scholarly journals Inactivation of mouse Hus1 results in genomic instability and impaired responses to genotoxic stress

2000 ◽  
Vol 14 (15) ◽  
pp. 1886-1898 ◽  
Author(s):  
Robert S. Weiss ◽  
Tamar Enoch ◽  
Philip Leder
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Replication ◽  
Chromosomal Abnormalities ◽  
Genotoxic Stress ◽  
Eukaryotic Cell ◽  
Cell Cycle Checkpoint ◽  
Cell Cycle Checkpoints ◽  
Genome Damage

The eukaryotic cell cycle is overseen by regulatory mechanisms, termed checkpoints, that respond to DNA damage, mitotic spindle defects, and errors in the ordering of cell cycle events. The DNA replication and DNA damage cell cycle checkpoints of the fission yeastSchizosaccharomyces pombe require the hus1+(hydroxyurea sensitive) gene. To determine the role of the mouse homolog of hus1+ in murine development and cell cycle checkpoint function, we produced a targeted disruption of mouse Hus1. Inactivation of Hus1results in mid-gestational embryonic lethality due to widespread apoptosis and defective development of essential extra-embryonic tissues. DNA damage-inducible genes are up-regulated inHus1-deficient embryos, and primary cells fromHus1-null embryos contain increased spontaneous chromosomal abnormalities, suggesting that loss of Hus1 leads to an accumulation of genome damage. Embryonic fibroblasts lackingHus1 fail to proliferate in vitro, but inactivation ofp21 allows for the continued growth of Hus1-deficient cells.Hus1−/−p21−/−cells display a unique profile of significantly heightened sensitivity to hydroxyurea, a DNA replication inhibitor, and ultraviolet light, but only slightly increased sensitivity to ionizing radiation. Taken together, these results indicate that mouse Hus1 functions in the maintenance of genomic stability and additionally identify an evolutionarily-conserved role for Hus1 in mediating cellular responses to genotoxins.

scholarly journals Human TopBP1 Ensures Genome Integrity during Normal S Phase

2005 ◽  
Vol 25 (24) ◽  
pp. 10907-10915 ◽  
Author(s):  
Ja-Eun Kim ◽  
Sarah A. McAvoy ◽  
David I. Smith ◽  
Junjie Chen
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Replication ◽  
Critical Role ◽  
Genotoxic Stress ◽  
S Phase ◽  
Fragile Sites ◽  
Cell Cycle Checkpoints ◽  
Exogenous Dna ◽  
Hydroxyurea Treatment

ABSTRACT Cell cycle checkpoints are essential for maintaining genomic integrity. Human topoisomerase II binding protein 1 (TopBP1) shares sequence similarity with budding yeast Dpb11, fission yeast Rad4/Cut5, and Xenopus Cut5, all of which are required for DNA replication and cell cycle checkpoints. Indeed, we have shown that human TopBP1 participates in the activation of replication checkpoint and DNA damage checkpoints, following hydroxyurea treatment and ionizing radiation. In this study, we address the physiological function of TopBP1 in S phase by using small interfering RNA. In the absence of exogenous DNA damage, TopBP1 is recruited to replicating chromatin. However, TopBP1 does not appear to be essential for DNA replication. TopBP1-deficient cells have increased H2AX phosphorylation and ATM-Chk 2 activation, suggesting the accumulation of DNA double-strand breaks in the absence of TopBP1. This leads to formation of gaps and breaks at fragile sites, 4N accumulation, and aberrant cell division. We propose that the cellular function of TopBP1 is to monitor ongoing DNA replication. By ensuring proper DNA replication, TopBP1 plays a critical role in the maintenance of genomic stability during normal S phase as well as following genotoxic stress.

