scholarly journals LIM Protein Ajuba associates with the RPA complex through direct and cell cycle-dependent interaction with the RPA70 subunit

2017 ◽  
Author(s):  
Sandy Fowler ◽  
Pascal Maguin ◽  
Sampada Kalan ◽  
Diego Loayza
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Genome Stability ◽  
Replication Stress ◽  
Cellular Transformation ◽  
Early Event ◽  
Lim Protein ◽  
Show Evidence ◽  
Dna Replication Stress ◽  
Cycle Dependent

AbstractDNA damage response pathways are essential for genome stability and cell survival. Specifically, the ATR kinase is activated by DNA replication stress. An early event in this activation is the recruitment and phosphorylation of RPA, a single stranded DNA binding complex composed of three subunits, RPA70,RPA32 and RPA14. We have previously shown that the LIM protein Ajuba associates with RPA, and that depletion of Ajuba leads to potent activation of ATR. In this study, we show evidence that the Ajuba-RPA interaction occurs through direct protein contact with RPA70, and that their association is cell cycle-regulated and is reduced upon DNA replication stress. We propose a model in which Ajuba negatively regulates the ATR pathway by directly interacting with RPA70, thereby preventing an inappropriate ATR activation. Our results provide a framework to understand the mechanism of regulation of ATR in human cells, which is important to prevent cellular transformation and tumorigenesis.

scholarly journals Dihydropyrimidinase protects from DNA replication stress caused by cytotoxic metabolites

2019 ◽  
Vol 48 (4) ◽  
pp. 1886-1904 ◽  
Author(s):  
Jihane Basbous ◽  
Antoine Aze ◽  
Laurent Chaloin ◽  
Rana Lebdy ◽  
Dana Hodroj ◽  
...  
Keyword(s):  
Dna Replication ◽  
Solid Tumors ◽  
Cancer Progression ◽  
Degradation Products ◽  
Replication Stress ◽  
Cellular Transformation ◽  
Chromosomal Dna ◽  
Normal Tissues ◽  
Dna Replication Stress ◽  
Pyrimidine Degradation

Abstract Imbalance in the level of the pyrimidine degradation products dihydrouracil and dihydrothymine is associated with cellular transformation and cancer progression. Dihydropyrimidines are degraded by dihydropyrimidinase (DHP), a zinc metalloenzyme that is upregulated in solid tumors but not in the corresponding normal tissues. How dihydropyrimidine metabolites affect cellular phenotypes remains elusive. Here we show that the accumulation of dihydropyrimidines induces the formation of DNA–protein crosslinks (DPCs) and causes DNA replication and transcriptional stress. We used Xenopus egg extracts to recapitulate DNA replication invitro. We found that dihydropyrimidines interfere directly with the replication of both plasmid and chromosomal DNA. Furthermore, we show that the plant flavonoid dihydromyricetin inhibits human DHP activity. Cellular exposure to dihydromyricetin triggered DPCs-dependent DNA replication stress in cancer cells. This study defines dihydropyrimidines as potentially cytotoxic metabolites that may offer an opportunity for therapeutic-targeting of DHP activity in solid tumors.

scholarly journals Budding yeast Dma1 and Dma2 participate in regulation of Swe1 levels and localization

2011 ◽  
Vol 22 (13) ◽  
pp. 2185-2197 ◽  
Author(s):  
Erica Raspelli ◽  
Corinne Cassani ◽  
Giovanna Lucchini ◽  
Roberta Fraschini
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Budding Yeast ◽  
Cell Cycle Control ◽  
Replication Stress ◽  
Down Regulation ◽  
Evolutionarily Conserved ◽  
Dna Replication Stress ◽  
Morphogenesis Checkpoint ◽  
The One

Timely down-regulation of the evolutionarily conserved protein kinase Swe1 plays an important role in cell cycle control, as Swe1 can block nuclear division through inhibitory phosphorylation of the catalytic subunit of cyclin-dependent kinase. In particular, Swe1 degradation is important for budding yeast cell survival in case of DNA replication stress, whereas it is inhibited by the morphogenesis checkpoint in response to alterations in actin cytoskeleton or septin structure. We show that the lack of the Dma1 and Dma2 ubiquitin ligases, which moderately affects Swe1 localization and degradation during an unperturbed cell cycle with no apparent phenotypic effects, is toxic for cells that are partially defective in Swe1 down-regulation. Moreover, Swe1 is stabilized, restrained at the bud neck, and hyperphosphorylated in dma1Δ dma2Δ cells subjected to DNA replication stress, indicating that the mechanism stabilizing Swe1 under these conditions is different from the one triggered by the morphogenesis checkpoint. Finally, the Dma proteins are required for proper Swe1 ubiquitylation. Taken together, the data highlight a previously unknown role of these proteins in the complex regulation of Swe1 and suggest that they might contribute to control, directly or indirectly, Swe1 ubiquitylation.

