scholarly journals Polo-like kinase 3 regulates CtIP during DNA double-strand break repair in G1

2014 ◽  
Vol 206 (7) ◽  
pp. 877-894 ◽  
Author(s):  
Olivia Barton ◽  
Steffen C. Naumann ◽  
Ronja Diemer-Biehs ◽  
Julia Künzel ◽  
Monika Steinlage ◽  
...  
Keyword(s):  
Large Scale ◽  
Strand Break ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Interacting Protein ◽  
Genomic Deletions ◽  
Cellular Processes ◽  
Dna Double Strand Break ◽  
Radiation Induced ◽  
Alternative Nhej

DNA double-strand breaks (DSBs) are repaired by nonhomologous end joining (NHEJ) or homologous recombination (HR). The C terminal binding protein–interacting protein (CtIP) is phosphorylated in G2 by cyclin-dependent kinases to initiate resection and promote HR. CtIP also exerts functions during NHEJ, although the mechanism phosphorylating CtIP in G1 is unknown. In this paper, we identify Plk3 (Polo-like kinase 3) as a novel DSB response factor that phosphorylates CtIP in G1 in a damage-inducible manner and impacts on various cellular processes in G1. First, Plk3 and CtIP enhance the formation of ionizing radiation-induced translocations; second, they promote large-scale genomic deletions from restriction enzyme-induced DSBs; third, they are required for resection and repair of complex DSBs; and finally, they regulate alternative NHEJ processes in Ku−/− mutants. We show that mutating CtIP at S327 or T847 to nonphosphorylatable alanine phenocopies Plk3 or CtIP loss. Plk3 binds to CtIP phosphorylated at S327 via its Polo box domains, which is necessary for robust damage-induced CtIP phosphorylation at S327 and subsequent CtIP phosphorylation at T847.

scholarly journals Ubiquitin-Specific-Processing Protease 47, A Novel 53BP1 regulator

2021 ◽  
Author(s):  
Doraid T. Sadideen ◽  
Baowei Chen ◽  
Manal Basili ◽  
Montaser Shaheen
Keyword(s):  
Strand Break ◽  
Dependent Manner ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Class Switch ◽  
Dna Double Strand Break ◽  
Radiation Induced ◽  
Non Homologous End Joining ◽  
Processing Protease

AbstractDNA double strand breaks (DSBs) are repair by homology-based repair or non-homologous end joining and multiple sub-pathways exist. 53BP1 is a key DNA double strand break repair protein that regulates repair pathway choice. It is key for joining DSBs during immunoglobulin heavy chain class switch recombination. Here we identify USP47 as a deubiquitylase that associates with and regulates 53BP1 function. USP47 loss results in 53BP1 instability in proteasome dependent manner, and defective 53BP1 ionizing radiation induced foci (IRIF). USP47 catalytic activity is required for maintaining 53BP1 protein level. Similar to 53BP1, USP47 depletion results in sensitivity to DNA DSB inducing agents and defective immunoglobulin CSR. Our findings establish a function for USP47 in DNA DSB repair at least partially through 53BP1.

scholarly journals Nej1(XLF) inhibits Sae2(CTIP)-Dna2(DNA2) mediated resection at DNA double strand breaks

2021 ◽  
Author(s):  
Aditya Mojumdar ◽  
Nancy Adam ◽  
Jennifer A Cobb
Keyword(s):  
Synthetic Lethality ◽  
Nuclease Activity ◽  
Strand Break ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Genomic Deletions ◽  
Dna Double Strand Break ◽  
Break Site ◽  
Dna End Resection ◽  
Non Homologous End Joining

