scholarly journals FANCD2 Binds Human Papillomavirus Genomes and Associates with a Distinct Set of DNA Repair Proteins to Regulate Viral Replication

mBio ◽  
2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Chelsey C. Spriggs ◽  
Laimonis A. Laimins
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Human Papillomavirus ◽  
Life Cycle ◽  
High Risk ◽  
Dna Damage Response ◽  
Fanconi Anemia ◽  
Repair Pathway ◽  
Damage Response ◽  
Dna Repair Proteins

ABSTRACTThe life cycle of human papillomavirus (HPV) is dependent on the differentiation state of its host cell. HPV genomes are maintained as low-copy episomes in basal epithelial cells and amplified to thousands of copies per cell in differentiated layers. Replication of high-risk HPVs requires the activation of the ataxia telangiectasia-mutated (ATM) and ATM and Rad3-related (ATR) DNA repair pathways. The Fanconi anemia (FA) pathway is a part of the DNA damage response and mediates cross talk between the ATM and ATR pathways. Our studies show that HPV activates the FA pathway, leading to the accumulation of a key regulatory protein, FANCD2, in large nuclear foci. These HPV-dependent foci colocalize with a distinct population of DNA repair proteins, including ATM components γH2AX and BRCA1, but infrequently with p-SMC1, which is required for viral genome amplification in differentiated cells. Furthermore, FANCD2 is found at viral replication foci, where it is preferentially recruited to viral genomes compared to cellular chromosomes and is required for maintenance of HPV episomes in undifferentiated cells. These findings identify FANCD2 as an important regulator of HPV replication and provide insight into the role of the DNA damage response in the differentiation-dependent life cycle of HPV.IMPORTANCEHigh-risk human papillomaviruses (HPVs) are the etiological agents of cervical cancer and are linked to the development of many other anogenital and oropharyngeal cancers. Identification of host cellular pathways involved in regulating the viral life cycle may be helpful in identifying treatments for HPV lesions. Mutations in genes of the Fanconi anemia (FA) DNA repair pathway lead to genomic instability in patients and a predisposition to HPV-associated malignancies. Our studies demonstrate that FA pathway component FANCD2 is recruited to HPV DNA, associates with members of the ATM DNA repair pathway, and is essential for the maintenance of viral episomes in basal epithelial cells. Disruption of the FA pathway may result in increased integration events and a higher incidence of HPV-related cancer. Our study identifies new links between HPV and the FA pathway that may help to identify new therapeutic targets for the treatment of existing HPV infections and cancers.

scholarly journals Werner Helicase Control of Human Papillomavirus 16 E1-E2 DNA Replication Is Regulated by SIRT1 Deacetylation

mBio ◽  
2019 ◽  
Vol 10 (2) ◽  
Author(s):  
Dipon Das ◽  
Molly L. Bristol ◽  
Nathan W. Smith ◽  
Claire D. James ◽  
Xu Wang ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Life Cycle ◽  
Dna Replication ◽  
Homologous Recombination ◽  
Dna Damage Response ◽  
Human Papillomaviruses ◽  
Repair Protein ◽  
Damage Response ◽  
Dna Repair Protein

