scholarly journals DNA damage signaling in response to double-strand breaks during mitosis

2010 ◽  
Vol 190 (2) ◽  
pp. 197-207 ◽  
Author(s):  
Simona Giunta ◽  
Rimma Belotserkovskaya ◽  
Stephen P. Jackson
Keyword(s):  
Dna Damage ◽  
Mitotic Cell ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Histone H2ax ◽  
Mrn Complex ◽  
Dna Damage Signaling ◽  
Dependent Protein ◽  
Mitotic Cells

The signaling cascade initiated in response to DNA double-strand breaks (DSBs) has been extensively investigated in interphase cells. Here, we show that mitotic cells treated with DSB-inducing agents activate a “primary” DNA damage response (DDR) comprised of early signaling events, including activation of the protein kinases ataxia telangiectasia mutated (ATM) and DNA-dependent protein kinase (DNA-PK), histone H2AX phosphorylation together with recruitment of mediator of DNA damage checkpoint 1 (MDC1), and the Mre11–Rad50–Nbs1 (MRN) complex to damage sites. However, mitotic cells display no detectable recruitment of the E3 ubiquitin ligases RNF8 and RNF168, or accumulation of 53BP1 and BRCA1, at DSB sites. Accordingly, we found that DNA-damage signaling is attenuated in mitotic cells, with full DDR activation only ensuing when a DSB-containing mitotic cell enters G1. Finally, we present data suggesting that induction of a primary DDR in mitosis is important because transient inactivation of ATM and DNA-PK renders mitotic cells hypersensitive to DSB-inducing agents.

scholarly journals MDC1 PST-repeat region promotes histone H2AX-independent chromatin association and DNA damage tolerance

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Israel Salguero ◽  
Rimma Belotserkovskaya ◽  
Julia Coates ◽  
Matylda Sczaniecka-Clift ◽  
Mukerrem Demir ◽  
...  
Keyword(s):  
Dna Damage ◽  
Double Strand Breaks ◽  
Repeat Region ◽  
Independent Function ◽  
Dna Double Strand Breaks ◽  
Dna Damage Tolerance ◽  
Strand Breaks ◽  
Histone H2ax ◽  
Independent Association ◽  
Dna Damage Signalling

AbstractHistone H2AX and MDC1 are key DNA repair and DNA-damage signalling proteins. When DNA double-strand breaks (DSBs) occur, H2AX is phosphorylated and then recruits MDC1, which in turn serves as a docking platform to promote the localization of other factors, including 53BP1, to DSB sites. Here, by using CRISPR-Cas9 engineered human cell lines, we identify a hitherto unknown, H2AX-independent, function of MDC1 mediated by its PST-repeat region. We show that the PST-repeat region directly interacts with chromatin via the nucleosome acidic patch and mediates DNA damage-independent association of MDC1 with chromatin. We find that this region is largely functionally dispensable when the canonical γH2AX-MDC1 pathway is operative but becomes critical for 53BP1 recruitment to DNA-damage sites and cell survival following DSB induction when H2AX is not available. Consequently, our results suggest a role for MDC1 in activating the DDR in areas of the genome lacking or depleted of H2AX.

scholarly journals Class switching and meiotic defects in mice lacking the E3 ubiquitin ligase RNF8

2010 ◽  
Vol 207 (5) ◽  
pp. 973-981 ◽  
Author(s):  
Margarida Almeida Santos ◽  
Michael S.Y. Huen ◽  
Mila Jankovic ◽  
Hua-Tang Chen ◽  
Andrés J. López-Contreras ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cellular Response ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Histone H2ax ◽  
Class Switch ◽  
Repair Defect ◽  
Dependent Pathway ◽  
Step Mechanism

