Deficiency of the Fanconi anemia E2 ubiqitin conjugase UBE2T only partially abrogates Alu-mediated recombination in a new model of homology dependent recombination

Lewis, Todd; Barthelemy, Joanna; Virts, Elizabeth L.; Kennedy, Felicia M.; Gadgil, Rujuta Yashodhan; Wiek, Constanze; Linka, René Martin; Zhang, Feng; Andreassen, Paul R.; Leffak, Michael

doi:10.1093/nar/gkz026

Cited by 13 publications

(11 citation statements)

References 129 publications

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“…“Cellular response to DNA damage stimulus” and “double-strand break repair” were among the mostly enriched ontology classes in both I-BET762- and I-BET762+TLZ-associated DEPs ( Figure 5 C). Among DEPs that are involved with DNA damage response, CHEK2, FANCC, NBN, OTUB1, UBE2B, and UBE2T have been reported to promote the HR-mediated DSBR process and they were down-regulated in both treatment groups [ 56 , 62 , 63 , 64 , 65 , 66 , 67 , 68 ] ( Figure S8 ). Furthermore, deficiencies in CHEK2, FANCC, or NBN have been shown to induce PARPi sensitivity in other tumor models [ 56 , 62 , 63 , 64 , 65 , 66 ].…”

Section: Resultsmentioning

confidence: 99%

BET-Inhibitor I-BET762 and PARP-Inhibitor Talazoparib Synergy in Small Cell Lung Cancer Cells

Fiorentino

Marchesi²,

Schröder³

et al. 2020

IJMS

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Small cell lung cancer (SCLC) is an aggressive type of lung cancer with high mortality that is caused by frequent relapses and acquired resistance. Despite that several target-based approaches with potential therapeutic impact on SCLC have been identified, numerous targeted drugs have not been successful in providing improvements in cancer patients when used as single agents. A combination of targeted therapies could be a strategy to induce maximum lethal effects on cancer cells. As a starting point in the development of new drug combination strategies for the treatment of SCLC, we performed a mid-throughput screening assay by treating a panel of SCLC cell lines with BETi or AKi in combination with PARPi or EZH2i. We observed drug synergy between I-BET762 and Talazoparib, BETi and PARPi, respectively, in SCLC cells. Combinatorial efficacy was observed in MYCs-amplified and MYCs-wt SCLC cells over SCLC cells with impaired MYC signaling pathway or non-tumor cells. We indicate that drug synergy between I-BET762 and Talazoparib is associated with the attenuation HR-DSBR process and the downregulation of various players of DNA damage response by BET inhibition, such as CHEK2, PTEN, NBN, and FANCC. Our results provide a rationale for the development of new combinatorial strategies for the treatment of SCLC.

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Section: Resultsmentioning

confidence: 99%

BET-Inhibitor I-BET762 and PARP-Inhibitor Talazoparib Synergy in Small Cell Lung Cancer Cells

Fiorentino

Marchesi²,

Schröder³

et al. 2020

IJMS

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“…In previous work in which the position of the I-Sce1 site was changed, we showed that homology directed repair that removes either the ectopic dTomato gene or the eGFP gene is 5 not inherently deleterious to cells (68). Therefore, the loss of the dTomato gene raised the possibility that the replication-dependent DSBs which were resistant to recombination and deleted all or part of chromosome 18 containing the dTomato gene were also inimical to cell survival.…”

Section: (Ctg/cag)n Repeat Length-dependent Dsbsmentioning

confidence: 99%

“…To identify factors affecting microsatellite DSBs and repair we developed a system in which a DSB between two chromosomal reporter genes could be detected by microscopy or flow cytometry ( Figure 1). In this system, (CTG/CAG)100 or (Pu/Py)88 microsatellites were individually integrated at a single copy FLP recombinase target (FRT) site at chromosome 18p11.22 in HeLa cells (64), bordered by the c-myc replication origin core (65)(66)(67), an I-Sce1 site, and two fluorescent protein marker genes (dTomato, eGFP) flanked by three identical AluYa5 elements ( Figure 1A) (68). Control cell lines were also constructed that contain the same starting construct except that the DF/myc cell line is missing the microsatellite sequences ( Figure 1B), and the DF cell line is additionally missing the c-myc origin core ( Figure 1C).…”

Section: A Dual Fluorescent Reporter System For Analysis Of Dna Dsbs mentioning

confidence: 99%

“…When these cells were transfected with the I-Sce1 expression vector, more than 40% of the cells lost the green reporter (generating red cells) or both reporters (resulting in double negative cells) by 4 days after treatment ( Figure 2C). The loss of the eGFP reporter gene is the result of intrachromosomal single-strand annealing between the second and third Alu elements, while the double negative cells result from SSA between the first and third Alu elements (68).…”

