scholarly journals Replication stress induces mitotic death through parallel pathways regulated by WAPL and telomere deprotection

2018 ◽  
Author(s):  
V. Pragathi Masamsetti ◽  
Ka Sin Mak ◽  
Ronnie Ren Jie Low ◽  
Chris D. Riffkin ◽  
Noa Lamm ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Spindle Assembly Checkpoint ◽  
Genome Instability ◽  
Replication Stress ◽  
Mitotic Arrest ◽  
Mitotic Catastrophe ◽  
Aurora B ◽  
Mitotic Death ◽  
Stressed Cells

ABSTRACTMitotic catastrophe is a broad descriptor encompassing unclear mechanisms of cell death. Here we investigate replication stress-driven mitotic catastrophe in human cells and identify that replication stress principally induces mitotic death signalled through two independent pathways. In p53-compromised cells we find that lethal replication stress confers WAPL-dependent centromere cohesion defects that maintain spindle assembly checkpoint-dependent mitotic arrest in the same cell cycle. Mitotic arrest then drives cohesion fatigue and triggers mitotic death through a primary pathway of BAX/BAK-dependent apoptosis. Simultaneously, a secondary mitotic death pathway is engaged through non-canonical telomere deprotection, regulated by TRF2, Aurora B and ATM. Additionally, we find that suppressing mitotic death promotes genome instability in replication stressed cells through diverse mechanisms depending upon how cell death is averted. These data demonstrate how replication stress-induced mitotic catastrophe signals cell death with implications for cancer treatment and genome instability.

scholarly journals Replication stress induces mitotic death through parallel pathways regulated by WAPL and telomere deprotection

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
V. Pragathi Masamsetti ◽  
Ronnie Ren Jie Low ◽  
Ka Sin Mak ◽  
Aisling O’Connor ◽  
Chris D. Riffkin ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Cancer Treatment ◽  
Spindle Assembly Checkpoint ◽  
Replication Stress ◽  
Mitotic Arrest ◽  
Mitotic Catastrophe ◽  
Aurora B ◽  
Mitotic Death ◽  
Stressed Cells

Abstract Mitotic catastrophe is a broad descriptor encompassing unclear mechanisms of cell death. Here we investigate replication stress-driven mitotic catastrophe in human cells and identify that replication stress principally induces mitotic death signalled through two independent pathways. In p53-compromised cells we find that lethal replication stress confers WAPL-dependent centromere cohesion defects that maintain spindle assembly checkpoint-dependent mitotic arrest in the same cell cycle. Mitotic arrest then drives cohesion fatigue and triggers mitotic death through a primary pathway of BAX/BAK-dependent apoptosis. Simultaneously, a secondary mitotic death pathway is engaged through non-canonical telomere deprotection, regulated by TRF2, Aurora B and ATM. Additionally, we find that suppressing mitotic death in replication stressed cells results in distinct cellular outcomes depending upon how cell death is averted. These data demonstrate how replication stress-induced mitotic catastrophe signals cell death with implications for cancer treatment and cancer genome evolution.

scholarly journals Antagonizing the spindle assembly checkpoint silencing enhances paclitaxel and Navitoclax-mediated apoptosis with distinct mechanistic

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ana C. Henriques ◽  
Patrícia M. A. Silva ◽  
Bruno Sarmento ◽  
Hassan Bousbaa
Keyword(s):  
Cell Death ◽  
Cancer Cells ◽  
Cell Fate ◽  
Spindle Assembly Checkpoint ◽  
Spindle Assembly ◽  
Time Lapse ◽  
Antimitotic Drugs ◽  
Mitotic Slippage ◽  
Mitotic Death ◽  
Chronic Activation

AbstractAntimitotic drugs arrest cells in mitosis through chronic activation of the spindle assembly checkpoint (SAC), leading to cell death. However, drug-treated cancer cells can escape death by undergoing mitotic slippage, due to premature mitotic exit. Therefore, overcoming slippage issue is a promising chemotherapeutic strategy to improve the effectiveness of antimitotics. Here, we antagonized SAC silencing by knocking down the MAD2-binding protein p31comet, to delay mitotic slippage, and tracked cancer cells treated with the antimitotic drug paclitaxel, over 3 days live-cell time-lapse analysis. We found that in the absence of p31comet, the duration of mitotic block was increased in cells challenged with nanomolar concentrations of paclitaxel, leading to an additive effects in terms of cell death which was predominantly anticipated during the first mitosis. As accumulation of an apoptotic signal was suggested to prevent mitotic slippage, when we challenged p31comet-depleted mitotic-arrested cells with the apoptosis potentiator Navitoclax (previously called ABT-263), cell fate was shifted to accelerated post-mitotic death. We conclude that inhibition of SAC silencing is critical for enhancing the lethality of antimitotic drugs as well as that of therapeutic apoptosis-inducing small molecules, with distinct mechanisms. The study highlights the potential of p31comet as a target for antimitotic therapies.

