scholarly journals MUS81 nuclease activity is essential for replication stress tolerance and chromosome segregation in BRCA2-deficient cells

2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Xianning Lai ◽  
Ronan Broderick ◽  
Valérie Bergoglio ◽  
Jutta Zimmer ◽  
Sophie Badie ◽  
...  
Keyword(s):  
Stress Tolerance ◽  
Chromosome Segregation ◽  
Replication Fork ◽  
Replication Stress ◽  
Nuclease Activity ◽  
Dna Lesions ◽  
Genome Replication ◽  
Replication Fork Progression ◽  
Nucleolytic Activity ◽  
Genomic Regions

AbstractFailure to restart replication forks stalled at genomic regions that are difficult to replicate or contain endogenous DNA lesions is a hallmark of BRCA2 deficiency. The nucleolytic activity of MUS81 endonuclease is required for replication fork restart under replication stress elicited by exogenous treatments. Here we investigate whether MUS81 could similarly facilitate DNA replication in the context of BRCA2 abrogation. Our results demonstrate that replication fork progression in BRCA2-deficient cells requires MUS81. Failure to complete genome replication and defective checkpoint surveillance enables BRCA2-deficient cells to progress through mitosis with under-replicated DNA, which elicits severe chromosome interlinking in anaphase. MUS81 nucleolytic activity is required to activate compensatory DNA synthesis during mitosis and to resolve mitotic interlinks, thus facilitating chromosome segregation. We propose that MUS81 provides a mechanism of replication stress tolerance, which sustains survival of BRCA2-deficient cells and can be exploited therapeutically through development of specific inhibitors of MUS81 nuclease activity.

scholarly journals Erratum: Corrigendum: MUS81 nuclease activity is essential for replication stress tolerance and chromosome segregation in BRCA2-deficient cells

2017 ◽  
Vol 8 (1) ◽  
Author(s):  
Xianning Lai ◽  
Ronan Broderick ◽  
Valérie Bergoglio ◽  
Jutta Zimmer ◽  
Sophie Badie ◽  
...  
Keyword(s):  
Stress Tolerance ◽  
Chromosome Segregation ◽  
Replication Stress ◽  
Nuclease Activity

scholarly journals Structural basis for the multi-activity factor Rad5 in replication stress tolerance

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Miaomiao Shen ◽  
Nalini Dhingra ◽  
Quan Wang ◽  
Chen Cheng ◽  
Songbiao Zhu ◽  
...  
Keyword(s):  
Stress Tolerance ◽  
Ubiquitin Ligase ◽  
Replication Fork ◽  
Replication Stress ◽  
Spatial Arrangement ◽  
Structural Basis ◽  
Dna Lesion ◽  
Activity Factor ◽  
Fork Regression ◽  
New Type

AbstractThe yeast protein Rad5 and its orthologs in other eukaryotes promote replication stress tolerance and cell survival using their multiple activities, including ubiquitin ligase, replication fork remodeling and DNA lesion targeting activities. Here, we present the crystal structure of a nearly full-length Rad5 protein. The structure shows three distinct, but well-connected, domains required for Rad5’s activities. The spatial arrangement of these domains suggest that different domains can have autonomous activities but also undergo intrinsic coordination. Moreover, our structural, biochemical and cellular studies demonstrate that Rad5’s HIRAN domain mediates interactions with the DNA metabolism maestro factor PCNA and contributes to its poly-ubiquitination, binds to DNA and contributes to the Rad5-catalyzed replication fork regression, defining a new type of HIRAN domains with multiple activities. Our work provides a framework to understand how Rad5 integrates its various activities in replication stress tolerance.

scholarly journals Mms1 and Mms22 stabilize the replisome during replication stress

2011 ◽  
Vol 22 (13) ◽  
pp. 2396-2408 ◽  
Author(s):  
Jessica A. Vaisica ◽  
Anastasija Baryshnikova ◽  
Michael Costanzo ◽  
Charles Boone ◽  
Grant W. Brown
Keyword(s):  
Dna Replication ◽  
Ubiquitin Ligase ◽  
Replication Fork ◽  
E3 Ubiquitin Ligase ◽  
Replication Stress ◽  
Replication Forks ◽  
Replication Fork Progression ◽  
Binding Partners ◽  
Efficient Recovery ◽  
Stalled Replication Forks