Reducing MCM Loading Causes Chromosomal Aneuploidy.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3349-3349
Author(s):  
Stephen J. Orr ◽  
Terry Gaymes ◽  
Rong Wang ◽  
Barbara Czepulkowski ◽  
Darius Ladon ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
T Cells ◽  
Dna Replication ◽  
Genomic Instability ◽  
Chromosomal Abnormalities ◽  
S Phase ◽  
Human T Cells ◽  
Dna Damage Responses ◽  
Mcm Proteins

Abstract Normal DNA replication must be accurate and occur only once per cell cycle. Sites of DNA replication are specified by binding the origin recognition complex, that includes minichromosome maintenance (MCM) proteins. Paradoxically, in higher eukaryotes MCM proteins are present in >20 fold excess of that required for DNA replication. They are also downregulated by elevated expression of proteins such as cyclin E that occurs in cancers, including AML and breast cancer. We investigated why human cells need “excess” MCM proteins and whether the reduction of MCM protein levels might contribute to a malignant phenotype. We determined the consequences of reducing the levels of MCM proteins in primary human T cells in which cell cycle controls and DNA damage responses are normal. Mass spectrometry sequencing of chromatin/nuclear matrix-bound proteins and western blotting identified that Mcm7 is not present in quiescent, normal primary human T cells. Mcm7 is induced in mid G1after the G0→G1 commitment point, the point beyond which T cells are committed to entering the cell cycle. Reduction of Mcm7 with siRNA to <5% of normal during G0→G1→S-phase reduces chromatin-binding of each of the MCM proteins that form the DNA helicase. However, these cells still enter S-phase and replicate DNA. Reducing MCM levels by titrating siRNA causes dose-dependent DNA-damage responses involving activation of ATR & ATM and Chk1 & Chk2. However, cells depleted of Mcm7 do not undergo apoptosis, rather reducing MCM levels even by 50% causes gross non-clonal chromosomal abnormalities normally found in genomic instability syndromes. M-FISH identified chromosome translocations, as well as loss and gain of individual chromosomes, which can occur individually or together in the same cell. Reducing MCM levels also causes misrepair by non-homologous end joining (NHEJ), and both NHEJ and homologous recombination (HR) are necessary for chromosomal abnormalities to occur. Therefore, “excess” MCM proteins that are present in a normal, proliferating cell are necessary for maintaining genome stability and reduction of MCM loading onto DNA that occurs in cancers is sufficient to cause genomic instability.

DNA damage and cell cycle checkpoints in hyperoxic lung injury: braking to facilitate repair

2001 ◽  
Vol 281 (2) ◽  
pp. L291-L305 ◽  
Author(s):  
Michael A. O'Reilly
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Genotoxic Stress ◽  
Supplemental Oxygen ◽  
Cell Cycle Checkpoints ◽  
Dna Integrity ◽  
Additional Time ◽  
Pulmonary Response ◽  
Arterial Blood ◽  
Hyperoxic Lung Injury

The beneficial use of supplemental oxygen therapies to increase arterial blood oxygen levels and reduce tissue hypoxia is offset by the knowledge that it injures and kills cells, resulting in increased morbidity and mortality. Although many studies have focused on understanding how hyperoxia kills cells, recent findings reveal that it also inhibits proliferation through activation of cell cycle checkpoints rather than through overt cytotoxicity. Cell cycle checkpoints are thought to be protective because they allow additional time for injured cells to repair damaged DNA and other essential molecules. During recovery in room air, the lung undergoes a burst of proliferation to replace injured and dead cells. Failure to terminate this proliferation has been associated with fibrosis. These observations suggest that growth-suppressive signals, which inhibit proliferation of injured cells and terminate proliferation when tissue repair has been completed, may play an important role in the pulmonary response to hyperoxia. Because DNA replication is coupled with DNA repair, activation of cell cycle checkpoints during hyperoxia may be a mechanism by which cells protect themselves from oxidant genotoxic stress. This review examines the effect of hyperoxia on DNA integrity, pulmonary cell proliferation, and cell cycle checkpoints activated by DNA damage.

scholarly journals LARP7 is a BRCA1 ubiquitinase substrate and regulates genome stability and tumorigenesis

