scholarly journals The nuclear RNAi factor, NRDE2, prevents the accumulation of DNA damage during mitosis in stressful growth conditions

2018 ◽  
Author(s):  
Aarati Asundi ◽  
Srivats Venkataramanan ◽  
Gina Caldas Cuellar ◽  
Atsushi Suzuki ◽  
Stephen N. Floor ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Mammalian Cells ◽  
Germ Line ◽  
Repair Process ◽  
Growth Conditions ◽  
Filamentous Actin ◽  
Heterochromatin Formation ◽  
C Elegans ◽  
Increased Risk

AbstractOrganisms have evolved multiple mechanisms to prevent and repair DNA damage to protect the integrity of the genome, particularly under stressful conditions. Unrepaired DNA damage leads to genomic instability, aneuploidy, and an increased risk for cancer. Before the cell can divide, it must repair damaged DNA and it is thought that this process requires global silencing of most transcription. In C. elegans, NRDE-2, in complex with other nuclear factors and guided by small RNA, directs heterochromatin formation and transcriptional silencing of targeted genes. Additionally, when C. elegans are cultivated at high temperatures, NRDE-2 is required to maintain germ line immortality. However, the role of NRDE-2 in maintaining the physical integrity of the genome is not understood. We show here that loss of NRDE2 in either nematode or human cells induces the accumulation of DNA damage specifically under conditions of stress, such as cultivation at a high temperature in C. elegans or Aurora B Kinase oncogenic overexpression in the MCF10A epithelial breast cell line. In addition, we found that NRDE2 interacts with β-actin in unstressed mammalian cells. This interaction is dramatically reduced upon DNA damage. Monomeric nuclear actin binds to heterochromatin remodeling factors and transcriptional activators while filamentous actin has been implicated in DNA repair processes. Thus, NRDE2 may dissociate from actin when it becomes filamentous as a result of DNA damage. In this way, heterochromatin factors may associate with the actin dependent DNA repair process to allow appropriate mitotic progression and maintain genomic integrity.

scholarly journals ANKLE1 as New Hotspot Mutation for Breast Cancer in Indian Population and Has a Role in DNA Damage and Repair in Mammalian Cells

2021 ◽  
Vol 11 ◽  
Author(s):  
Divya Bakshi ◽  
Archana Katoch ◽  
Souneek Chakraborty ◽  
Ruchi Shah ◽  
Bhanu Sharma ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Dna Damage ◽  
Dna Repair ◽  
Mammalian Cells ◽  
Nuclear Architecture ◽  
Cancer Cell Line ◽  
Repair Process ◽  
Dna Damage And Repair ◽  
Women In India

Breast cancer has replaced cervical cancer as being the most common and having the highest mortality among women in India. ANKLE gene is conserved among organisms during evolutionary succession and is a member of LEM family proteins in lower metazoans and is involved in critical functions in the nuclear architecture, gene expression and cell signaling. ANKLE1 is the human orthologous of LEM-3 and is involved in DNA damage response and DNA repair. Whole Exome Sequencing (WES) of paired breast cancer samples was performed and ANKLE1 was found to be a new possible hotspot for predisposition of breast cancer. The mass array genotyping for breast cancer variant rs2363956 further confirmed the ANKLE1 association with the studied population of breast cancer. To elucidate the role of ANKLE1 in DNA damage, it was knocked down in MCF-7 breast cancer cell line and the expression of γH2AX was assessed. ANKLE1 knockdown cells displayed elevated levels of γ-H2AX foci in response to the cisplatin induced replication stress. The localization pattern of ANKLE1 further emphasized the role of ANKLE1 in DNA repair process. We observed that ANKLE1 is required for maintaining genomic stability and plays a role in DNA damage and repair process. These findings provided a molecular basis for the suspected role of ANKLE1 in human breast cancer and suggested an important role of this gene in controlling breast cancer development among women in India.

