scholarly journals A complex of BRCA2 and PP2A-B56 is required for DNA repair by homologous recombination

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Sara M. Ambjørn ◽  
Julien P. Duxin ◽  
Emil P. T. Hertz ◽  
Isha Nasa ◽  
Joana Duro ◽  
...  
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Suppressor Gene ◽  
Tumour Suppressor Gene ◽  
Parp Inhibition ◽  
Binding Motif ◽  
Filament Formation ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Atr Kinases

AbstractMutations in the tumour suppressor gene BRCA2 are associated with predisposition to breast and ovarian cancers. BRCA2 has a central role in maintaining genome integrity by facilitating the repair of toxic DNA double-strand breaks (DSBs) by homologous recombination (HR). BRCA2 acts by controlling RAD51 nucleoprotein filament formation on resected single-stranded DNA, but how BRCA2 activity is regulated during HR is not fully understood. Here, we delineate a pathway where ATM and ATR kinases phosphorylate a highly conserved region in BRCA2 in response to DSBs. These phosphorylations stimulate the binding of the protein phosphatase PP2A-B56 to BRCA2 through a conserved binding motif. We show that the phosphorylation-dependent formation of the BRCA2-PP2A-B56 complex is required for efficient RAD51 filament formation at sites of DNA damage and HR-mediated DNA repair. Moreover, we find that several cancer-associated mutations in BRCA2 deregulate the BRCA2-PP2A-B56 interaction and sensitize cells to PARP inhibition. Collectively, our work uncovers PP2A-B56 as a positive regulator of BRCA2 function in HR with clinical implications for BRCA2 and PP2A-B56 mutated cancers.

scholarly journals A complex of BRCA2 and PP2A-B56 is required for DNA repair by homologous recombination

2021 ◽  
Author(s):  
Sara Marie Ambjoern ◽  
Julien P Duxin ◽  
Emil PT Hertz ◽  
Isha Nasa ◽  
Joana Duro ◽  
...  
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Suppressor Gene ◽  
Tumour Suppressor Gene ◽  
Double Strand Breaks ◽  
Parp Inhibition ◽  
Binding Motif ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Atr Kinases

Mutations in the tumour suppressor gene BRCA2 are associated with predisposition to breast and ovarian cancers. BRCA2 has a central role in maintaining genome integrity by facilitating the repair of toxic DNA double-strand breaks (DSBs) by homologous recombination (HR). BRCA2 acts by promoting RAD51 nucleoprotein filament formation on resected single-stranded DNA, but how BRCA2 activity is regulated during HR is not fully understood. Here, we delineate a pathway where ATM and ATR kinases phosphorylate a highly conserved region in BRCA2 in response to DSBs. These phosphorylations stimulate the binding of the protein phosphatase PP2A-B56 to BRCA2 through a conserved binding motif. We show that the phosphorylation-dependent formation of the BRCA2-PP2A-B56 complex is required for efficient RAD51 loading to sites of DNA damage and HR-mediated DNA repair. Moreover, we find that several cancer-associated mutations in BRCA2 deregulate the BRCA2-PP2A-B56 interaction and sensitize cells to PARP inhibition. Collectively, our work uncovers PP2A-B56 as a positive regulator of BRCA2 function in HR with clinical implications for BRCA2 and PP2A-B56 mutated cancers.

scholarly journals End-resection at DNA double-strand breaks in the three domains of life

2013 ◽  
Vol 41 (1) ◽  
pp. 314-320 ◽  
Author(s):  
John K. Blackwood ◽  
Neil J. Rzechorzek ◽  
Sian M. Bray ◽  
Joseph D. Maman ◽  
Luca Pellegrini ◽  
...  
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Homologous Chromosomes ◽  
Domains Of Life ◽  
Strand Invasion ◽  
End Resection

During DNA repair by HR (homologous recombination), the ends of a DNA DSB (double-strand break) must be resected to generate single-stranded tails, which are required for strand invasion and exchange with homologous chromosomes. This 5′–3′ end-resection of the DNA duplex is an essential process, conserved across all three domains of life: the bacteria, eukaryota and archaea. In the present review, we examine the numerous and redundant helicase and nuclease systems that function as the enzymatic analogues for this crucial process in the three major phylogenetic divisions.