ABSTRACTHuman papillomaviruses (HPV) are double-stranded DNA viruses causative in a host of human diseases, including several cancers. Following infection, two viral proteins, E1 and E2, activate viral replication in association with cellular factors and stimulate the DNA damage response (DDR) during the replication process. E1-E2 uses homologous recombination (HR) to facilitate DNA replication, but an understanding of host factors involved in this process remains incomplete. Previously, we demonstrated that the class III deacetylase SIRT1, which can regulate HR, is recruited to E1-E2-replicating DNA and regulates the level of replication. Here, we demonstrate that SIRT1 promotes the fidelity of E1-E2 replication and that the absence of SIRT1 results in reduced recruitment of the DNA repair protein Werner helicase (WRN) to E1-E2-replicating DNA. CRISPR/Cas9 editing demonstrates that WRN, like SIRT1, regulates the quantity and fidelity of E1-E2 replication. This is the first report of WRN regulation of E1-E2 DNA replication, or a role for WRN in the HPV life cycle. In the absence of SIRT1 there is an increased acetylation and stability of WRN, but a reduced ability to interact with E1-E2-replicating DNA. We present a model in which E1-E2 replication turns on the DDR, stimulating SIRT1 deacetylation of WRN. This deacetylation promotes WRN interaction with E1-E2-replicating DNA to control the quantity and fidelity of replication. As well as offering a crucial insight into HPV replication control, this system offers a unique model for investigating the link between SIRT1 and WRN in controlling replication in mammalian cells.IMPORTANCEHPV16 is the major viral human carcinogen responsible for between 3 and 4% of all cancers worldwide. Following infection, this virus activates the DNA damage response (DDR) to promote its life cycle and recruits DDR proteins to its replicating DNA in order to facilitate homologous recombination during replication. This promotes the production of viable viral progeny. Our understanding of how HPV16 replication interacts with the DDR remains incomplete. Here, we demonstrate that the cellular deacetylase SIRT1, which is a part of the E1-E2 replication complex, regulates recruitment of the DNA repair protein WRN to the replicating DNA. We demonstrate that WRN regulates the level and fidelity of E1-E2 replication. Overall, the results suggest a mechanism by which SIRT1 deacetylation of WRN promotes its interaction with E1-E2-replicating DNA to control the levels and fidelity of that replication.

scholarly journals Aberrant DNA Damage Response and DNA Repair Pathway in High Glucose Conditions

2018 ◽  
Vol 7 (3) ◽  
pp. 64-74 ◽  
Author(s):  
Amy Zhong ◽  
Melissa Chang ◽  
Theresa Yu ◽  
Raymond Gau ◽  
Daniel J. Riley ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Dna Damage Response ◽  
High Glucose ◽  
Repair Pathway ◽  
Damage Response

scholarly journals Inhibition of USP28 overcomes Cisplatin-Resistance of Squamous Tumors by Suppression of the Fanconi Anemia Pathway

2020 ◽  
Author(s):  
Cristian Prieto-Garcia ◽  
Oliver Hartmann ◽  
Michaela Reissland ◽  
Thomas Fischer ◽  
Carina R. Maier ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Tumor Cells ◽  
Dna Damage Response ◽  
Fanconi Anemia ◽  
Cisplatin Resistance ◽  
Chemotherapy Resistance ◽  
Fanconi Anemia Pathway ◽  
Platinum Based Chemotherapy ◽  
Damage Response

AbstractSquamous cell carcinomas (SCC) frequently have a limited response to or develop resistance to platinum-based chemotherapy, and have an exceptionally high tumor mutational burden. As a consequence, overall survival is limited and novel therapeutic strategies are urgently required, especially in light of a rising incidences. SCC tumors express ΔNp63, a potent regulator of the Fanconi Anemia (FA) DNA-damage response pathway during chemotherapy, thereby directly contributing to chemotherapy-resistance. Here we report that the deubiquitylase USP28 affects the FA DNA repair pathway during cisplatin treatment in SCC, thereby influencing therapy outcome. In an ATR-dependent fashion, USP28 is phosphorylated and activated to positively regulate the DNA damage response. Inhibition of USP28 reduces recombinational repair via an ΔNp63-Fanconi Anemia pathway axis, and weakens the ability of tumor cells to accurately repair DNA. Our study presents a novel mechanism by which tumor cells, and in particular ΔNp63 expressing SCC, can be targeted to overcome chemotherapy resistance.SignificanceLimited treatment options and low response rates to chemotherapy are particularly common in patients with squamous cancer. The SCC specific transcription factor ΔNp63 enhances the expression of Fanconi Anemia genes, thereby contributing to recombinational DNA repair and Cisplatin resistance. Targeting the USP28-ΔNp63 axis in SCC tones down this DNA damage response pathways, thereby sensitizing SCC cells to cisplatin treatment.