Expression of the DNA Repair Gene RAD51 Is Associated with FLT3 ITD Transcript Level and Is Down-Regulated by PKC412 Resulting in Suppression of DNA Repair.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 283-283
Author(s):  
Claire H. Seedhouse ◽  
Hannah M. Hunter ◽  
Ian Carter ◽  
Anne-Marie Massip ◽  
Monica Pallis ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Cell Line ◽  
Mammalian Cells ◽  
Transcript Level ◽  
Hl60 Cell ◽  
Prognostic Impact ◽  
Dna Repair Gene ◽  
Increased Risk ◽  
Hl60 Cell Line

Abstract The presence of internal tandem duplication (ITD) mutations in the FLT3 receptor tyrosine kinase confer an adverse prognosis in AML due to an increased risk of disease relapse. However the mechanisms underlying this increased relapse risk are unclear. We have investigated whether AML cells with FLT3 ITD mutations have an enhanced capacity for DNA repair following cytotoxic drug exposure. RAD51 is a key protein in the high-fidelity homologous recombination double strand break repair pathway and is the limiting factor for this pathway in mammalian cells. Using quantitative real-time PCR we found that the level of RAD51 transcripts are significantly correlated with the level of FLT3 transcripts in FLT3 ITD cells (n=27; p=0.017) but not in FLT3 WT cells (n=57; p=0.58). Clinically FLT3 ITDs have the most significant prognostic impact in AML patients with normal cytogenetics and if this group is studied the association between FLT3 ITD and RAD51 transcript levels are particularly pronounced (n=12; p=0.003). To establish whether increases in RAD51 expression correlate with enhanced DNA repair activity we have adopted the comet assay and studied the MV4-11 cell line (FLT3 ITD), HL60 cell line (FLT3 WT) and a number of AML patient cells. The background level of DNA damage in untreated FLT3 ITD AML patients and the MV4-11 cell line was significantly lower than in FLT3 WT patients and the HL60 cell line (n=10; p=0.02), suggesting a constitutive up-regulation of DNA repair in cells harbouring the FLT3 ITD mutation resulting in lower background levels of DNA damage. To test whether the increase in RAD51 and DNA repair was a consequence of the FLT3 ITD, we treated cells with the FLT3 inhibitor PKC412 and then examined the response of the cells to sub-toxic doses of daunorubicin. The MV4-11 FLT3 ITD cells, but not the FLT3 WT HL60 cells, demonstrated a statistically significant suppression of early DNA repair when treated with PKC412 and daunorubicin (p<0.001). Similar results were obtained in primary AML cells, with a loss of early DNA repair in the FLT3 ITD cells and no effect on the WT FLT3 cells. Furthermore the reduction in daunorubicin-induced DNA repair seen in PKC412 treated FLT3 ITD cells was associated with down-regulation of RAD51 expression. There was a statistically significant decrease in RAD51 transcript level following PKC412 treatment in the FLT3 ITD patients and MV4-11 cell line but not in the FLT3 WT patients or HL60 cell line (p<0.05). The decrease in the expression of RAD51 is closely associated with the reduction in DNA repair function in the inhibitor treated FLT3 ITD cells. This work suggests that high expression levels of FLT3 transcripts in AML cells with a FLT3 ITD up-regulate RAD51 resulting in more efficient DNA repair following chemotherapy treatment. This may lead to resistance to cytotoxic therapies and genomic instability, ultimately resulting in the manifestation of a more resistant disease and a greater likelihood of relapse. The use of FLT3 inhibitors concurrently with or following AML therapy may suppress enhanced DNA repair in FLT3 mutated cells.

AMPK blocks chromatin activation and consequent R-loop formation to protect genome stability following acute starvation

2021 ◽  
Author(s):  
Bing Sun ◽  
McLean Sherrin ◽  
Richard Roy
Keyword(s):  
Dna Damage ◽  
Genome Stability ◽  
Germ Line ◽  
Critical Role ◽  
Significant Proportion ◽  
Loop Formation ◽  
C Elegans ◽  
Chromatin Activation ◽  
Acute Starvation

Abstract During periods of starvation organisms must modify both gene expression and metabolic pathways to adjust to the energy stress. We previously reported that C. elegans that lack AMPK have transgenerational reproductive defects that result from abnormally elevated H3K4me3 levels in the germ line following recovery from acute starvation1. Here we show that H3K4me3 is dramatically increased at promoters, driving aberrant transcription elongation that results in the accumulation of R-loops in the starved AMPK mutants. DRIP-seq analysis demonstrated that a significant proportion of the genome was affected by R-loop formation with a dramatic expansion in the number of R-loops at numerous loci, most pronounced at the promoter-TSS regions of genes in the starved AMPK mutants. The R-loops are transmissible into subsequent generations, likely contributing to the transgenerational reproductive defects typical of these mutants following starvation. Strikingly, AMPK null germ lines show considerably more RAD-51 foci at sites of R-loop formation, potentially sequestering it from its critical role at meiotic breaks and/or at sites of induced DNA damage. Our study reveals a previously unforeseen role of AMPK in maintaining genome stability following starvation, where in its absence R-loops accumulate, resulting in reproductive compromise and DNA damage hypersensitivity.