2019 ◽  
Author(s):  
Fang Zhang ◽  
Pengyi Yan ◽  
Huijing Yu ◽  
Huangying Le ◽  
Zixuan li ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Rna Binding ◽  
Tumor Therapy ◽  
Genotoxic Stress ◽  
Therapy Resistance ◽  
List Type ◽  
M Cell

SummaryAttenuated DNA repair leads to genomic instability and tumorigenesis. BRCA1/BARD1 are the best known tumor suppressors that promote homology recombination (HR) and arrest cell cycle at G2/M checkpoint. As E3 ubiquitin ligases, their ubiquitinase activity has been known to involve in the HR and tumor suppression, but the mechanism remains ambiguous. Here, we demonstrated upon genotoxic stress, BRCA1 together with BARD1 catalyzed the K48 ployubiquitination on LARP7, a 7SK RNA binding protein known to control RNAPII pausing, and thereby degraded it through 26S ubiquitin-proteasome pathway. Depleting LARP7 suppressed the expression of CDK1 complex, arrested cell at G2/M DNA damage checkpoint and reduced BRCA2 phosphorylation which thereby facilitated RAD51 recruitment to damaged DNA to enhance HR. Importantly, LARP7 depletion observed in breast patients lead to the chemoradiotherapy resistance both in vitro and in vivo. Together, this study unveils a mechanism by which BRCA1/BARD1 utilizes their E3 ligase activity to control HR and cell cycle, and highlights LARP7 as a potential target for cancer prevention and therapy.HighlightsDNA damage response downregulates LARP7 through BRCA1/BARD1BRCA1/BARD1 catalyzes the K48 polyubiquitination on LARP7LARP7 promotes G2/M cell cycle transition and tumorigenesis via CDK1 complexLARP7 disputes homology-directed repair that leads to tumor therapy resistance

scholarly journals A kinetochore-based ATM/ATR-independent DNA damage checkpoint maintains genomic integrity in trypanosomes

2019 ◽  
Vol 47 (15) ◽  
pp. 7973-7988 ◽  
Author(s):  
Qing Zhou ◽  
Kieu T M Pham ◽  
Huiqing Hu ◽  
Yasuhiro Kurasawa ◽  
Ziyin Li
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Spindle Assembly Checkpoint ◽  
Cell Cycle Checkpoint ◽  
Cell Cycle Checkpoints ◽  
Genomic Integrity ◽  
Metaphase Arrest ◽  
M Cell ◽  
Independent Manner ◽  
Mitotic Cyclin

Abstract DNA damage-induced cell cycle checkpoints serve as surveillance mechanisms to maintain genomic stability, and are regulated by ATM/ATR-mediated signaling pathways that are conserved from yeast to humans. Trypanosoma brucei, an early divergent microbial eukaryote, lacks key components of the conventional DNA damage-induced G2/M cell cycle checkpoint and the spindle assembly checkpoint, and nothing is known about how T. brucei controls its cell cycle checkpoints. Here we discover a kinetochore-based, DNA damage-induced metaphase checkpoint in T. brucei. MMS-induced DNA damage triggers a metaphase arrest by modulating the abundance of the outer kinetochore protein KKIP5 in an Aurora B kinase- and kinetochore-dependent, but ATM/ATR-independent manner. Overexpression of KKIP5 arrests cells at metaphase through stabilizing the mitotic cyclin CYC6 and the cohesin subunit SCC1, mimicking DNA damage-induced metaphase arrest, whereas depletion of KKIP5 alleviates the DNA damage-induced metaphase arrest and causes chromosome mis-segregation and aneuploidy. These findings suggest that trypanosomes employ a novel DNA damage-induced metaphase checkpoint to maintain genomic integrity.

scholarly journals New Insights into p53 Signaling and Cancer Cell Response to DNA Damage: Implications for Cancer Therapy