53BP1 is a well-known mediator of the cellular response to DNA damage. Two alternative mechanisms have been proposed to explain 53BP1’s interaction with DNA double-strand breaks (DSBs), one by binding to methylated histones and the other via an RNF8 E3 ligase–dependent ubiquitylation pathway. The formation of RNF8 and 53BP1 irradiation-induced foci are both dependent on histone H2AX. To evaluate the contribution of the RNF8-dependent pathway to 53BP1 function, we generated RNF8 knockout mice. We report that RNF8 deficiency results in defective class switch recombination (CSR) and accumulation of unresolved immunoglobulin heavy chain–associated DSBs. The CSR DSB repair defect is milder than that observed in the absence of 53BP1 but similar to that found in H2AX−/− mice. Moreover, similar to H2AX but different from 53BP1 deficiency, RNF8−/− males are sterile, and this is associated with defective ubiquitylation of the XY chromatin. Combined loss of H2AX and RNF8 does not cause further impairment in CSR, demonstrating that the two genes function epistatically. Importantly, although 53BP1 foci formation is RNF8 dependent, its binding to chromatin is preserved in the absence of RNF8. This suggests a two-step mechanism for 53BP1 association with chromatin in which constitutive loading is dependent on interactions with methylated histones, whereas DNA damage–inducible RNF8-dependent ubiquitylation allows its accumulation at damaged chromatin.

scholarly journals Changes in chromatin structure and mobility in living cells at sites of DNA double-strand breaks

2006 ◽  
Vol 172 (6) ◽  
pp. 823-834 ◽  
Author(s):  
Michael J. Kruhlak ◽  
Arkady Celeste ◽  
Graham Dellaire ◽  
Oscar Fernandez-Capetillo ◽  
Waltraud G. Müller ◽  
...  
Keyword(s):  
Dna Damage ◽  
Living Cells ◽  
Double Strand Breaks ◽  
Dna Integrity ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Limited Mobility ◽  
Energy Filtering ◽  
Dna Damage Signaling ◽  
Cell Biol

The repair of DNA double-strand breaks (DSBs) is facilitated by the phosphorylation of H2AX, which organizes DNA damage signaling and chromatin remodeling complexes in the vicinity of the lesion (Pilch, D.R., O.A. Sedelnikova, C. Redon, A. Celeste, A. Nussenzweig, and W.M. Bonner. 2003. Biochem. Cell Biol. 81:123–129; Morrison, A.J., and X. Shen. 2005. Cell Cycle. 4:568–571; van Attikum, H., and S.M. Gasser. 2005. Nat. Rev. Mol. Cell. Biol. 6:757–765). The disruption of DNA integrity induces an alteration of chromatin architecture that has been proposed to activate the DNA damage transducing kinase ataxia telangiectasia mutated (ATM; Bakkenist, C.J., and M.B. Kastan. 2003. Nature. 421:499–506). However, little is known about the physical properties of damaged chromatin. In this study, we use a photoactivatable version of GFP-tagged histone H2B to examine the mobility and structure of chromatin containing DSBs in living cells. We find that chromatin containing DSBs exhibits limited mobility but undergoes an energy-dependent local expansion immediately after DNA damage. The localized expansion observed in real time corresponds to a 30–40% reduction in the density of chromatin fibers in the vicinity of DSBs, as measured by energy-filtering transmission electron microscopy. The observed opening of chromatin occurs independently of H2AX and ATM. We propose that localized adenosine triphosphate–dependent decondensation of chromatin at DSBs establishes an accessible subnuclear environment that facilitates DNA damage signaling and repair.

Persistent DNA damage signaling and DNA polymerase theta promote broken chromosome segregation

2021 ◽  
Vol 220 (12) ◽  
Author(s):  
Delisa E. Clay ◽  
Heidi S. Bretscher ◽  
Erin A. Jezuit ◽  
Korie B. Bush ◽  
Donald T. Fox
Keyword(s):  
Dna Damage ◽  
Dna Polymerase ◽  
Chromosome Segregation ◽  
Genome Instability ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Dna Damage Signaling ◽  
Alternative End Joining