ILF2-YB1 Protein Interaction Modulates RNA Splicing to Induce Resistance to Chemotherapy in High Risk Multiple Myeloma

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 359-359
Author(s):  
Matteo Marchesini ◽  
Yamini Ogoti ◽  
Elena Fiorini ◽  
Marianna D'anca ◽  
Paola Storti ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Dna Damage Response ◽  
Copy Number ◽  
Research Funding ◽  
Repair Pathway ◽  
Dna Breaks ◽  
Double Strand Dna Breaks ◽  
Damage Response ◽  
Cellular Replication

Abstract The 1q21 amplification, which occurs in approximately 40% of de novo and 70% of relapsed MM, is among the most frequent chromosomal aberrations in multiple myeloma (MM) patients and is considered a very high-risk genetic feature that is especially correlated with disease progression and drug resistance. To uncover novel 1q21 MM-critical genes, we first identified a list of 78 potential 1q21 drivers, which were located in the minimal common region of amplification of 254 MM samples and showed copy number-driven expression. These 78 candidates were then subjected to an shRNA screen to identify those genes involved in selective death and/or growth inhibition of MM cells carrying the 1q21 amplification. Using this approach, we identified and functionally validated the Interleukin-2 enhancer binding factor 2 (ILF2) as one of key 1q21 amplification-specific genes. ILF2 downregulation in 1q21-amplified MM cells resulted in multinucleated phenotypes and abnormal nuclear morphologies, findings that are consistent with the DNA damage-induced genomic instability that is associated with DNA repair defects that occur during cellular replication. Correspondingly, ILF2 downregulation was associated with a significant increase in the activation of the ATM (but not ATR or DNA-PK) pathway and accumulation of gH2AX foci, which are indicative of double-strand DNA breaks, and resulted in caspase 3-mediated apoptosis. Therefore, we sought to determine whether ILF2 is involved in the genome damage repair that occurs during cellular replication. To this end, we evaluated whether ILF2 depletion could affect the efficiency of non-homologous end joining (NHEJ) or homologous recombination (HR), the two major repair pathways in mammalian cells. We observed a profound impairment of HR in ILF2-depleted cells (p=0.038), whereas NHEJ was unaltered after ILF2 downregulation. Conversely, enforced ILF2 expression significantly enhanced HR efficiency in MM cells (p=0.008). To further support the role of ILF2 in the regulation of the DNA repair pathway in MM cells, we evaluated whether ILF2 downregulation increased MM sensitivity to DNA-damaging agents routinely used in the treatment of MM. Employing the interstrand crosslinker melphalan as an instigator of double-strand DNA breaks, we found that ILF2-depleted MM cells subjected to continuous melphalan treatment showed increased accumulation of γH2AX and apoptosis. Consistent with these findings, elevated ILF2 expression significantly correlated with poor survival in MM patients treated with high-dose melphalan followed by tandem autologous transplantation (n=256, p=0.01). Mechanistically, mass spectrometry analysis showed that ILF2 interacted with numerous RNA binding proteins directly involved in the regulation of DNA damage response by modulating alternative splicing of specific pre-mRNAs. RNA-sequencing experiments confirmed that ILF2 depletion resulted in aberrant splicing of genes involved in the DNA repair pathway, including ERCC1, FANCD2, and EXO1. RNA immunoprecipitation sequencing experiments showed that ILF2 directly bound to transcripts involved in the regulation of the HR pathway, including components of BRCA1 protein complex. Furthermore, in an attempt to dissect the ILF2 protein interacting network involved in the DNA repair regulation in response to DNA damage activation, we found that ILF2 mediated drug resistance in a dose-dependent manner by modulating YB-1 nuclear localization and interaction with the splicing factor U2AF65 to promote mRNA processing and stabilization of DNA repair genes, including FANCD2 and EXO1, in response to DNA damage. In conclusion, our study reveals an intimate relationship among 1q21 amplification, mRNA splicing, and DNA repair in the control of DNA damage response in MM. Given that 1q21 amplification is one of the most frequent copy number alterations in cancer, synthetic lethality approaches based on targeting gain-of-functions associated with ILF2 may have a broad spectrum of applications to potentiate the sensitivity of cancer cells to chemotherapeutic agents. Disclosures Giuliani: Janssen: Research Funding; Celgene: Research Funding.