The two major pathways of DNA double strand break (DSB) repair, non-homologous end-joining (NHEJ) and homologous recombination (HR), are highly conserved from yeast to mammals. Regulated 5 DNA end resection is important for repair pathway choice and repair outcomes. Nej1 was first identified as a canonical NHEJ factor involved in stimulating the ligation of broken DNA ends and, more recently, it was shown to be important for DNA end-bridging and inhibiting Dna2-Sgs1 mediated 5 resection. Dna2 localizes to DSBs in the absence of Sgs1 through interactions with Mre11 and Sae2 and DNA damage sensitivity is greater in cells lacking Dna2 nuclease activity compared to sgs1∆ mutants. Dna2-Sae2 mediated 5 resection is down-regulated by Nej1, which itself interacts with Sae2. The resection defect of sae2∆ and the synthetic lethality of sae2∆ sgs1∆ are reversed by deletion of NEJ1 and dependent on Dna2 nuclease activity. Our work demonstrates the importance of Nej1 in inhibiting short-range resection at a DSB by Dna2-Sae2, a critical regulatory mechanism that prevents the formation of genomic deletions at the break site.

scholarly journals RAP80 suppresses the vulnerability of R-loops during DNA double-strand break repair

2021 ◽  
Author(s):  
Takaaki Yasuhara ◽  
Reona Kato ◽  
Motohiro Yamauchi ◽  
Yuki Uchihara ◽  
Lee Zou ◽  
...  
Keyword(s):  
Active Regions ◽  
Strand Break ◽  
Double Strand Breaks ◽  
Critical Component ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Dsb Repair ◽  
Strand Breaks ◽  
Chromosome Translocations ◽  
Dna Double Strand Break

AbstractR-loops, consisting of ssDNA and DNA-RNA hybrids, are potentially vulnerable unless they are appropriately processed. Recent evidence suggests that R-loops can form in the proximity of DNA double-strand breaks (DSBs) within transcriptionally active regions. Yet, how the vulnerability of R-loops is overcome during DSB repair remains unclear. Here, we identify RAP80 as a factor suppressing the vulnerability of ssDNA in R-loops and chromosome translocations and deletions during DSB repair. Mechanistically, RAP80 prevents unscheduled nucleolytic processing of ssDNA in R-loops by CtIP. This mechanism promotes efficient DSB repair via transcription-associated end-joining dependent on BRCA1, Polθ, and LIG1/3. Thus, RAP80 suppresses the vulnerability of R-loops during DSB repair, thereby precluding genomic abnormalities in a critical component of the genome caused by deleterious R-loop processing.

scholarly journals Regulation of DNA Double-Strand Break Repair by Non-Coding RNAs

2018 ◽  
Author(s):  
Roopa Thapar
Keyword(s):  
Strand Break ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Dna Double Strand Break ◽  
Protein Dna Interactions ◽  
Damage Response ◽  
Non Coding Rnas ◽  
Non Homologous End Joining

DNA double-strand breaks (DSBs) are deleterious lesions that are generated in response to ionizing radiation or replication fork collapse that can lead to genomic instability and cancer.  Eukaryotes have evolved two major pathways, namely homologous recombination (HR) and non-homologous end joining (NHEJ) to repair DSBs.  Whereas the roles of protein-DNA interactions in HR and NHEJ have been fairly well defined, the functions of small and long non-coding RNAs and RNA-DNA hybrids in the DNA damage response is just beginning to be elucidated.  This review summarizes recent discoveries on the identification of non-coding RNAs and RNA-mediated regulation of DSB repair

scholarly journals SIRT6 is a DNA double-strand break sensor

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Lior Onn ◽  
Miguel Portillo ◽  
Stefan Ilic ◽  
Gal Cleitman ◽  
Daniel Stein ◽  
...  
Keyword(s):  
Dna Damage ◽  
Binding Capacity ◽  
Critical Factor ◽  
Strand Break ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Downstream Signaling ◽  
The Road ◽  
Dna Double Strand Break ◽  
Non Homologous End Joining

DNA double-strand breaks (DSB) are the most deleterious type of DNA damage. In this work, we show that SIRT6 directly recognizes DNA damage through a tunnel-like structure that has high affinity for DSB. SIRT6 relocates to sites of damage independently of signaling and known sensors. It activates downstream signaling for DSB repair by triggering ATM recruitment, H2AX phosphorylation and the recruitment of proteins of the homologous recombination and non-homologous end joining pathways. Our findings indicate that SIRT6 plays a previously uncharacterized role as a DNA damage sensor, a critical factor in initiating the DNA damage response (DDR). Moreover, other Sirtuins share some DSB-binding capacity and DDR activation. SIRT6 activates the DDR before the repair pathway is chosen, and prevents genomic instability. Our findings place SIRT6 as a sensor of DSB, and pave the road to dissecting the contributions of distinct DSB sensors in downstream signaling.