scholarly journals The emerging role of RNAs in DNA damage repair

2017 ◽  
Vol 24 (4) ◽  
pp. 580-587 ◽  
Author(s):  
Ben R Hawley ◽  
Wei-Ting Lu ◽  
Ania Wilczynska ◽  
Martin Bushell
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Critical Role ◽  
Repair Process ◽  
Rna Molecules ◽  
Processing Enzymes ◽  
Damage Repair ◽  
New Findings ◽  
Integral Role ◽  
Repair Mechanisms

Abstract Many surveillance and repair mechanisms exist to maintain the integrity of our genome. All of the pathways described to date are controlled exclusively by proteins, which through their enzymatic activities identify breaks, propagate the damage signal, recruit further protein factors and ultimately resolve the break with little to no loss of genetic information. RNA is known to have an integral role in many cellular pathways, but, until very recently, was not considered to take part in the DNA repair process. Several reports demonstrated a conserved critical role for RNA-processing enzymes and RNA molecules in DNA repair, but the biogenesis of these damage-related RNAs and their mechanisms of action remain unknown. We will explore how these new findings challenge the idea of proteins being the sole participants in the response to DNA damage and reveal a new and exciting aspect of both DNA repair and RNA biology.

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

2020 ◽  
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.

scholarly journals A Review on DNA Repair Inhibition by PARP Inhibitors in Cancer Therapy

Folia Medica ◽  
2018 ◽  
Vol 60 (1) ◽  
pp. 39-47 ◽  
Author(s):  
Ashish P. Shah ◽  
Chhagan N. Patel ◽  
Dipen K. Sureja ◽  
Kirtan P. Sanghavi
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Cancer Therapy ◽  
Repair Process ◽  
Parp Inhibitors ◽  
Brca Mutations ◽  
Development Of Resistance ◽  
Damage Repair ◽  
Future Therapy

AbstractThe DNA repair process protects the cells from DNA damaging agent by multiple pathways. Majority of the cancer therapy cause DNA damage which leads to apoptosis. The cell has natural ability to repair this damage which ultimately leads to development of resistance of drugs. The key enzymes involved in DNA repair process are poly(ADP-ribose) (PAR) and poly(ADP-ribose) polymerases (PARP). Tumor cells repair their defective gene via defective homologues recombination (HR) in the presence of enzyme PARP. PARP inhibitors inhibit the enzyme poly(ADP-ribose) polymerases (PARPs) which lead to apoptosis of cancer cells. Current clinical data shows the role of PARP inhibitors is not restricted to BRCA mutations but also effective in HR dysfunctions related tumors. Therefore, investigation in this area could be very helpful for future therapy of cancer. This review gives detail information on the role of PARP in DNA damage repair, the role of PARP inhibitors and chemistry of currently available PARP inhibitors.

scholarly journals Advances and perspectives of PARP inhibitors

2019 ◽  
Vol 8 (1) ◽  
Author(s):  
Ming Yi ◽  
Bing Dong ◽  
Shuang Qin ◽  
Qian Chu ◽  
Kongming Wu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Cancer Cells ◽  
Genome Instability ◽  
Single Gene ◽  
High Sensitivity ◽  
Lethal Effect ◽  
Single Strand Break ◽  
Synthetic Lethal ◽  
Increased Risk

Abstract DNA damage repair deficiency leads to the increased risk of genome instability and oncogenic transformation. In the meanwhile, this deficiency could be exploited for cancer treatment by inducing excessive genome instability and catastrophic DNA damage. Continuous DNA replication in cancer cells leads to higher demand of DNA repair components. Due to the oncogenic loss of some DNA repair effectors (e.g. BRCA) and incomplete DNA repair repertoire, some cancer cells are addicted to certain DNA repair pathways such as Poly (ADP-ribose) polymerase (PARP)-related single-strand break repair pathway. The interaction between BRCA and PARP is a form of synthetic lethal effect which means the simultaneously functional loss of two genes lead to cell death, while defect in any single gene has a slight effect on cell viability. Based on synthetic lethal theory, Poly (ADP-ribose) polymerase inhibitor (PARPi) was developed aiming to selectively target cancer cells harboring BRCA1/2 mutations. Recently, a growing body of evidence indicated that a broader population of patients could benefit from PARPi therapy far beyond those with germline BRCA1/2 mutated tumors. Numerous biomarkers including homologous recombination deficiency and high level of replication pressure also herald high sensitivity to PARPi treatment. Besides, a series of studies indicated that PARPi-involved combination therapy such as PARPi with additional chemotherapy therapy, immune checkpoint inhibitor, as well as targeted agent had a great advantage in overcoming PARPi resistance and enhancing PARPi efficacy. In this review, we summarized the advances of PARPi in clinical application. Besides, we highlighted multiple promising PARPi-based combination strategies in preclinical and clinical studies.

