy structure of the human core TFIIH-XPA-DNA complex at 3.6 Å resolution. The structure represents the lesion-scanning intermediate on the NER pathway and rationalizes the distinct phenotypes of disease mutations. It reveals that XPB and XPD bind double- and single-stranded DNA, respectively, consistent with their translocase and helicase activities. XPA forms a bridge between XPB and XPD, and retains the DNA at the 5’-edge of the repair bubble. Biochemical data and comparisons with prior structures6,7 explain how XPA and XPG can switch TFIIH from a transcription factor to a DNA repair factor. During transcription, the kinase module inhibits the repair helicase XPD8. For DNA repair, XPA dramatically rearranges the core TFIIH structure, which reorients the ATPases, releases the kinase module and displaces a ‘plug’ element from the DNA-binding pore in XPD. This enables XPD to move by ~80 Å, engage with DNA, and scan for the lesion in a XPG-stimulated manner. Our results provide the basis for a detailed mechanistic analysis of the NER mechanism.

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Systematic analysis of mutational spectra associated with DNA repair deficiency in C. elegans

10.1101/2020.06.04.133306 ◽

2020 ◽

Cited By ~ 1

Author(s):

B Meier ◽

NV Volkova ◽

Y Hong ◽

S Bertolini ◽

V González-Huici ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Excision Repair ◽

Structural Variants ◽

Systematic Analysis ◽

C Elegans ◽

Mutational Spectra ◽

Base Substitutions ◽

Repair Pathways ◽

Complex Structural

AbstractGenome integrity is particularly important in germ cells to faithfully preserve genetic information across generations. As yet little is known about the contribution of various DNA repair pathways to prevent mutagenesis. Using the C. elegans model we analyse mutational spectra that arise in wild-type and 61 DNA repair and DNA damage response mutants cultivated over multiple generations. Overall, 44% of lines show >2-fold increased mutagenesis with a broad spectrum of mutational outcomes including changes in single or multiple types of base substitutions induced by defects in base excision or nucleotide excision repair, or elevated levels of 50-400 bp deletions in translesion polymerase mutants rev-3(pol ζ) and polh-1(pol η). Mutational signatures associated with defective homologous recombination fall into two classes: 1) mutants lacking brc-1/BRCA1 or rad-51/RAD51 paralogs show elevated base substitutions, indels and structural variants, while 2) deficiency for MUS-81/MUS81 and SLX-1/SLX1 nucleases, and HIM-6/BLM, HELQ-1/HELQ and RTEL-1/RTEL1 helicases primarily cause structural variants. Genome-wide investigation of mutagenesis patterns identified elevated rates of tandem duplications often associated with inverted repeats in helq-1 mutants, and a unique pattern of ‘translocation’ events involving homeologous sequences in rip-1 paralog mutants. atm-1/ATM DNA damage checkpoint mutants harboured complex structural variants enriched in subtelomeric regions, and chromosome end-to-end fusions. Finally, while inactivation of the p53-like gene cep-1 did not affect mutagenesis, combined brc-1 cep-1 deficiency displayed increased, locally clustered mutagenesis. In summary, we provide a global view of how DNA repair pathways prevent germ cell mutagenesis.

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Immune checkpoint inhibitor sensitivity of DNA repair deficient tumors

Immunotherapy ◽

10.2217/imt-2021-0024 ◽

2021 ◽

Vol 13 (14) ◽

pp. 1205-1213

Author(s):

Pauline Rochefort ◽

Françoise Desseigne ◽

Valérie Bonadona ◽

Sophie Dussart ◽

Clélia Coutzac ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Immune Checkpoint ◽

Genome Stability ◽

Immune Checkpoint Inhibitor ◽

Checkpoint Inhibitor ◽

Excision Repair ◽

Checkpoint Inhibitors ◽

Repair Deficiency ◽

Dna Repair Deficiency

Faithful DNA replication is necessary to maintain genome stability and implicates a complex network with several pathways depending on DNA damage type: homologous repair, nonhomologous end joining, base excision repair, nucleotide excision repair and mismatch repair. Alteration in components of DNA repair machinery led to DNA damage accumulation and potentially carcinogenesis. Preclinical data suggest sensitivity to immune checkpoint inhibitors in tumors with DNA repair deficiency. Here, we review clinical studies that explored the use of immune checkpoint inhibitor in patient harboring tumor with DNA repair deficiency.

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Mutational signatures are jointly shaped by DNA damage and repair

Nature Communications ◽

10.1038/s41467-020-15912-7 ◽

2020 ◽

Vol 11 (1) ◽

Cited By ~ 19

Author(s):

Nadezda V. Volkova ◽

Bettina Meier ◽

Víctor González-Huici ◽

Simone Bertolini ◽

Santiago Gonzalez ◽

...

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Excision Repair ◽

Point Mutations ◽

Mutational Signatures ◽

Dna Damage And Repair ◽

C Elegans ◽

Dna Repair Deficiency ◽

Dna Alterations ◽

Repair Pathways

AbstractCells possess an armamentarium of DNA repair pathways to counter DNA damage and prevent mutation. Here we use C. elegans whole genome sequencing to systematically quantify the contributions of these factors to mutational signatures. We analyse 2,717 genomes from wild-type and 53 DNA repair defective backgrounds, exposed to 11 genotoxins, including UV-B and ionizing radiation, alkylating compounds, aristolochic acid, aflatoxin B1, and cisplatin. Combined genotoxic exposure and DNA repair deficiency alters mutation rates or signatures in 41% of experiments, revealing how different DNA alterations induced by the same genotoxin are mended by separate repair pathways. Error-prone translesion synthesis causes the majority of genotoxin-induced base substitutions, but averts larger deletions. Nucleotide excision repair prevents up to 99% of point mutations, almost uniformly across the mutation spectrum. Our data show that mutational signatures are joint products of DNA damage and repair and suggest that multiple factors underlie signatures observed in cancer genomes.

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