Non-canonical function of DGCR8 in DNA double-strand break repair signaling and tumor radioresistance

2020 ◽  
Author(s):  
Qinglei Hang ◽  
Liyong Zeng ◽  
Li Wang ◽  
Litong Nie ◽  
Fan Yao ◽  
...  
Keyword(s):  
Radiation Treatment ◽  
Critical Role ◽  
Strand Break ◽  
Histone H2a ◽  
Tumor Resistance ◽  
Dna Double Strand Breaks ◽  
Canonical Function ◽  
Dna Double Strand Break ◽  
Radiation Induced ◽  
Protein Recruitment

Abstract Cells respond to cytotoxic DNA double-strand breaks (DSBs) by recruiting repair proteins to the damaged sites. During the DNA damage response, ubiquitin signaling plays a critical role in coordinating protein recruitment. Here, we find that the microRNA biogenesis factor DGCR8 promotes tumor resistance to X-ray radiation independently of its Drosha-binding ability. In response to radiation, the deubiquitinase USP51 and the kinase ATM mediate the stabilization and activation of DGCR8 through deubiquitination and phosphorylation, respectively. While radiation-induced USP51 binds, deubiquitinates, and stabilizes DGCR8, ATM-dependent phosphorylation of DGCR8 at serine 677 leads to the recruitment of DGCR8 and DGCR8’s binding partner RNF168 to MDC1 and RNF8. This, in turn, promotes ubiquitination of histone H2A, repair of DSBs, and radioresistance. Altogether, these findings reveal the non-canonical function of DGCR8 in DSB repair and suggest that radiation treatment may result in therapy-induced tumor radioresistance through USP51- and ATM-mediated upregulation and activation of DGCR8.

scholarly journals Structural snapshots of human DNA polymerase μ engaged on a DNA double-strand break

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Andrea M. Kaminski ◽  
John M. Pryor ◽  
Dale A. Ramsden ◽  
Thomas A. Kunkel ◽  
Lars C. Pedersen ◽  
...  
Keyword(s):  
Chromosomal Rearrangements ◽  
Strand Break ◽  
Nonhomologous End Joining ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Template Strand ◽  
Single Strand Breaks ◽  
Dna Double Strand Break ◽  
Break Site

Abstract Genomic integrity is threatened by cytotoxic DNA double-strand breaks (DSBs), which must be resolved efficiently to prevent sequence loss, chromosomal rearrangements/translocations, or cell death. Polymerase μ (Polμ) participates in DSB repair via the nonhomologous end-joining (NHEJ) pathway, by filling small sequence gaps in broken ends to create substrates ultimately ligatable by DNA Ligase IV. Here we present structures of human Polμ engaging a DSB substrate. Synapsis is mediated solely by Polμ, facilitated by single-nucleotide homology at the break site, wherein both ends of the discontinuous template strand are stabilized by a hydrogen bonding network. The active site in the quaternary Pol μ complex is poised for catalysis and nucleotide incoporation proceeds in crystallo. These structures demonstrate that Polμ may address complementary DSB substrates during NHEJ in a manner indistinguishable from single-strand breaks.

scholarly journals RAD-51-Dependent and -Independent Roles of a Caenorhabditis elegans BRCA2-Related Protein during DNA Double-Strand Break Repair

2005 ◽  
Vol 25 (8) ◽  
pp. 3127-3139 ◽  
Author(s):  
Julie S. Martin ◽  
Nicole Winkelmann ◽  
Mark I. R. Petalcorin ◽  
Michael J. McIlwraith ◽  
Simon J. Boulton
Keyword(s):  
Caenorhabditis Elegans ◽  
Strand Break ◽  
Double Strand Break ◽  
Nonhomologous End Joining ◽  
Related Protein ◽  
End Joining ◽  
Dsb Repair ◽  
Phenotypic Differences ◽  
Dna Double Strand Break ◽  
Radiation Induced

