scholarly journals MDC1 regulates intra-S-phase checkpoint by targeting NBS1 to DNA double-strand breaks

2008 ◽  
Vol 105 (32) ◽  
pp. 11200-11205 ◽  
Author(s):  
L. Wu ◽  
K. Luo ◽  
Z. Lou ◽  
J. Chen
Keyword(s):  
S Phase ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
S Phase Checkpoint

scholarly journals Mitotic cells can repair DNA double-strand breaks via a homology-directed pathway

2020 ◽  
Vol 62 (1) ◽  
pp. 25-33
Author(s):  
Yuki Sakamoto ◽  
Tetsuya Kokuta ◽  
Ai Teshigahara ◽  
Kenta Iijima ◽  
Hiroyuki Kitao ◽  
...  
Keyword(s):  
Mitotic Cell ◽  
S Phase ◽  
Double Strand Breaks ◽  
G1 Phase ◽  
Dna Double Strand Breaks ◽  
Major Pathway ◽  
Strand Breaks ◽  
Homology Directed Repair ◽  
Repair Efficiency ◽  
M Phase

Abstract The choice of repair pathways of DNA double-strand breaks (DSBs) is dependent upon the cell cycle phases. While homologous recombination repair (HRR) is active between the S and G2 phases, its involvement in mitotic DSB repair has not been examined in detail. In the present study, we developed a new reporter assay system to detect homology-directed repair (HDR), a major pathway used for HRR, in combination with an inducible DSB-generation system. As expected, the maximal HDR activity was observed in the late S phase, along with minimal activity in the G1 phase and at the G1/S boundary. Surprisingly, significant HDR activity was observed in M phase, and the repair efficiency was similar to that observed in late S phase. HDR was also confirmed in metaphase cells collected with continuous colcemid exposure. ChIP assays revealed the recruitment of RAD51 to the vicinity of DSBs in M phase. In addition, the ChIP assay for gamma-H2AX and phosphorylated DNA-PKcs indicated that a part of M-phase cells with DSBs could proceed into the next G1 phase. These results provide evidence showing that a portion of mitotic cell DSBs are undoubtedly repaired through action of the HDR repair pathway.

scholarly journals WRN Is Required for ATM Activation and the S-Phase Checkpoint in Response to Interstrand Cross-Link–Induced DNA Double-Strand Breaks

2008 ◽  
Vol 19 (9) ◽  
pp. 3923-3933 ◽  
Author(s):  
Wen-Hsing Cheng ◽  
Diana Muftic ◽  
Meltem Muftuoglu ◽  
Lale Dawut ◽  
Christa Morris ◽  
...  
Keyword(s):  
Kinase Inhibitor ◽  
Werner Syndrome ◽  
Genetic Disorder ◽  
S Phase ◽  
Double Strand Breaks ◽  
Premature Aging ◽  
Cross Link ◽  
Strand Breaks ◽  
In Cells ◽  
S Phase Checkpoint

Werner syndrome (WS) is a human genetic disorder characterized by extensive clinical features of premature aging. Ataxia-telengiectasia (A-T) is a multisystem human genomic instability syndrome that includes premature aging in some of the patients. WRN and ATM, the proteins defective in WS and A-T, respectively, play significant roles in the maintenance of genomic stability and are involved in several DNA metabolic pathways. A role for WRN in DNA repair has been proposed; however, this study provides evidence that WRN is also involved in ATM pathway activation and in a S-phase checkpoint in cells exposed to DNA interstrand cross-link–induced double-strand breaks. Depletion of WRN in such cells by RNA interference results in an intra-S checkpoint defect, and interferes with activation of ATM as well as downstream phosphorylation of ATM target proteins. Treatment of cells under replication stress with the ATM kinase inhibitor KU 55933 results in a S-phase checkpoint defect similar to that observed in WRN shRNA cells. Moreover, γH2AX levels are higher in WRN shRNA cells than in control cells 6 and 16 h after exposure to psoralen DNA cross-links. These results suggest that WRN and ATM participate in a replication checkpoint response, in which WRN facilitates ATM activation in cells with psoralen DNA cross-link–induced collapsed replication forks.

ANKHD1 Silencing Delays S Phase Progression in Multiple Myeloma Cells Via Activation of ATM/ATR -CDC25a Pathway

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5624-5624
Author(s):  
Dhyani Anamika ◽  
Patricia Favaro ◽  
Sara Teresinha Olalla Saad
Keyword(s):  
Cell Cycle ◽  
Multiple Myeloma ◽  
Dna Damage ◽  
S Phase ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Myeloma Cells ◽  
Checkpoint Kinase ◽  
Strand Breaks ◽  
Phase Progression