Section: Replication-dependent Dsbs Are Not Repaired In the Same Way mentioning

confidence: 99%

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Replication stress at microsatellites causes DNA double-strand breaks and break-induced replication

Gadgil

Romer

Goodman

et al. 2020

Journal of Biological Chemistry

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Short tandemly repeated DNA sequences, termed microsatellites, are abundant in the human genome. These microsatellites exhibit length instability and susceptibility to DNA double strand breaks (DSBs) due to their tendency to form stable non-B DNA structures. Replication-dependent microsatellite DSBs are linked to genome instability signatures in human developmental diseases and cancers. To probe the causes and consequences of microsatellite DSBs, we designed a dual fluorescence reporter system to detect DSBs at expanded (CTG/CAG)n and polypurine/polypyrimidine (Pu/Py) mirror repeat structures alongside the c-myc replication origin integrated at a single ectopic chromosomal site. Restriction cleavage near the (CTG/CAG)100 microsatellite leads to homology directed single strand annealing between flanking AluY elements, and reporter gene deletion that can be detected by flow cytometry. However, in the absence of restriction cleavage, endogenous and exogenous replication stressors induce DSBs at the (CTG/CAG)100 and Pu/Py microsatellites. DSBs map to a narrow region at the downstream edge of the (CTG)100 lagging strand template. (CTG/CAG)n chromosome fragility is repeat length dependent, while instability at the (Pu/Py) microsatellites depends on replication polarity. Strikingly, restriction-generated DSBs and replication-dependent DSBs are not repaired by the same mechanism. Knockdown of DNA damage response proteins increase (Rad18, Pol η, Pol κ) or decrease (Mus81) the sensitivity of the (CTG/CAG)100 microsatellites to replication stress. Replication stress and DSBs at the ectopic (CTG/CAG)100 microsatellite lead to break induced replication and high frequency mutagenesis at a flanking thymidine kinase gene. Our results show that non-B structure-prone microsatellites are susceptible to replication-dependent DSBs that cause genome instability.

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“…Ubiquitin signaling is a fundamental eukaryotic regulatory system, controlling diverse cellular functions, and once there are mutations or impairment of the E2 genes, severe disease states can occur [ 21 ]. Various studies reported that UBE2T influences tumors through the Fanconi anemia pathway [ 25 , 26 ].…”

Section: Discussionmentioning

confidence: 99%

Identification of Biomarkers Based on Bioinformatics Analysis: The Expression of Ubiquitin-Conjugating Enzyme E2T (UBE2T) in the Carcinogenesis and Progression of Hepatocellular Carcinoma

et al. 2020

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Background The purpose of this study was to screen and identify key genes in the occurrence and development of hepatocellular carcinoma (HCC) based on bioinformatics analysis. Material/Methods Three Gene Expression Omnibus (GEO) series (GSE) – GSE121248, GSE87630, and GSE84598 – were downloaded from the GEO database. GEO2R was used to screen different genes and a Venn diagram was drawn to screen coexpressed differentially expressed genes (DEGs). Coexpressed DEGs were obtained by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis, a protein-protein interaction network diagram was produced by Cytoscape, and module genes were calculated by the Molecular Complex Detection Cytoscape plug-in. Finally, overall survival, progression-free survival, and relapse-free survival analysis of the key genes selected were performed using the online Kaplan-Meier plotter. For the target genes, the online network UCSC Cancer Genome Browser was used to analyze the gene expression profiles of the grade and vascular invasion of HCC. Results A total of 296 coexpressed DEGs were obtained from the 3 GSEs and 12 key genes were obtained from the modular analysis. Survival analysis showed that the upregulated genes UBE2T and FBLN5 were involved in the poor prognosis of HCC. Furthermore, the expression of UBE2T was significantly related to the grade and vascular invasion of HCC. Conclusions The expression of the UBE2T gene was significantly upregulated in HCC tissue compared to in normal liver tissue. UBE2T may be a new marker for the diagnosis and subsequent therapy of HCC.

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Deficiency of the Fanconi anemia E2 ubiqitin conjugase UBE2T only partially abrogates Alu-mediated recombination in a new model of homology dependent recombination

Cited by 13 publications

References 129 publications

BET-Inhibitor I-BET762 and PARP-Inhibitor Talazoparib Synergy in Small Cell Lung Cancer Cells

BET-Inhibitor I-BET762 and PARP-Inhibitor Talazoparib Synergy in Small Cell Lung Cancer Cells

Replication stress at microsatellites causes DNA double-strand breaks and break-induced replication

Identification of Biomarkers Based on Bioinformatics Analysis: The Expression of Ubiquitin-Conjugating Enzyme E2T (UBE2T) in the Carcinogenesis and Progression of Hepatocellular Carcinoma

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