scholarly journals Uniform Widespread Nuclear Phosphorylation of Histone H2AX Is an Indicator of Lethal DNA Replication Stress

Cancers ◽  
2019 ◽  
Vol 11 (3) ◽  
pp. 355 ◽  
Author(s):  
Eric Moeglin ◽  
Dominique Desplancq ◽  
Sascha Conic ◽  
Mustapha Oulad-Abdelghani ◽  
Audrey Stoessel ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Cancer Cells ◽  
Drug Efficacy ◽  
Replication Stress ◽  
Strand Break ◽  
Histone H2ax ◽  
C Terminus ◽  
Dna Replication Stress ◽  
Stressed Cells

Phosphorylated histone H2AX (γ-H2AX), a central player in the DNA damage response (DDR), serves as a biomarker of DNA double-strand break repair. Although DNA damage is generally visualized by the formation of γ-H2AX foci in injured nuclei, it is unclear whether the widespread uniform nuclear γ-H2AX (called pan-nuclear) pattern occurring upon intense replication stress (RS) is linked to DDR. Using a novel monoclonal antibody that binds exclusively to the phosphorylated C-terminus of H2AX, we demonstrate that H2AX phosphorylation is systematically pan-nuclear in cancer cells stressed with RS-inducing drugs just before they die. The pan-nuclear γ-H2AX pattern is abolished by inhibition of the DNA-PK kinase. Cell death induction of cancer cells treated with increasing combinations of replication and kinase (ATR and Chk1) inhibitory drugs was proportional to the appearance of pan-nuclear γ-H2AX pattern. Delivery of labeled anti-γ-H2AX Fabs in stressed cells demonstrated at a single cell level that pan-nuclear γ-H2AX formation precedes irreversible cell death. Moreover, we show that H2AX is not required for RS-induced cell death in HeLa cells. Thus, the nuclear-wide formation of γ-H2AX is an incident of RS-induced cell death and, thus, the pan nuclear H2AX pattern should be regarded as an indicator of lethal RS-inducing drug efficacy.

scholarly journals Involvement of p53 in cell death following cell cycle arrest and mitotic catastrophe induced by rotenone

2011 ◽  
Vol 1813 (3) ◽  
pp. 492-499 ◽  
Author(s):  
António Pedro Gonçalves ◽  
Valdemar Máximo ◽  
Jorge Lima ◽  
Keshav K. Singh ◽  
Paula Soares ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Death ◽  
Cell Cycle Arrest ◽  
Mitotic Catastrophe ◽  
Cycle Arrest

scholarly journals C1orf109L promote R-loop accumulation induced DNA damage to inhibit cell growth

2019 ◽  
Author(s):  
Dou Peng ◽  
Li Yiqun ◽  
Xie Wanqiu ◽  
Zhang Xiaoqing ◽  
Zhang Dandan ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Cell Death ◽  
Cell Growth ◽  
Genome Instability ◽  
Cell Death Pathway ◽  
Unknown Gene ◽  
Lower Expression

AbstractAs a function unknown gene, C1orf109 is lower expression in various cells. Here, we reported that C1orf109L, the longest variant of C1orf109, which interacted with R-loop-regulating proteins to trigger R-loop, a three-stranded nucleic acid structure frequently mediated genome instability, accumulation. C1orf109L induce chronic DNA damage to promote P21 upregulation and strongly inhibits cell growth in vitro and in vivo by arresting the cell cycle in the G2 phase. With camptothecin (CPT), an R-loop activator, treatment, C1orf109L further triggers R-loop accumulation-induced DNA damage and promotes cell death by activating cell-death pathway. Furthermore, CPT treatment increases C1orf109L ubiquitination and turnover, which inhibits cell death and promotes the G0/G1 phase of the cell cycle. Therefore, our data illustrated the mechanisms underlying C1orf109L-related cell growth inhibition and provide feasibility and limitations for C1orf109L as a potential target for cancer therapy.

scholarly journals Growth hormone induces mitotic catastrophe of podocytes and contributes to proteinuria