scholarly journals Identification of MCM8IP, an interactor of MCM8-9 and RPA1 that promotes homologous recombination and DNA synthesis in response to DNA damage

2019 ◽  
Author(s):  
Jen-Wei Huang ◽  
Angelo Taglialatela ◽  
Ananya Acharya ◽  
Giuseppe Leuzzi ◽  
Tarun S. Nambiar ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Homologous Recombination ◽  
Dna Synthesis ◽  
Dna Recombination ◽  
Direct Interaction ◽  
Response To Treatment ◽  
Parp Inhibition ◽  
Dna Double Strand Breaks ◽  
Crosslinking Agents

ABSTRACTHomologous recombination (HR) mediates the error-free repair of DNA double-strand breaks to maintain genomic stability. HR is carried out by a complex network of DNA repair factors. Here we identify C17orf53/MCM8IP, an OB-fold containing protein that binds ssDNA, as a novel DNA repair factor involved in HR. MCM8IP-deficient cells exhibit HR defects, especially in long-tract gene conversion, occurring downstream of RAD51 loading, consistent with a role for MCM8IP in HR-dependent DNA synthesis. Moreover, loss of MCM8IP confers cellular sensitivity to crosslinking agents and PARP inhibition. Importantly, we identify a direct interaction with MCM8-9, a putative helicase complex mutated in Primary Ovarian Insufficiency, that is crucial for MCM8IP’s ability to promote resistance to DNA damaging agents. In addition to its association with MCM8-9, MCM8IP also binds directly to RPA1. We show that the interactions of MCM8IP with both MCM8-9 and RPA are required to maintain replication fork progression in response to treatment with crosslinking agents. Collectively, our work identifies MCM8IP as a key regulator of DNA damage-associated DNA synthesis during DNA recombination and replication.

scholarly journals RNF19A-mediated ubiquitination of BARD1 prevents BRCA1/BARD1-dependent homologous recombination

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Qian Zhu ◽  
Jinzhou Huang ◽  
Hongyang Huang ◽  
Huan Li ◽  
Peiqiang Yi ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Dna Repair ◽  
Homologous Recombination ◽  
Cancer Cell ◽  
Nuclear Export ◽  
Molecular Mechanisms ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Polymerase Inhibitors

AbstractBRCA1-BARD1 heterodimers act in multiple steps during homologous recombination (HR) to ensure the prompt repair of DNA double strand breaks. Dysfunction of the BRCA1 pathway enhances the therapeutic efficiency of poly-(ADP-ribose) polymerase inhibitors (PARPi) in cancers, but the molecular mechanisms underlying this sensitization to PARPi are not fully understood. Here, we show that cancer cell sensitivity to PARPi is promoted by the ring between ring fingers (RBR) protein RNF19A. We demonstrate that RNF19A suppresses HR by ubiquitinating BARD1, which leads to dissociation of BRCA1-BARD1 complex and exposure of a nuclear export sequence in BARD1 that is otherwise masked by BRCA1, resulting in the export of BARD1 to the cytoplasm. We provide evidence that high RNF19A expression in breast cancer compromises HR and increases sensitivity to PARPi. We propose that RNF19A modulates the cancer cell response to PARPi by negatively regulating the BRCA1-BARD1 complex and inhibiting HR-mediated DNA repair.

STEM-22. TARGETING REPLICATION STRESS RESPONSE FOR GLIOMA STEM CELL SPECIFIC CYTOTOXICITY

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi25-vi26
Author(s):  
Emily Clough ◽  
Tracy Ballinger ◽  
Colin Semple ◽  
Karen Strathdee ◽  
ross carruthers
Keyword(s):  
Dna Repair ◽  
Replication Stress ◽  
Nuclear Bodies ◽  
Parp Inhibition ◽  
Dna Double Strand Breaks ◽  
Origin Firing ◽  
Strand Breaks ◽  
Specific Cytotoxicity ◽  
Dna Replication Stress