scholarly journals E2F1: Cause and Consequence of DNA Replication Stress

2021 ◽  
Vol 7 ◽  
Author(s):  
Shahd Fouad ◽  
David Hauton ◽  
Vincenzo D'Angiolella
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Tumor Suppressor ◽  
Signaling Pathways ◽  
Mammalian Cells ◽  
Replication Stress ◽  
Checkpoint Control ◽  
Oncogenic Signaling ◽  
Dna Replication Stress ◽  
Oncogenic Signaling Pathways

In mammalian cells, cell cycle entry occurs in response to the correct stimuli and is promoted by the transcriptional activity of E2F family members. E2F proteins regulate the transcription of S phase cyclins and genes required for DNA replication, DNA repair, and apoptosis. The activity of E2F1, the archetypal and most heavily studied E2F family member, is tightly controlled by the DNA damage checkpoints to modulate cell cycle progression and initiate programmed cell death, when required. Altered tumor suppressor and oncogenic signaling pathways often result in direct or indirect interference with E2F1 regulation to ensure higher rates of cell proliferation independently of external cues. Despite a clear link between dysregulated E2F1 activity and cancer progression, literature on the contribution of E2F1 to DNA replication stress phenotypes is somewhat scarce. This review discusses how dysfunctional tumor suppressor and oncogenic signaling pathways promote the disruption of E2F1 transcription and hence of its transcriptional targets, and how such events have the potential to drive DNA replication stress. In addition to the involvement of E2F1 upstream of DNA replication stress, this manuscript also considers the role of E2F1 as a downstream effector of the response to this type of cellular stress. Lastly, the review introduces some reflections on how E2F1 activity is integrated with checkpoint control through post-translational regulation, and proposes an exploitable tumor weakness based on this axis.

scholarly journals Protein phosphatase 2A (PP2A) promotes anaphase entry after DNA replication stress in budding yeast

2021 ◽  
Author(s):  
Cory Haluska ◽  
Fengzhi Jin ◽  
Yanchang Wang
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Dna Synthesis ◽  
Protein Phosphatase ◽  
Budding Yeast ◽  
Replication Stress ◽  
S Phase ◽  
Viability Loss ◽  
Phosphatase 2A ◽  
Dna Replication Stress

DNA replication stress activates the S-phase checkpoint that arrests the cell cycle, but it is poorly understood how cells recover from this arrest. Cyclin-dependent kinase (CDK) and Protein Phosphatase 2A (PP2A) are key cell cycle regulators, and Cdc55 is a regulatory subunit of PP2A in budding yeast. We found that yeast cells lacking functional PP2ACdc55 showed slow growth in the presence of hydroxyurea (HU), a DNA synthesis inhibitor, without obvious viability loss. Moreover, PP2A mutants exhibited delayed anaphase entry and sustained levels of anaphase inhibitor Pds1 after HU treatment. A DNA damage checkpoint Chk1 phosphorylates and stabilizes Pds1. We showed that chk1Δ and mutation of the Chk1 phosphorylation sites in Pds1 largely restored efficient anaphase entry in PP2A mutants after HU treatment. In addition, deletion of SWE1 that encodes the inhibitory kinase for CDK or mutation of the Swe1 phosphorylation site in CDK ( cdc28F19) also suppressed the anaphase entry delay in PP2A mutants after HU treatment. Our genetic data suggest that Swe1/CDK acts upstream of Pds1. Surprisingly, cdc55Δ showed significant suppression to the viability loss of S-phase checkpoint mutants during DNA synthesis block. Together, our results uncover a PP2A-Swe1-CDK-Chk1-Pds1 axis that promotes recovery from DNA replication stress.

scholarly journals ARID1A regulates R-loop associated DNA replication stress

2020 ◽  
Author(s):  
Shuhe Tsai ◽  
Emily Yun-chia Chang ◽  
Louis-Alexandre Fournier ◽  
James P. Wells ◽  
Sean W. Minaker ◽  
...  
Keyword(s):  
Dna Replication ◽  
Genome Stability ◽  
Clear Cell ◽  
Replication Stress ◽  
Clear Cell Ovarian Carcinoma ◽  
Dna Replication Stress ◽  
Cancer Types ◽  
The Impact ◽  
Oncogenic Gene ◽  
Transcription And Replication