2019 ◽  
Author(s):  
Rajkishor Nishad ◽  
Dhanunjay Mukhi ◽  
Ashish Kumar Singh ◽  
Kumaraswami Chintala ◽  
Prasad Tammineni ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Growth Hormone ◽  
Cell Death ◽  
Diabetic Nephropathy ◽  
Notch Signaling ◽  
Mitotic Catastrophe ◽  
Podocyte Injury ◽  
Tgf Β1 ◽  
Notch Activation

AbstractPodocytes are integral members of the filtration barrier in the kidney and are crucial for glomerular permselectivity. Podocytes are highly differentiated and vulnerable to an array of noxious stimuli during various clinical conditions whereas podocyte loss plays a key role in progressive glomerular diseases. Elevated circulating growth hormone (GH) levels are associated with podocyte injury and proteinuria in diabetics. Previous studies have shown that podocytes express GH receptors (GHR), and induce Notch signaling when exposed to GH. However, the precise mechanism(s) by which excess GH elicits podocytopathy remains to be elucidated. In the present study, we demonstrate that GH induces cognate TGF-β1 signaling and provokes cell cycle re-entry of otherwise quiescent podocytes. Though, differentiated podocytes re-enter the cell cycle in response to GH and TGF-β1 unable to accomplish cytokinesis, despite nuclear division. Owing to this aberrant cell-cycle events significant amount of GH or TGF-β1 treated cells remain binucleated and undergo mitotic catastrophe. Importantly, inhibition of GHR, TGFBR1, or Notch signaling prevented cell cycle re-entry and protects podocyte from cell death. Furthermore, inhibition of Notch activation prevents GH-dependent podocyte injury and proteinuria. Kidney biopsy sections from patients with diabetic nephropathy show activation of Notch signaling and bi-nucleated podocytes. All these data confirm that excess GH induces Notch1 signaling via TGF-β1 and contributes to the mitotic catastrophe of podocytes. This study highlights the role of aberrant GH signaling in the podocytopathy and the potential application of inhibitors of TGF-β1 or Notch inhibitors as a therapeutic agent for diabetic nephropathy.Significance StatementElevated circulating levels of growth hormone (GH) associated with glomerular hypertrophy and proteinuria. Whereas decreased GH action protected against proteinuria. Podocytes are highly differentiated cells that play a vital role in glomerular filtration and curb protein loss. The direct role of GH in podocytes is the focus of our study. We found that GH induces TGF-β1 and both provoke cell cycle re-entry of podocytes in Notch1 dependent manner. Notch activation enables the podocytes to accomplish karyokinesis, but not cytokinesis owing to which podocytes remain binucleated. Binucleated podocytes that were observed during GH/TGF-β1 treatment are susceptible to cell death. Our study highlighted the fact that enforcing the differentiated podocytes to re-enter the cell cycle results in mitotic catastrophe and permanent loss.

scholarly journals Polyomavirus Small T Antigen Induces Apoptosis in Mammalian Cells through the UNC5B Pathway in a PP2A-Dependent Manner

2020 ◽  
Vol 94 (14) ◽  
Author(s):  
Sameer Ahmed Bhat ◽  
Zarka Sarwar ◽  
Syed Qaaifah Gillani ◽  
Misbah Un Nisa ◽  
Irfana Reshi ◽  
...  
Keyword(s):  
Cell Death ◽  
Mammalian Cells ◽  
Mitotic Arrest ◽  
T Antigen ◽  
Mitotic Catastrophe ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Antibody Treatment ◽  
Small T Antigen ◽  
Apoptotic Activity