Mms1 and Mms22 form a Cul4Ddb1-like E3 ubiquitin ligase with the cullin Rtt101. In this complex, Rtt101 is bound to the substrate-specific adaptor Mms22 through a linker protein, Mms1. Although the Rtt101Mms1/Mms22ubiquitin ligase is important in promoting replication through damaged templates, how it does so has yet to be determined. Here we show that mms1Δ and mms22Δ cells fail to properly regulate DNA replication fork progression when replication stress is present and are defective in recovery from replication fork stress. Consistent with a role in promoting DNA replication, we find that Mms1 is enriched at sites where replication forks have stalled and that this localization requires the known binding partners of Mms1—Rtt101 and Mms22. Mms1 and Mms22 stabilize the replisome during replication stress, as binding of the fork-pausing complex components Mrc1 and Csm3, and DNA polymerase ε, at stalled replication forks is decreased in mms1Δ and mms22Δ. Taken together, these data indicate that Mms1 and Mms22 are important for maintaining the integrity of the replisome when DNA replication forks are slowed by hydroxyurea and thereby promote efficient recovery from replication stress.

scholarly journals Topologically associating domain boundaries are enriched in early firing origins and restrict replication fork progression

2020 ◽  
Author(s):  
Emilia Puig Lombardi ◽  
Madalena Tarsounas
Keyword(s):  
Binding Sites ◽  
Replication Fork ◽  
Replication Timing ◽  
Replication Initiation ◽  
Ctcf Binding ◽  
Origin Firing ◽  
Replication Fork Progression ◽  
Fork Stalling ◽  
Genomic Regions ◽  
The Impact

ABSTRACTTopologically associating domains (TADs) are units of the genome architecture defined by binding sites for the CTCF transcription factor and cohesin-mediated loop extrusion. Genomic regions containing DNA replication initiation sites have been mapped in the proximity of TAD boundaries. However, the factors that determine this positioning have not been identified. Moreover, the impact of TADs on the directionality of replication fork progression remains unknown. Here we use EdU-seq technology to map origin firing sites at 10 kb resolution and to monitor replication fork progression after restart from hydroxyurea arrest. We show that origins firing in early/mid S-phase within TAD boundaries map to two distinct peaks flanking the centre of the boundary, which is occupied by CTCF and cohesin. When transcription is inhibited chemically or deregulated by oncogene overexpression, replication origins become repositioned to the centre of the TAD. Furthermore, we demonstrate the strikingly asymmetric fork progression initiating from origins located within TAD boundaries. Divergent CTCF binding sites and neighbouring TADs with different replication timing (RT) cause fork stalling in regions external to the TAD. Thus, our work assigns for the first time a role to transcription within TAD boundaries in promoting replication origin firing and demonstrates how genomic regions adjacent to the TAD boundaries could restrict replication progression.

scholarly journals CHK1 inhibition exacerbates replication stress induced by IGF blockade

Oncogene ◽  
2021 ◽  
Author(s):  
Xiaoning Wu ◽  
Elena Seraia ◽  
Stephanie B. Hatch ◽  
Xiao Wan ◽  
Daniel V. Ebner ◽  
...  
Keyword(s):  
Ribonucleotide Reductase ◽  
Replication Fork ◽  
Synthetic Lethality ◽  
Growth Factor Receptor ◽  
Replication Stress ◽  
Regulatory Subunit ◽  
Breast Cancer Cell Lines ◽  
Replication Fork Progression ◽  
Deoxynucleotide Triphosphate