ABSTRACT The BRCA2 tumor suppressor is implicated in DNA double-strand break (DSB) repair by homologous recombination (HR), where it regulates the RAD51 recombinase. We describe a BRCA2-related protein of Caenorhabditis elegans (CeBRC-2) that interacts directly with RAD-51 via a single BRC motif and that binds preferentially to single-stranded DNA through an oligonucleotide-oligosaccharide binding fold. Cebrc-2 mutants fail to repair meiotic or radiation-induced DSBs by HR due to inefficient RAD-51 nuclear localization and a failure to target RAD-51 to sites of DSBs. Genetic and cytological comparisons of Cebrc-2 and rad-51 mutants revealed fundamental phenotypic differences that suggest a role for Cebrc-2 in promoting the use of an alternative repair pathway in the absence of rad-51 and independent of nonhomologous end joining (NHEJ). Unlike rad-51 mutants, Cebrc-2 mutants also accumulate RPA-1 at DSBs, and abnormal chromosome aggregates that arise during the meiotic prophase can be rescued by blocking the NHEJ pathway. CeBRC-2 also forms foci in response to DNA damage and can do so independently of rad-51. Thus, CeBRC-2 not only regulates RAD-51 during HR but can also function independently of rad-51 in DSB repair processes.

scholarly journals Akt/PKB suppresses DNA damage processing and checkpoint activation in late G2

2010 ◽  
Vol 190 (3) ◽  
pp. 297-305 ◽  
Author(s):  
Naihan Xu ◽  
Nadia Hegarat ◽  
Elizabeth J. Black ◽  
Mary T. Scott ◽  
Helfrid Hochegger ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Protein A ◽  
Strand Break ◽  
Checkpoint Activation ◽  
Interacting Protein ◽  
Dna Double Strand Break ◽  
Radiation Induced ◽  
G2 Cells ◽  
Tumor Types

Using chemical genetics to reversibly inhibit Cdk1, we find that cells arrested in late G2 are unable to delay mitotic entry after irradiation. Late G2 cells detect DNA damage lesions and form γ-H2AX foci but fail to activate Chk1. This reflects a lack of DNA double-strand break processing because late G2 cells fail to recruit RPA (replication protein A), ATR (ataxia telangiectasia and Rad3 related), Rad51, or CtIP (C-terminal interacting protein) to sites of radiation-induced damage, events essential for both checkpoint activation and initiation of DNA repair by homologous recombination. Remarkably, inhibition of Akt/PKB (protein kinase B) restores DNA damage processing and Chk1 activation after irradiation in late G2. These data demonstrate a previously unrecognized role for Akt in cell cycle regulation of DNA repair and checkpoint activation. Because Akt/PKB is frequently activated in many tumor types, these findings have important implications for the evolution and therapy of such cancers.

Regulation of the cellular DNA double-strand break response

2007 ◽  
Vol 85 (6) ◽  
pp. 663-674 ◽  
Author(s):  
Kendra L. Cann ◽  
Geoffrey G. Hicks
Keyword(s):  
Mammalian Cells ◽  
Strand Break ◽  
Double Strand Break ◽  
Double Strand Breaks ◽  
Cell Cycle Checkpoints ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Dna Double Strand Break ◽  
Cellular Dna

DNA double-strand breaks occur frequently in cycling cells, and are also induced by exogenous sources, including ionizing radiation. Cells have developed integrated double-strand break response pathways to cope with these lesions, including pathways that initiate DNA repair (either via homologous recombination or nonhomologous end joining), the cell-cycle checkpoints (G1–S, intra-S phase, and G2–M) that provide time for repair, and apoptosis. However, before any of these pathways can be activated, the damage must first be recognized. In this review, we will discuss how the response of mammalian cells to DNA double-strand breaks is regulated, beginning with the activation of ATM, the pinnacle kinase of the double-strand break signalling cascade.

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