2012 ◽  
Vol 2012 ◽  
pp. 1-16 ◽  
Author(s):  
Razmik Mirzayans ◽  
Bonnie Andrais ◽  
April Scott ◽  
David Murray
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Apoptotic Cell ◽  
Cell Types ◽  
Cell Cycle Checkpoint ◽  
Cell Cycle Checkpoints ◽  
Cancer Therapies ◽  
Wild Type ◽  
P53 Signaling ◽  
Upstream Kinase

Activation of the p53 signaling pathway by DNA-damaging agents was originally proposed to result either in cell cycle checkpoint activation to promote survival or in apoptotic cell death. This model provided the impetus for numerous studies focusing on the development of p53-based cancer therapies. According to recent evidence, however, most p53 wild-type human cell types respond to ionizing radiation by undergoing stress-induced premature senescence (SIPS) and not apoptosis. SIPS is a sustained growth-arrested state in which cells remain viable and secrete factors that may promote cancer growth and progression. Thep21WAF1(hereafter p21) protein has emerged as a key player in the p53 pathway. In addition to its well-studied role in cell cycle checkpoints, p21 regulates p53 and its upstream kinase (ATM), controls gene expression, suppresses apoptosis, and induces SIPS. Herein, we review these and related findings with human solid tumor-derived cell lines, report new data demonstrating dynamic behaviors of p53 and p21 in the DNA damage response, and examine the gain-of-function properties of cancer-associated p53 mutations. We point out obstacles in cancer-therapeutic strategies that are aimed at reactivating the wild-type p53 function and highlight some alternative approaches that target the apoptotic threshold in cancer cells with differing p53 status.

scholarly journals Cdc25A Serine 123 Phosphorylation Couples Centrosome Duplication with DNA Replication and Regulates Tumorigenesis

2008 ◽  
Vol 28 (24) ◽  
pp. 7442-7450 ◽  
Author(s):  
Sathyavageeswaran Shreeram ◽  
Weng Kee Hee ◽  
Dmitry V. Bulavin
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Replication ◽  
Cell Cycle Progression ◽  
Phosphorylation Site ◽  
Centrosome Duplication ◽  
Cycle Progression ◽  
Cdc25a Phosphatase

ABSTRACT The cell division cycle 25A (Cdc25A) phosphatase is a critical regulator of cell cycle progression under normal conditions and after stress. Stress-induced degradation of Cdc25A has been proposed as a major way of delaying cell cycle progression. In vitro studies pointed toward serine 123 as a key site in regulation of Cdc25A stability after exposure to ionizing radiation (IR). To address the role of this phosphorylation site in vivo, we generated a knock-in mouse in which alanine was substituted for serine 123. The Cdc25 S123A knock-in mice appeared normal, and, unexpectedly, cells derived from them exhibited unperturbed cell cycle and DNA damage responses. In turn, we found that Cdc25A was present in centrosomes and that Cdc25A levels were not reduced after IR in knock-in cells. This resulted in centrosome amplification due to lack of induction of Cdk2 inhibitory phosphorylation after IR specifically in centrosomes. Further, Cdc25A knock-in animals appeared sensitive to IR-induced carcinogenesis. Our findings indicate that Cdc25A S123 phosphorylation is crucial for coupling centrosome duplication to DNA replication cycles after DNA damage and therefore is likely to play a role in the regulation of tumorigenesis.

DNA replication and damage checkpoints and meiotic cell cycle controls in the fission and budding yeasts

2000 ◽  
Vol 349 (1) ◽  
pp. 1-12 ◽  
Author(s):  
Hiroshi MURAKAMI ◽  
Paul NURSE
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Replication ◽  
Fission Yeast ◽  
Molecular Mechanisms ◽  
Cell Cycle Checkpoint ◽  
Cell Cycle Regulators ◽  
Meiotic Cell ◽  
Checkpoint Proteins ◽  
Dna Damage Checkpoints