scholarly journals Mutational signatures are jointly shaped by DNA damage and repair

2020 ◽  
Vol 11 (1) ◽  
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.

scholarly journals Caenorhabditis elegans Deficient in DOT-1.1 Exhibit Increases in H3K9me2 at Enhancer and Certain RNAi-Regulated Regions

Cells ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 1846 ◽  
Author(s):  
Ruben Esse ◽  
Alla Grishok
Keyword(s):  
Small Rnas ◽  
Mammalian Cells ◽  
Leukemic Cells ◽  
Rnai Pathway ◽  
Open Chromatin ◽  
Loss Of Function ◽  
Promoter Regions ◽  
Heterochromatin Formation ◽  
C Elegans ◽  
H3k79 Methylation

The methylation of histone H3 at lysine 79 is a feature of open chromatin. It is deposited by the conserved histone methyltransferase DOT1. Recently, DOT1 localization and H3K79 methylation (H3K79me) have been correlated with enhancers in C. elegans and mammalian cells. Since earlier research implicated H3K79me in preventing heterochromatin formation both in yeast and leukemic cells, we sought to inquire whether a H3K79me deficiency would lead to higher levels of heterochromatic histone modifications, specifically H3K9me2, at developmental enhancers in C. elegans. Therefore, we used H3K9me2 ChIP-seq to compare its abundance in control and dot-1.1 loss-of-function mutant worms, as well as in rde-4; dot-1.1 and rde-1; dot-1.1 double mutants. The rde-1 and rde-4 genes are components of the RNAi pathway in C. elegans, and RNAi is known to initiate H3K9 methylation in many organisms, including C. elegans. We have previously shown that dot-1.1(−) lethality is rescued by rde-1 and rde-4 loss-of-function. Here we found that H3K9me2 was elevated in enhancer, but not promoter, regions bound by the DOT-1.1/ZFP-1 complex in dot-1.1(−) worms. We also found increased H3K9me2 at genes targeted by the ALG-3/4-dependent small RNAs and repeat regions. Our results suggest that ectopic H3K9me2 in dot-1.1(−) could, in some cases, be induced by small RNAs.

Mutations in CREBBP and EP300 genes affect DNA repair of oxidative damage in Rubinstein-Taybi syndrome cells

2019 ◽  
Vol 41 (3) ◽  
pp. 257-266
Author(s):  
Ilaria Dutto ◽  
Claudia Scalera ◽  
Micol Tillhon ◽  
Giulio Ticli ◽  
Gianluca Passaniti ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Genome Stability ◽  
Oxidative Dna Damage ◽  
Excision Repair ◽  
Control Cell ◽  
Nuclear Antigen ◽  
Autosomal Dominant Disorder ◽  
Cell Extracts ◽  
Increased Risk

Abstract Rubinstein-Taybi syndrome (RSTS) is an autosomal-dominant disorder characterized by intellectual disability, skeletal abnormalities, growth deficiency and an increased risk of tumors. RSTS is predominantly caused by mutations in CREBBP or EP300 genes encoding for CBP and p300 proteins, two lysine acetyl-transferases (KAT) playing a key role in transcription, cell proliferation and DNA repair. However, the efficiency of these processes in RSTS cells is still largely unknown. Here, we have investigated whether pathways involved in the maintenance of genome stability are affected in lymphoblastoid cell lines (LCLs) obtained from RSTS patients with mutations in CREBBP or in EP300 genes. We report that RSTS LCLs with mutations affecting CBP or p300 protein levels or KAT activity, are more sensitive to oxidative DNA damage and exhibit defective base excision repair (BER). We have found reduced OGG1 DNA glycosylase activity in RSTS compared to control cell extracts, and concomitant lower OGG1 acetylation levels, thereby impairing the initiation of the BER process. In addition, we report reduced acetylation of other BER factors, such as DNA polymerase β and Proliferating Cell Nuclear Antigen (PCNA), together with acetylation of histone H3. We also show that complementation of CBP or p300 partially reversed RSTS cell sensitivity to DNA damage. These results disclose a mechanism of defective DNA repair as a source of genome instability in RSTS cells.

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