scholarly journals Homologous Recombination Repair Factors Rad51 and BRCA1 Are Necessary for Productive Replication of Human Papillomavirus 31

2015 ◽  
Vol 90 (5) ◽  
pp. 2639-2652 ◽  
Author(s):  
William H. Chappell ◽  
Dipendra Gautam ◽  
Suzan T. Ok ◽  
Bryan A. Johnson ◽  
Daniel C. Anacker ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Human Papillomavirus ◽  
Homologous Recombination ◽  
Viral Replication ◽  
Dna Damage Response ◽  
Double Strand Breaks ◽  
Strand Breaks ◽  
Damage Response ◽  
Productive Replication

ABSTRACTHigh-risk human papillomavirus 31 (HPV31)-positive cells exhibit constitutive activation of the ATM-dependent DNA damage response (DDR), which is necessary for productive viral replication. In response to DNA double-strand breaks (DSBs), ATM activation leads to DNA repair through homologous recombination (HR), which requires the principal recombinase protein Rad51, as well as BRCA1. Previous studies from our lab demonstrated that Rad51 and BRCA1 are expressed at high levels in HPV31-positive cells and localize to sites of viral replication. These results suggest that HPV may utilize ATM activity to increase HR activity as a means to facilitate viral replication. In this study, we demonstrate that high-risk HPV E7 expression alone is sufficient for the increase in Rad51 and BRCA1 protein levels. We have found that this increase occurs, at least in part, at the level of transcription. Studies analyzing protein stability indicate that HPV may also protect Rad51 and BRCA1 from turnover, contributing to the overall increase in cellular levels. We also demonstrate that Rad51 is bound to HPV31 genomes, with binding increasing per viral genome upon productive replication. We have found that depletion of Rad51 and BRCA1, as well as inhibition of Rad51's recombinase activity, abrogates productive viral replication upon differentiation. Overall, these results indicate that Rad51 and BRCA1 are required for the process of HPV31 genome amplification and suggest that productive replication occurs in a manner dependent upon recombination.IMPORTANCEProductive replication of HPV31 requires activation of an ATM-dependent DNA damage response, though how ATM activity contributes to replication is unclear. Rad51 and BRCA1 play essential roles in repair of double-strand breaks, as well as the restart of stalled replication forks through homologous recombination (HR). Given that ATM activity is required to initiate HR repair, coupled with the requirement of Rad51 and BRCA1 for productive viral replication, our findings suggest that HPV may utilize ATM activity to ensure localization of recombination factors to productively replicating viral genomes. The finding that E7 increases the levels of Rad51 and BRCA1 suggests that E7 contributes to productive replication by providing DNA repair factors required for viral DNA synthesis. Our studies not only imply a role for recombination in the regulation of productive HPV replication but provide further insight into how HPV manipulates the DDR to facilitate the productive phase of the viral life cycle.

scholarly journals Beyond the Double-Strand Breaks: The Role of DNA Repair Proteins in Cancer Stem-Cell Regulation

Cancers ◽  
2021 ◽  
Vol 13 (19) ◽  
pp. 4818
Author(s):  
Jacqueline Nathansen ◽  
Felix Meyer ◽  
Luise Müller ◽  
Marc Schmitz ◽  
Kerstin Borgmann ◽  
...  
Keyword(s):  
Stem Cells ◽  
Dna Damage ◽  
Dna Repair ◽  
Dna Damage Response ◽  
Tumor Evolution ◽  
Regulation Of Transcription ◽  
Mesenchymal Transition ◽  
Damage Response ◽  
Dna Repair Proteins