The cell cycle checkpoint mechanisms ensure the order of cell cycle events to preserve genomic integrity. Among these, the DNA-replication and DNA-damage checkpoints prevent chromosome segregation when DNA replication is inhibited or DNA is damaged. Recent studies have identified an outline of the regulatory networks for both of these controls, which apparently operate in all eukaryotes. In addition, it appears that these checkpoints have two arrest points, one is just before entry into mitosis and the other is prior to chromosome separation. The former point requires the central cell-cycle regulator Cdc2 kinase, whereas the latter involves several key regulators and substrates of the ubiquitin ligase called the anaphase promoting complex. Linkages between these cell-cycle regulators and several key checkpoint proteins are beginning to emerge. Recent findings on post-translational modifications and protein-protein interactions of the checkpoint proteins provide new insights into the checkpoint responses, although the functional significance of these biochemical properties often remains unclear. We have reviewed the molecular mechanisms acting at the DNA-replication and DNA-damage checkpoints in the fission yeast Schizosaccharomyces pombe, and the modifications of these controls during the meiotic cell cycle. We have made comparisons with the controls in fission yeast and other organisms, mainly the distantly related budding yeast.

scholarly journals Critical role of SMG7 in activation of the ATR-CHK1 axis in response to genotoxic stress

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Kathleen Ho ◽  
Hongwei Luo ◽  
Wei Zhu ◽  
Yi Tang
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Replication ◽  
Cell Cycle Progression ◽  
Critical Role ◽  
Genotoxic Stress ◽  
Dna Damage Checkpoint ◽  
Cycle Progression ◽  
Essential Step

AbstractCHK1 is a crucial DNA damage checkpoint kinase and its activation, which requires ATR and RAD17, leads to inhibition of DNA replication and cell cycle progression. Recently, we reported that SMG7 stabilizes and activates p53 to induce G1 arrest upon DNA damage; here we show that SMG7 plays a critical role in the activation of the ATR-CHK1 axis. Following genotoxic stress, SMG7-null cells exhibit deficient ATR signaling, indicated by the attenuated phosphorylation of CHK1 and RPA32, and importantly, unhindered DNA replication and fork progression. Through its 14-3-3 domain, SMG7 interacts directly with the Ser635-phosphorylated RAD17 and promotes chromatin retention of the 9-1-1 complex by the RAD17-RFC, an essential step to CHK1 activation. Furthermore, through maintenance of CHK1 activity, SMG7 controls G2-M transition and facilitates orderly cell cycle progression during recovery from replication stress. Taken together, our data reveals SMG7 as an indispensable signaling component in the ATR-CHK1 pathway during genotoxic stress response.

scholarly journals Cell cycle checkpoints cooperate to suppress DNA and RNA associated molecular pattern recognition and anti-tumor immune responses

2020 ◽  
Author(s):  
Jie Chen ◽  
Shane M Harding ◽  
Ramakrishnan Natesan ◽  
Lei Tian ◽  
Joseph L Benci ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Pattern Recognition ◽  
Dna Damage ◽  
Immune Responses ◽  
Genotoxic Stress ◽  
Cell Cycle Checkpoints ◽  
Dependent Manner ◽  
Dna And Rna ◽  
Molecular Pattern ◽  
Immune Mediated

SummaryThe DNA dependent pattern recognition receptor, cGAS mediates communication between genotoxic stress and the immune system. Mitotic chromosome missegregation is an established stimulator of cGAS activity, however, it is unclear if progression through mitosis is required for cancer cell intrinsic activation of immune mediated anti-tumor responses. Moreover, it is unknown if disruption of cell cycle checkpoints can restore responses in cancer cells that are recalcitrant to DNA damage induced inflammation. Here we demonstrate that prolonged cell cycle arrest at the G2-mitosis boundary from either CDK1 inhibition or excessive DNA damage prevents inflammatory stimulated gene expression and immune mediated destruction of distal tumors. Remarkably, DNA damage induced inflammatory signaling is restored in a cGAS-and RIG-I-dependent manner upon concomitant disruption of p53 and the G2 checkpoint. These findings link aberrant cell progression and p53 loss to an expanded spectrum of damage associated molecular pattern recognition and have implications for the design of rational approaches to augment antitumor immune responses.

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