ABSTRACTARID1A is lost in up to 7% of all cancers, and this frequency increases in certain cancer types, such as clear cell ovarian carcinoma where ARID1A protein is lost in about 50% of cases. While the impact of ARID1A loss on the function of the BAF chromatin remodeller complexes is likely to drive oncogenic gene expression programs in specific contexts, ARID1A also binds genome stability regulators such as ATR and TOP2. Here we show that ARID1A loss leads to DNA replication stress associated with R-loops and transcription-replication conflicts in human cells. These effects correlate with altered transcription and replication dynamics in ARID1A knockout cells and to reduced TOP2A binding at R-loop sites. Together this work extends mechanisms of replication stress in ARID1A deficient cells with implications for targeting ARID1A deficient cancers.

scholarly journals Interferon alpha-inducible protein 6 regulates NRASQ61K-induced melanomagenesis and growth

eLife ◽  
2016 ◽  
Vol 5 ◽  
Author(s):  
Romi Gupta ◽  
Matteo Forloni ◽  
Malik Bisserier ◽  
Shaillay Kumar Dogra ◽  
Qiaohong Yang ◽  
...  
Keyword(s):  
Dna Replication ◽  
Interferon Alpha ◽  
Replication Stress ◽  
Cellular Transformation ◽  
Mek Inhibitor ◽  
Cytotoxic Effects ◽  
Dna Replication Stress ◽  
Increased Sensitivity ◽  
Inducible Protein ◽  
Melanoma Growth

Mutations in the NRAS oncogene are present in up to 20% of melanoma. Here, we show that interferon alpha-inducible protein 6 (IFI6) is necessary for NRASQ61K-induced transformation and melanoma growth. IFI6 was transcriptionally upregulated by NRASQ61K, and knockdown of IFI6 resulted in DNA replication stress due to dysregulated DNA replication via E2F2. This stress consequentially inhibited cellular transformation and melanoma growth via senescence or apoptosis induction depending on the RB and p53 pathway status of the cells. NRAS-mutant melanoma were significantly more resistant to the cytotoxic effects of DNA replication stress-inducing drugs, and knockdown of IFI6 increased sensitivity to these drugs. Pharmacological inhibition of IFI6 expression by the MEK inhibitor trametinib, when combined with DNA replication stress-inducing drugs, blocked NRAS-mutant melanoma growth. Collectively, we demonstrate that IFI6, via E2F2 regulates DNA replication and melanoma development and growth, and this pathway can be pharmacologically targeted to inhibit NRAS-mutant melanoma.

scholarly journals Centrosome impairment causes DNA replication stress through MLK3/MK2 signaling and R-loop formation

2020 ◽  
Author(s):  
Zainab Tayeh ◽  
Kim Stegmann ◽  
Antonia Kleeberg ◽  
Mascha Friedrich ◽  
Josephine Ann Mun Yee Choo ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Replication Stress ◽  
List Type ◽  
Replication Forks ◽  
Loop Formation ◽  
Animal Cells ◽  
Replication Fork Progression ◽  
Primordial Dwarfism ◽  
Dna Replication Stress

AbstractCentrosomes function as organizing centers of microtubules and support accurate mitosis in many animal cells. However, it remains to be explored whether and how centrosomes also facilitate the progression through different phases of the cell cycle. Here we show that impairing the composition of centrosomes, by depletion of centrosomal components or by inhibition of polo-like kinase 4 (PLK4), reduces the progression of DNA replication forks. This occurs even when the cell cycle is arrested before damaging the centrosomes, thus excluding mitotic failure as the source of replication stress. Mechanistically, the kinase MLK3 associates with centrosomes. When centrosomes are disintegrated, MLK3 activates the kinases p38 and MK2/MAPKAPK2. Transcription-dependent RNA:DNA hybrids (R-loops) are then causing DNA replication stress. Fibroblasts from patients with microcephalic primordial dwarfism (Seckel syndrome) harbouring defective centrosomes showed replication stress and diminished proliferation, which were each alleviated by inhibition of MK2. Thus, centrosomes not only facilitate mitosis, but their integrity is also supportive in DNA replication.HighlightsCentrosome defects cause replication stress independent of mitosis.MLK3, p38 and MK2 (alias MAPKAPK2) are signalling between centrosome defects and DNA replication stress through R-loop formation.Patient-derived cells with defective centrosomes display replication stress, whereas inhibition of MK2 restores their DNA replication fork progression and proliferation.Graphical abstract

scholarly journals DNA replication stress induces deregulation of the cell cycle events in root meristems of Allium cepa

2012 ◽  
Vol 110 (8) ◽  
pp. 1581-1591 ◽  
Author(s):  
Aneta Żabka ◽  
Justyna Teresa Polit ◽  
Janusz Maszewski
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Allium Cepa ◽  
Replication Stress ◽  
Root Meristems ◽  
Dna Replication Stress
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