AbstractWe recently reported that genetic or pharmacological inhibition of insulin-like growth factor receptor (IGF-1R) slows DNA replication and induces replication stress by downregulating the regulatory subunit RRM2 of ribonucleotide reductase, perturbing deoxynucleotide triphosphate (dNTP) supply. Aiming to exploit this effect in therapy we performed a compound screen in five breast cancer cell lines with IGF neutralising antibody xentuzumab. Inhibitor of checkpoint kinase CHK1 was identified as a top screen hit. Co-inhibition of IGF and CHK1 caused synergistic suppression of cell viability, cell survival and tumour growth in 2D cell culture, 3D spheroid cultures and in vivo. Investigating the mechanism of synthetic lethality, we reveal that CHK1 inhibition in IGF-1R depleted or inhibited cells further downregulated RRM2, reduced dNTP supply and profoundly delayed replication fork progression. These effects resulted in significant accumulation of unreplicated single-stranded DNA and increased cell death, indicative of replication catastrophe. Similar phenotypes were induced by IGF:WEE1 co-inhibition, also via exacerbation of RRM2 downregulation. Exogenous RRM2 expression rescued hallmarks of replication stress induced by co-inhibiting IGF with CHK1 or WEE1, identifying RRM2 as a critical target of the functional IGF:CHK1 and IGF:WEE1 interactions. These data identify novel therapeutic vulnerabilities and may inform future trials of IGF inhibitory drugs.

scholarly journals SUMO-Based Regulation of Nuclear Positioning to Spatially Regulate Homologous Recombination Activities at Replication Stress Sites

Genes ◽  
2021 ◽  
Vol 12 (12) ◽  
pp. 2010
Author(s):  
Kamila Schirmeisen ◽  
Sarah A. E. Lambert ◽  
Karol Kramarz
Keyword(s):  
Homologous Recombination ◽  
Replication Fork ◽  
Replication Stress ◽  
Dna Lesions ◽  
Nuclear Pore ◽  
Nuclear Pore Complexes ◽  
Replication Forks ◽  
Nuclear Positioning ◽  
Nuclear Compartment ◽  
Pore Complexes

DNA lesions have properties that allow them to escape their nuclear compartment to achieve DNA repair in another one. Recent studies uncovered that the replication fork, when its progression is impaired, exhibits increased mobility when changing nuclear positioning and anchors to nuclear pore complexes, where specific types of homologous recombination pathways take place. In yeast models, increasing evidence points out that nuclear positioning is regulated by small ubiquitin-like modifier (SUMO) metabolism, which is pivotal to maintaining genome integrity at sites of replication stress. Here, we review how SUMO-based pathways are instrumental to spatially segregate the subsequent steps of homologous recombination during replication fork restart. In particular, we discussed how routing towards nuclear pore complex anchorage allows distinct homologous recombination pathways to take place at halted replication forks.

scholarly journals MIF is a 3’ flap nuclease that facilitates DNA replication and promotes tumor growth

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yijie Wang ◽  
Yan Chen ◽  
Chenliang Wang ◽  
Mingming Yang ◽  
Yanan Wang ◽  
...  
Keyword(s):  
Dna Replication ◽  
Tumor Growth ◽  
Dna Synthesis ◽  
Cancer Cells ◽  
Macrophage Migration Inhibitory Factor ◽  
Replication Fork ◽  
Replication Stress ◽  
Nuclease Activity ◽  
Cell Growth Inhibition ◽  
Poor Overall Survival

AbstractHow cancer cells cope with high levels of replication stress during rapid proliferation is currently unclear. Here, we show that macrophage migration inhibitory factor (MIF) is a 3’ flap nuclease that translocates to the nucleus in S phase. Poly(ADP-ribose) polymerase 1 co-localizes with MIF to the DNA replication fork, where MIF nuclease activity is required to resolve replication stress and facilitates tumor growth. MIF loss in cancer cells leads to mutation frequency increases, cell cycle delays and DNA synthesis and cell growth inhibition, which can be rescued by restoring MIF, but not nuclease-deficient MIF mutant. MIF is significantly upregulated in breast tumors and correlates with poor overall survival in patients. We propose that MIF is a unique 3’ nuclease, excises flaps at the immediate 3’ end during DNA synthesis and favors cancer cells evading replication stress-induced threat for their growth.

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