Abstract BACKGROUND Evidence suggests treatment resistant glioma stem cells (GSCs) drive glioblastoma (GBM) recurrence. Current treatments fail to eradicate GSC and novel GSC targeting therapies are a priority. GSC exhibit elevated DNA replication stress (RS) versus non GSC tumour cells driving constitutive DNA damage response (DDR) activation and efficient DNA repair. We previously demonstrated that targeting RS response with combined ATR and PARP inhibition (CAiPi) (VE821 and Olaparib) provides potent GSC specific cytotoxicity. In this study we investigated the underlying DDR phenotype which determines this vulnerability. RESULTS Paired GSC enriched (‘GSC’) and GSC depleted, differentiated (‘bulk’) populations were cultured from GBM specimens in neurobasal media with growth factors or serum containing media respectively. GSC exhibited reduced survival following exposure to CAiPi versus bulk. CAiPi significantly increased 53BP1 G1 phase nuclear bodies (53BP1NBs) in GSC, which are known to shield under-replicated DNA in actively transcribed genes. Mapping the genomic distribution of endogenously occurring replication dependent DNA double strand breaks via Breaks Ligation In Situ Sequencing (BLISS), revealed reduced intragenic DSB in long actively transcribed genes in GSC versus bulk at baseline, suggesting a reliance upon transcription coupled DSB repair in GSC. RNA seq demonstrated CAiPi-induced transcriptomic alterations in GSC including replication regulation and initiation. DNA fibre assay showed that CAiPi increased GSC new origin firing which correlated with PARP trapping. GSCs were rescued from CAiPi by roscovitine induced inhibition of excess origin firing. CAiPi is potently radiosensitizing by clonogenic assay and we demonstrated murine blood brain barrier penetration of CAiPi utilising VE822 and pamiparib in vivo. CONCLUSION Dysregulation of origin firing by CAiPi exposes a GSC specific vulnerability which results in DNA under-replication and abrogation of proficient DNA repair seen at long actively transcribed genes and has potential to be clinically translated as a GSC specific cytotoxic therapy.

scholarly journals E2F1 acetylation directs p300/CBP-mediated histone acetylation at DNA double-strand breaks to facilitate repair

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Swarnalatha Manickavinayaham ◽  
Renier Vélez-Cruz ◽  
Anup K. Biswas ◽  
Ella Bedford ◽  
Brianna J. Klein ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Dna Damage ◽  
Dna Repair ◽  
Histone Acetylation ◽  
Double Strand Breaks ◽  
Tumor Suppressor Protein ◽  
Binding Motif ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Expression Of Genes

Abstract E2F1 and retinoblastoma (RB) tumor-suppressor protein not only regulate the periodic expression of genes important for cell proliferation, but also localize to DNA double-strand breaks (DSBs) to promote repair. E2F1 is acetylated in response to DNA damage but the role this plays in DNA repair is unknown. Here we demonstrate that E2F1 acetylation creates a binding motif for the bromodomains of the p300/KAT3B and CBP/KAT3A acetyltransferases and that this interaction is required for the recruitment of p300 and CBP to DSBs and the induction of histone acetylation at sites of damage. A knock-in mutation that blocks E2F1 acetylation abolishes the recruitment of p300 and CBP to DSBs and also the accumulation of other chromatin modifying activities and repair factors, including Tip60, BRG1 and NBS1, and renders mice hypersensitive to ionizing radiation (IR). These findings reveal an important role for E2F1 acetylation in orchestrating the remodeling of chromatin structure at DSBs to facilitate repair.

scholarly journals Switch-Like Phosphorylation of Wrn Integrates End-Resection With Repair of Dsbs at Replication Forks

2018 ◽  
Author(s):  
Valentina Palermo ◽  
Eva Malacaria ◽  
Massimo Sanchez ◽  
Annapaola Franchitto ◽  
Pietro Pichierri
Keyword(s):  
Homologous Recombination ◽  
Long Range ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Strand Invasion ◽  
Wrn Helicase ◽  
End Resection ◽  
Atr Kinases ◽  
Dna Ends

ABSTRACTReplication-dependent DNA double-strand breaks are harmful lesions preferentially repaired by homologous recombination, a process that requires processing of DNA ends to allow RAD51-mediated strand invasion. End-resection and subsequent repair are two intertwined processes, but the mechanism underlying their execution is still poorly appreciated. The WRN helicase is one of the crucial factors for the end-resection and is instrumental to select the proper repair pathway. Here, we reveal that ordered phosphorylation of WRN by the CDK1, ATM and ATR kinases define a complex regulatory layer that is essential for correct long-range end-resection connecting it to repair by homologous recombination. We establish that long-range end-resection requires an ATM-dependent phosphorylation of WRN at Ser1058 and that phosphorylation at Ser1141, together with dephosphorylation at the CDK1 site Ser1133, is needed to conclude long-range end-resection and support RAD51-dependent repair. Collectively, our findings suggest that regulation of WRN by multiple kinases functions as molecular switch to allow a timely execution of end-resection and repair at replication-dependent DNA double-strand breaks.