Abstract Ankyrin repeat and KH domain-containing protein 1, ANKHD1, is highly expressed in myeloma cells and plays an important role in multiple myeloma (MM) progression and growth. ANKHD1 is found to be overexpressed in S phase of cell cycle in MM cells and silencing of ANKHD1 expression leads to accumulation of cells in S phase, suggesting a role in S phase progression (1). Earlier studies by our group reported that ANKHD1 silencing downregulates all replication dependent histones and that this downregulation may be associated with replication stress and DNA damage (2). We observed increased expression of γH2AX protein (phosphorylated histone H2A variant, H2AX, at Serine 139), a marker for DNA double strand breaks (DSBs) and an early sign of DNA damage induced by replication stress, in ANKHD1 silenced MM cells. In the present study we further sought to investigate the mechanisms underlying the induction of DNA damage on ANKHD1 silencing. We first confirmed the increased expression of γH2AX by flow cytometry analysis and observed that both the mean fluorescence intensity as well as percentage of γH2AX positive cells were higher in ANKHD1 silenced MM cells as compared to control cells. Phosphorylation of histone 2AX requires activation of the phosphatidylinositol-3-OH-kinase-like family of protein kinases, DNA-PKcs (DNA-dependent protein kinase), ATM (ataxia telangiectasia mutated)andATR (ATM-Rad3-related) that serves as central components of the signaling cascade initiated by DSBs. Hence, we checked for the expression of these kinases and observed increased phosphorylation of both ATM and ATR kinases in ANKHD1 silenced MM cells. There was no difference in the expressions of DNA-PKcs in control and ANKHD1 silenced cells by western blot. We next checked for the expression of CHK1 (checkpoint kinase 1) and CHK2 (checkpoint kinase 2), essential serine threonine kinases downstream of ATM and ATR. We observed a decrease in pCHK2 (phosphorylated CHK2 at Thr 68), with no change in expression of pCHK1 (phosphorylated CHK1 at Ser 345) total CHK1 or total CHK2. We also checked for expression of CDC25a (a member of the CDC25 family of dual-specificity phosphatases), that is specifically degraded in response to DNA damage (DSBs) and delays S phase progression via activation of ATM /ATR-CHK2 signaling pathway. Expression of CDC25a was significantly decreased in ANKHD1 silencing cells, confirming the induction of DSBs, and probably accounting for S phase delay on ANKHD1 silencing. Since there was decrease in active CHK2 (pCHK2) and no change in CHK1 required for degradation of CDC25a, we assume that decrease in CDC25a in ANKHD1 silenced MM cells may be via activation of ATM/ ATR pathway independent of CHK2/CHK1. Expression of several other downstream factors of DSBs induced DNA damage response and repair such as BRCA1, PTEN, DNMT1, SP1, HDAC2 were also found to be modulated in ANKHD1 silenced MM cells. In conclusion, ANKHD1 silencing in MM cells leads to DNA damage and modulates expression of several genes implicated in DNA damage and repair. DNA damage induced after ANKHD1 silencing in MM cells activates ATM/ ATR-CDC25a pathway which may lead to the activation of S phase checkpoint in MM cells. Results however are preliminary and further studies are required to understand the role of ANKHD1 in intra S phase check point. References: 1) ANKHD1 regulates cell cycle progression and proliferation in multiple myeloma cells. Dhyani et al. FEBS letters 2012; 586: 4311-18. 2) ANKHD1 is essential for repair of DNA double strand breaks in multiple myeloma. Dhyani et al. ASH Abstract, Blood 2015; 126:1762. Disclosures No relevant conflicts of interest to declare.

scholarly journals A minority of foci or pan-nuclear apoptotic staining of  H2AX in the S phase after UV damage contain DNA double-strand breaks

2010 ◽  
Vol 107 (15) ◽  
pp. 6870-6875 ◽  
Author(s):  
S. de Feraudy ◽  
I. Revet ◽  
V. Bezrookove ◽  
L. Feeney ◽  
J. E. Cleaver
Keyword(s):  
S Phase ◽  
Double Strand Breaks ◽  
Uv Damage ◽  
Dna Double Strand Breaks ◽  
Strand Breaks

scholarly journals DNA damage triggers increased mobility of chromosomes in G1-phase cells

2019 ◽  
Vol 30 (21) ◽  
pp. 2620-2625 ◽  
Author(s):  
Michael J. Smith ◽  
Eric E. Bryant ◽  
Fraulin J. Joseph ◽  
Rodney Rothstein
Keyword(s):  
Dna Damage ◽  
S Phase ◽  
Double Strand Breaks ◽  
Homology Search ◽  
G1 Phase ◽  
Dna Damage Checkpoint ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Global Mobility ◽  
Break Site

During S phase in Saccharomyces cerevisiae, chromosomal loci become mobile in response to DNA double-strand breaks both at the break site (local mobility) and throughout the nucleus (global mobility). Increased nuclear exploration is regulated by the recombination machinery and the DNA damage checkpoint and is likely an important aspect of homology search. While mobility in response to DNA damage has been studied extensively in S phase, the response in interphase has not, and the question of whether homologous recombination proceeds to completion in G1 phase remains controversial. Here, we find that global mobility is triggered in G1 phase. As in S phase, global mobility in G1 phase is controlled by the DNA damage checkpoint and the Rad51 recombinase. Interestingly, despite the restriction of Rad52 mediator foci to S phase, Rad51 foci form at high levels in G1 phase. Together, these observations indicate that the recombination and checkpoint machineries promote global mobility in G1 phase, supporting the notion that recombination can occur in interphase diploids.

scholarly journals The scaffold protein Nde1 safeguards the brain genome during S phase of early neural progenitor differentiation

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Shauna L Houlihan ◽  
Yuanyi Feng
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Developmental Disorders ◽  
Neural Progenitor ◽  
Cortical Layer ◽  
S Phase ◽  
Double Strand Breaks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Genomic Disorders

Successfully completing the S phase of each cell cycle ensures genome integrity. Impediment of DNA replication can lead to DNA damage and genomic disorders. In this study, we show a novel function for NDE1, whose mutations cause brain developmental disorders, in safeguarding the genome through S phase during early steps of neural progenitor fate restrictive differentiation. Nde1 mutant neural progenitors showed catastrophic DNA double strand breaks concurrent with the DNA replication. This evoked DNA damage responses, led to the activation of p53-dependent apoptosis, and resulted in the reduction of neurons in cortical layer II/III. We discovered a nuclear pool of Nde1, identified the interaction of Nde1 with cohesin and its associated chromatin remodeler, and showed that stalled DNA replication in Nde1 mutants specifically occurred in mid-late S phase at heterochromatin domains. These findings suggest that NDE1-mediated heterochromatin replication is indispensible for neuronal differentiation, and that the loss of NDE1 function may lead to genomic neurological disorders.

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