Cycling cells must respond to DNA double-strand breaks (DSBs) to avoid genome instability. Missegregation of chromosomes with DSBs during mitosis results in micronuclei, aberrant structures linked to disease. How cells respond to DSBs during mitosis is incompletely understood. We previously showed that Drosophilamelanogaster papillar cells lack DSB checkpoints (as observed in many cancer cells). Here, we show that papillar cells still recruit early acting repair machinery (Mre11 and RPA3) and the Fanconi anemia (FA) protein Fancd2 to DSBs. These proteins persist as foci on DSBs as cells enter mitosis. Repair foci are resolved in a stepwise manner during mitosis. DSB repair kinetics depends on both monoubiquitination of Fancd2 and the alternative end-joining protein DNA polymerase θ. Disruption of either or both of these factors causes micronuclei after DNA damage, which disrupts intestinal organogenesis. This study reveals a mechanism for how cells with inactive DSB checkpoints can respond to DNA damage that persists into mitosis.

scholarly journals The chromatin remodeler p400 ATPase facilitates Rad51-mediated repair of DNA double-strand breaks

2012 ◽  
Vol 199 (7) ◽  
pp. 1067-1081 ◽  
Author(s):  
Céline Courilleau ◽  
Catherine Chailleux ◽  
Alain Jauneau ◽  
Fanny Grimal ◽  
Sébastien Briois ◽  
...  
Keyword(s):  
Dna Damage ◽  
Chromatin Remodeling ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Dsb Repair ◽  
Strand Breaks ◽  
Homology Directed Repair ◽  
Dna Damage Signaling ◽  
Dna Dsbs ◽  
Dependent Processes

DNA damage signaling and repair take place in a chromatin context. Consequently, chromatin-modifying enzymes, including adenosine triphosphate–dependent chromatin remodeling enzymes, play an important role in the management of DNA double-strand breaks (DSBs). Here, we show that the p400 ATPase is required for DNA repair by homologous recombination (HR). Indeed, although p400 is not required for DNA damage signaling, DNA DSB repair is defective in the absence of p400. We demonstrate that p400 is important for HR-dependent processes, such as recruitment of Rad51 to DSB (a key component of HR), homology-directed repair, and survival after DNA damage. Strikingly, p400 and Rad51 are present in the same complex and both favor chromatin remodeling around DSBs. Altogether, our data provide a direct molecular link between Rad51 and a chromatin remodeling enzyme involved in chromatin decompaction around DNA DSBs.

scholarly journals Alterations in hormonal signals spatially coordinate distinct responses to DNA double-strand breaks in Arabidopsis roots

2021 ◽  
Vol 7 (25) ◽  
pp. eabg0993
Author(s):  
Naoki Takahashi ◽  
Soichi Inagaki ◽  
Kohei Nishimura ◽  
Hitoshi Sakakibara ◽  
Ioanna Antoniadi ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Root Apex ◽  
Double Strand Breaks ◽  
Root Tip ◽  
Dna Double Strand Breaks ◽  
Cytokinin Signaling ◽  
Strand Breaks ◽  
Dna Damage Signaling ◽  
Cycle Arrest

Plants have a high ability to cope with changing environments and grow continuously throughout life. However, the mechanisms by which plants strike a balance between stress response and organ growth remain elusive. Here, we found that DNA double-strand breaks enhance the accumulation of cytokinin hormones through the DNA damage signaling pathway in the Arabidopsis root tip. Our data showed that activation of cytokinin signaling suppresses the expression of some of the PIN-FORMED genes that encode efflux carriers of another hormone, auxin, thereby decreasing the auxin signals in the root tip and causing cell cycle arrest at G2 phase and stem cell death. Elevated cytokinin signaling also promotes an early transition from cell division to endoreplication in the basal part of the root apex. We propose that plant hormones spatially coordinate differential DNA damage responses, thereby maintaining genome integrity and minimizing cell death to ensure continuous root growth.

scholarly journals DROSHA is recruited to DNA damage sites by the MRN complex to promote non-homologous end-joining

2021 ◽  
pp. jcs.249706
Author(s):  
Matteo Cabrini ◽  
Marco Roncador ◽  
Alessandro Galbiati ◽  
Lina Cipolla ◽  
Antonio Maffia ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
End Joining ◽  
Strand Breaks ◽  
Mrn Complex ◽  
Break Site ◽  
Non Homologous End Joining ◽  
Dna Ends