ABSTRACT UNC5B is a dependence receptor that promotes survival in the presence of its ligand, netrin-1, while inducing cell death in its absence. The receptor has an important role in the development of the nervous and vascular systems. It is also involved in the normal turnover of intestinal epithelium. Netrin-1 and UNC5B are deregulated in multiple cancers, including colorectal, neuroblastoma, and breast tumors. However, the detailed mechanism of UNC5B function is not fully understood. We have utilized the murine polyomavirus small T antigen (PyST) as a tool to study UNC5B-mediated apoptosis. PyST is known to induce mitotic arrest followed by extensive cell death in mammalian cells. Our results show that the expression of PyST increases mRNA levels of UNC5B by approximately 3-fold in osteosarcoma cells (U2OS) and also stabilizes UNC5B at the posttranslational level. Furthermore, UNC5B is upregulated predominantly in those cells that undergo mitotic arrest upon PyST expression. Interestingly, although its expression was previously reported to be regulated by p53, our data show that the increase in UNC5B levels by PyST is p53 independent. The posttranslational stabilization of UNC5B by PyST is regulated by the interaction of PyST with PP2A. We also show that netrin-1 expression, which is known to inhibit UNC5B apoptotic activity, promotes survival of PyST-expressing cells. Our results thus suggest an important role of UNC5B in small-T antigen-induced mitotic catastrophe that also requires PP2A. IMPORTANCE UNC5B, PP2A, and netrin-1 are deregulated in a variety of cancers. UNC5B and PP2A are regarded as tumor suppressors, as they promote apoptosis and are deleted or mutated in many cancers. In contrast, netrin-1 promotes survival by inhibiting dependence receptors, including UNC5B, and is upregulated in many cancers. Here, we show that UNC5B-mediated apoptosis can occur independently of p53 but in a PP2A-dependent manner. A substantial percentage of cancers arise due to p53 mutations and are insensitive to chemotherapeutic treatments that activate p53. Unexpectedly, treatment of cancers having functional p53 with many conventional drugs leads to the upregulation of netrin-1 through activated p53, which is counterintuitive. Therefore, understanding the p53-independent mechanisms of the netrin-UNC5B axis, such as those involving PP2A, assumes greater clinical significance. Anticancer strategies utilizing anti-netrin-1 antibody treatment are already in clinical trials.

scholarly journals Intrinsic checkpoint deficiency during cell cycle re-entry from quiescence

2019 ◽  
Vol 218 (7) ◽  
pp. 2169-2184 ◽  
Author(s):  
Jacob Peter Matson ◽  
Amy M. House ◽  
Gavin D. Grant ◽  
Huaitong Wu ◽  
Joanna Perez ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Live Cell Imaging ◽  
Genome Instability ◽  
Replication Stress ◽  
S Phase ◽  
Minichromosome Maintenance ◽  
Cell Cycle Entry ◽  
Origin Licensing ◽  
Dna Replication Origins ◽  
Phase Entry

To maintain tissue homeostasis, cells transition between cell cycle quiescence and proliferation. An essential G1 process is minichromosome maintenance complex (MCM) loading at DNA replication origins to prepare for S phase, known as origin licensing. A p53-dependent origin licensing checkpoint normally ensures sufficient MCM loading before S phase entry. We used quantitative flow cytometry and live cell imaging to compare MCM loading during the long first G1 upon cell cycle entry and the shorter G1 phases in the second and subsequent cycles. We discovered that despite the longer G1 phase, the first G1 after cell cycle re-entry is significantly underlicensed. Consequently, the first S phase cells are hypersensitive to replication stress. This underlicensing results from a combination of slow MCM loading with a severely compromised origin licensing checkpoint. The hypersensitivity to replication stress increases over repeated rounds of quiescence. Thus, underlicensing after cell cycle re-entry from quiescence distinguishes a higher-risk first cell cycle that likely promotes genome instability.

scholarly journals PP2A-Cdc55 is responsible for mitotic arrest in DNA re-replicating cells in S. cerevisiae

2020 ◽  
Author(s):  
Shoily Khondker ◽  
Amy E. Ikui
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Spindle Assembly Checkpoint ◽  
Mitotic Arrest ◽  
Cell Cycle Checkpoints ◽  
Genome Integrity ◽  
Cyclin Dependent Kinase ◽  
Metaphase Arrest ◽  
Daughter Cells ◽  
Origin Licensing

AbstractThe cell cycle is an ordered process in which cells replicate their DNA in S-phase and divide them into two identical daughter cells in mitosis. DNA replication takes place only once per cell cycle to preserve genome integrity, which is tightly regulated by Cyclin Dependent Kinase (CDK). Formation of the pre-replicative complex, a platform for origin licensing, is inhibited through CDK-dependent phosphorylation. Failure of this control leads to re-licensing, re-replication and DNA damage. Eukaryotic cells have evolved surveillance mechanisms to maintain genome integrity, termed cell cycle checkpoints. It has been shown that the DNA damage checkpoint is activated upon the induction of DNA re-replication and arrests cell cycle in mitosis in S. cerevisiae. In this study, we show that PP2A-Cdc55 is responsible for the metaphase arrest induced by DNA re-replication, leading to dephosphorylation of APC component, Exclusion of Cdc55 from the nucleus bypassed the mitotic arrest and resulted in enhanced cell lethality in re-replicating cells. The metaphase arrest in re-replication cells was retained in the absence of Mad2, a key component of the spindle assembly checkpoint. Moreover, re-replicating cells showed the same rate of DNA damage induction in the presence or absence of Cdc55. These results indicate that PP2A-Cdc55 maintains metaphase arrest upon DNA re-replication and DNA damage through APC inhibition.

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