Cancer stem cells (CSCs) are pluripotent and highly tumorigenic cells that can re-populate a tumor and cause relapses even after initially successful therapy. As with tissue stem cells, CSCs possess enhanced DNA repair mechanisms. An active DNA damage response alleviates the increased oxidative and replicative stress and leads to therapy resistance. On the other hand, mutations in DNA repair genes cause genomic instability, therefore driving tumor evolution and developing highly aggressive CSC phenotypes. However, the role of DNA repair proteins in CSCs extends beyond the level of DNA damage. In recent years, more and more studies have reported the unexpected role of DNA repair proteins in the regulation of transcription, CSC signaling pathways, intracellular levels of reactive oxygen species (ROS), and epithelial–mesenchymal transition (EMT). Moreover, DNA damage signaling plays an essential role in the immune response towards tumor cells. Due to its high importance for the CSC phenotype and treatment resistance, the DNA damage response is a promising target for individualized therapies. Furthermore, understanding the dependence of CSC on DNA repair pathways can be therapeutically exploited to induce synthetic lethality and sensitize CSCs to anti-cancer therapies. This review discusses the different roles of DNA repair proteins in CSC maintenance and their potential as therapeutic targets.

scholarly journals Werner helicase control of human papillomavirus 16 E1-E2 DNA replication is regulated by SIRT1 deacetylation

2018 ◽  
Author(s):  
Dipon Das ◽  
Molly L Bristol ◽  
Nathan W Smith ◽  
Xu Wang ◽  
Pietro Pichierri ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Life Cycle ◽  
Dna Replication ◽  
Dna Damage Response ◽  
Mammalian Cells ◽  
Human Papillomaviruses ◽  
Repair Protein ◽  
Damage Response ◽  
Dna Repair Protein

AbstractHuman papillomaviruses (HPV) are double stranded DNA viruses causative in a host of human diseases including several cancers. Following infection two viral proteins, E1 and E2, activate viral replication in association with cellular factors, and stimulate the DNA damage response (DDR) during the replication process. E1-E2 uses homologous replication (HR) to facilitate DNA replication, but an understanding of host factors involved in this process remains incomplete. Previously we demonstrated that the class III deacetylase SIRT1, which can regulate HR, is recruited to E1-E2 replicating DNA and regulates the level of replication. Here we demonstrate that SIRT1 promotes the fidelity of E1-E2 replication and that the absence of SIRT1 results in reduced recruitment of the DNA repair protein Werner helicase (WRN) to E1-E2 replicating DNA. CRISPR/Cas9 editing demonstrates that WRN, like SIRT1, regulates the quantity and fidelity of E1-E2 replication. This is the first report of WRN regulation of E1-E2 DNA replication, or a role for WRN in the HPV life cycle. In the absence of SIRT1 there is an increased acetylation and stability of WRN, but a reduced ability to interact with E1-E2 replicating DNA. We present a model in which E1-E2 replication turns on the DDR stimulating SIRT1 deacetylation of WRN. This deacetylation promotes WRN interaction with E1-E2 replicating DNA to control the quantity and fidelity of replication. As well as offering a crucial insight into HPV replication control, this system offers a unique model for investigating the link between SIRT1 and WRN in controlling replication in mammalian cells.ImportanceHPV16 is the major viral human carcinogen, responsible for between 3 and 4% of all cancers worldwide. Following infection this virus activates the DNA damage response (DDR) to promote its life cycle, and recruits DDR proteins to its replicating DNA in order to facilitate homologous recombination during replication. This promotes the production of viable viral progeny. Our understanding of how HPV16 replication interacts with the DDR remains incomplete. Here we demonstrate that the cellular deacetylase SIRT1, which is a part of the E1-E2 replication complex, regulates recruitment of the DNA repair protein WRN to the replicating DNA. We demonstrate that WRN regulates the level and fidelity of E1-E2 replication. Overall the results suggest a mechanism where SIRT1 deacetylation of WRN promotes its interaction with E1-E2 replicating DNA to control the levels and fidelity of that replication.

scholarly journals Crosstalk between the nucleolus and the DNA damage response

2017 ◽  
Vol 13 (3) ◽  
pp. 443-455 ◽  
Author(s):  
L. M. Ogawa ◽  
S. J. Baserga
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Dna Damage Response ◽  
Ribosome Biogenesis ◽  
Damage Response ◽  
Dna Repair Proteins

We review the role for conventional DNA repair proteins in ribosome biogenesis and ribosome biogenesis factors in DNA repair.

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