scholarly journals A PALB2-interacting domain in RNF168 couples homologous recombination to DNA break-induced chromatin ubiquitylation

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Martijn S Luijsterburg ◽  
Dimitris Typas ◽  
Marie-Christine Caron ◽  
Wouter W Wiegant ◽  
Diana van den Heuvel ◽  
...  
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Direct Interaction ◽  
Histone H2a ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Histone Ubiquitylation ◽  
G2 Cells ◽  
G1 Cells ◽  
Interacting Domain

DNA double-strand breaks (DSB) elicit a ubiquitylation cascade that controls DNA repair pathway choice. This cascade involves the ubiquitylation of histone H2A by the RNF168 ligase and the subsequent recruitment of RIF1, which suppresses homologous recombination (HR) in G1 cells. The RIF1-dependent suppression is relieved in S/G2 cells, allowing PALB2-driven HR to occur. With the inhibitory impact of RIF1 relieved, it remains unclear how RNF168-induced ubiquitylation influences HR. Here, we uncover that RNF168 links the HR machinery to H2A ubiquitylation in S/G2 cells. We show that PALB2 indirectly recognizes histone ubiquitylation by physically associating with ubiquitin-bound RNF168. This direct interaction is mediated by the newly identified PALB2-interacting domain (PID) in RNF168 and the WD40 domain in PALB2, and drives DNA repair by facilitating the assembly of PALB2-containing HR complexes at DSBs. Our findings demonstrate that RNF168 couples PALB2-dependent HR to H2A ubiquitylation to promote DNA repair and preserve genome integrity.

scholarly journals REV7/FANCV Binds to CHAMP1 and Promotes Homologous Recombination Repair

2021 ◽  
Author(s):  
Feng Li ◽  
Prabha Sarangi ◽  
Hanrong Feng ◽  
Lisa Moreau ◽  
Huy Nguyen ◽  
...  
Keyword(s):  
Dna Repair ◽  
Homologous Recombination ◽  
Parp Inhibitor ◽  
Dna Double Strand Breaks ◽  
Critical Determinant ◽  
Strand Breaks ◽  
Positive Regulator ◽  
Recombination Repair ◽  
Nhej Repair ◽  
Inhibitor Resistance

A critical determinant of DNA repair pathway choice is the HORMA protein REV7, a small abundant adaptor which binds to various DNA repair proteins through its C-terminal seatbelt domain. The REV7 seatbelt binds to the REV3 polymerase to form the Polymerase ζ complex, a positive regulator of translesion synthesis (TLS) repair. Alternatively, the REV7 seatbelt binds to SHLD3 in the Shieldin complex, a positive regulator of NHEJ repair. Recent studies have identified another novel REV7 seatbelt-binding protein, CHAMP1 (Chromosome Alignment-Maintaining Phosphoprotein, though its role in DNA repair is unknown. Here, we show that the REV7-CHAMP1 complex promotes homologous recombination (HR) repair by sequestering REV7 from the Shieldin complex. CHAMP1 competes directly with the SHLD3 subunit of the Shieldin complex for a limited pool of C-REV7, thereby inhibiting the REV7-mediated recruitment of the SHLD2 and SHLD1 effector subunits to DNA double strand breaks. CHAMP1 thereby channels DNA repair away from error-prone NHEJ and towards the competing error-free HR pathway. Similarly, CHAMP1 competes with the REV3 component of the POLζ complex, thereby reducing the level of mutagenic TLS repair. CHAMP1 interacts with POGZ in a heterochromatin complex further promoting HR repair. Importantly, in human tumors, CHAMP1 overexpression promotes HR, confers PARP inhibitor resistance, and correlates with poor prognosis. Thus, by binding to either REV3, SHLD3, or CHAMP1 through its seatbelt, the REV7 protein can promote either TLS repair, NHEJ repair, or HR repair respectively.

Sign in / Sign up

Export Citation Format

Share Document