The DNA damage response (DDR) is the signaling cascade that recognizes DNA double-strand breaks (DSB) and promotes their resolution via the DNA repair pathways of Non-Homologous End Joining (NHEJ) or Homologous Recombination (HR). We and others have shown that DDR activation requires DROSHA. However, whether DROSHA exerts its functions by associating with damage sites, what controls its recruitment and how DROSHA influences DNA repair, remains poorly understood. Here we show that DROSHA associates to DSBs independently from transcription. Neither H2AX, nor ATM nor DNA-PK kinase activities are required for its recruitment to break site. Rather, DROSHA interacts with RAD50 and inhibition of MRN by Mirin treatment abolishes this interaction. MRN inactivation by RAD50 knockdown or mirin treatment prevents DROSHA recruitment to DSB and, as a consequence, also 53BP1 recruitment. During DNA repair, DROSHA inactivation reduces NHEJ and boosts HR frequency. Indeed, DROSHA knockdown also increase the association of downstream HR factors such as RAD51 to DNA ends. Overall, our results demonstrate that DROSHA is recruited at DSBs by the MRN complex and direct DNA repair toward NHEJ.

CBIO-08. ENDOGENOUS DNA DOUBLE STRAND BREAKS ACTIVATE HETEROGENOUS DNA DAMAGE SIGNALING IN IDH1/2 MUTANT GLIOMAS

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi28-vi29
Author(s):  
Gaspar Kitange ◽  
Rachael Vaubel ◽  
Jann Sarkaria
Keyword(s):  
Dna Damage ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Isocitrate Dehydrogenase 1 ◽  
Strand Breaks ◽  
Dna Damage Signaling ◽  
Alternative Lengthening ◽  
Systematic Effort ◽  
Patient Specimen

Abstract Isocitrate dehydrogenase 1/2 (IDH1/2) mutations are common in astrocytic glioma and are frequently coupled with TP53 and ATRX mutations. Collectively, these alterations cause genomic instability leading to high basal DNA double strand breaks (DSBs). Understanding how IDH/TP53/ATRX mutant cells process endogenous DSBs may help exploit inhibitors of DNA damage response (DDR) for the treatment of patients with IDH mutant gliomas. Through systematic effort to uncover the mechanisms involved in repair of endogenous DSBs in IDH1/2 mutant GBMs, we have discovered that high basal phosphorylated DNA-PK (p-DNA-PK) was characteristic of an IDH1/TP53/ATRX mutant GBM164 patient derived xenograft (PDX) but not in another IDH1 mutant GBM196 PDX. Immunofluorescence (IF) studies in patient specimen from which GBM164 was derived showed that p-DNA-PK co-localized with g-H2AX, 53BP1 or H4K20me2 (but not p-RPA) the known surrogates of DSBs. In contrast, p-DNA-PK was absent in the patient specimen from which GBM196 was derived, which otherwise had equally intense g-H2AX immunostaining colocalized with p-RPA. An independent IF study involving 11 IDH1 wild-type (WT) and 11 IDH1 mutant GBM patient samples, the p-DNA-PK was observed in 3 (27%) of 11 IDH1 mutant samples while IDH1 WT tumors were negative for p-DNA-PK. A telomere specific fluorescence in situ hybridization (Tel-FISH) confirmed elevated alternative lengthening of telomere (ALT) activity in GBM196 (but not in GBM164) indicative of HR proficiency. Consistently, HR related genes, including BRCA1 and MRE11A, were found upregulated in ALT-positive GBM196 as compared to those in GBM164. Interestingly, ALT+ GBM196 cells were highly vulnerable to inhibitors of ATM and ATR pathways. In conclusion, IDH1/TP53/ATRX mutant gliomas can be subdivided into HR-mediated ALT-positive group, which repairs the endogenous DSBs by HR (e.g. GBM196) and an ALT-negative/p-DNA-PK group, which repairs DSBs by c-NHEJ (e.g. GBM164) and this subdivision can be developed as a prescient biomarker of sensitivity to DDR inhibitors.

Sign in / Sign up

Export Citation Format

Share Document