Mec1 Is Activated at the Onset of Normal S Phase by Low-dNTP Pools Impeding DNA Replication

Forey, Romain; Poveda, Ana; Sharma, Sushma; Barthe, Antoine; Padioleau, Ismaël; Renard, Claire-Angélique; Lambert, Robin; Skrzypczak, Magdalena; Ginalski, Krzysztof; Lengronne, Armelle; Chabes, Andrei; Pardo, Benjamín; Pasero, Philippe

doi:10.1016/j.molcel.2020.02.021

Cited by 51 publications

(54 citation statements)

References 80 publications

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“…Similarly, a recent study by the Lindqvist laboratory has shown that active DNA replication restricts the activation of CDK1 and PLK1 mitotic kinases through CHK1 [58]. Another recent study in yeast has shown that the basal activation of the Mec1-Rad53 (the functional homologs of ATR-CHK1) pathway results from the spontaneous replication stress triggered by an insufficient level of dNTPs in the early S phase that contributes to the coordination of the origin activity with dNTP synthesis, a function conserved in mammals [47,59,60]. Interestingly, however, the essential function of Mec1-Rad53 is linked to the maintenance of replication fork integrity, which prevents cells bearing under-replicated and damaged chromosomes from progressing to mitosis [59].…”

Section: Mechanisms Protecting Against Replication Stress and Under-rmentioning

confidence: 99%

“…Another recent study in yeast has shown that the basal activation of the Mec1-Rad53 (the functional homologs of ATR-CHK1) pathway results from the spontaneous replication stress triggered by an insufficient level of dNTPs in the early S phase that contributes to the coordination of the origin activity with dNTP synthesis, a function conserved in mammals [47,59,60]. Interestingly, however, the essential function of Mec1-Rad53 is linked to the maintenance of replication fork integrity, which prevents cells bearing under-replicated and damaged chromosomes from progressing to mitosis [59]. The stabilization of the replication fork protects the stalled fork from degradation and maintains its competence to resume DNA synthesis upon removal or bypass of the replication obstacle [45,61].…”

Section: Mechanisms Protecting Against Replication Stress and Under-rmentioning

confidence: 99%

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DNA Replication Stress and Chromosomal Instability: Dangerous Liaisons

Wilhelm

Said

Naim

2020

Genes

104

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Chromosomal instability (CIN) is associated with many human diseases, including neurodevelopmental or neurodegenerative conditions, age-related disorders and cancer, and is a key driver for disease initiation and progression. A major source of structural chromosome instability (s-CIN) leading to structural chromosome aberrations is “replication stress”, a condition in which stalled or slowly progressing replication forks interfere with timely and error-free completion of the S phase. On the other hand, mitotic errors that result in chromosome mis-segregation are the cause of numerical chromosome instability (n-CIN) and aneuploidy. In this review, we will discuss recent evidence showing that these two forms of chromosomal instability can be mechanistically interlinked. We first summarize how replication stress causes structural and numerical CIN, focusing on mechanisms such as mitotic rescue of replication stress (MRRS) and centriole disengagement, which prevent or contribute to specific types of structural chromosome aberrations and segregation errors. We describe the main outcomes of segregation errors and how micronucleation and aneuploidy can be the key stimuli promoting inflammation, senescence, or chromothripsis. At the end, we discuss how CIN can reduce cellular fitness and may behave as an anticancer barrier in noncancerous cells or precancerous lesions, whereas it fuels genomic instability in the context of cancer, and how our current knowledge may be exploited for developing cancer therapies.

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Section: Mechanisms Protecting Against Replication Stress and Under-rmentioning

confidence: 99%

Section: Mechanisms Protecting Against Replication Stress and Under-rmentioning

confidence: 99%

DNA Replication Stress and Chromosomal Instability: Dangerous Liaisons

Wilhelm

Said

Naim

2020

Genes

104

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“…Importantly, this model assumes a heterogeneous probability of origin firing and suggests that Chk1 exerts a global origin inhibitory action during unperturbed S phase ( Platel et al 2015 ). On the other hand, the constancy of the initial number of limiting factors N 0 in the presence or absence of UCN-01 suggests that Chk1 does not actively control origins before S phase actually starts ( Lupardus et al 2002 Stokes et al 2002 Forey et al 2020 ). These observations indicate that MM4 can deliver a reliable, minimally complex picture of origin firing regulation in Xenopus egg extracts.…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, a recent study in unperturbed yeast cells suggests that dNTPs are limiting at the entry into S phase, so that, similar to Xenopus ( Zou 2007 ), the firing of the earliest origins creates a replication stress that activates the Rad53 checkpoint which prevents further origin firing. Rad53 activation also stimulates dNTP synthesis, which in turn down regulates the checkpoint and allows later origin firing ( Forey et al 2020 ). However, it remains uncertain if this feed-back loop does also exist in Xenopus egg extracts which contain an abundant pool of dNTPs.…”

Section: Discussionmentioning

confidence: 99%

“…It is now well accepted that the intra-S phase checkpoint regulates origin firing during both unperturbed and artificially perturbed S phase ( Marheineke and Hyrien 2004 Ge and Blow, 2010 Guo et al, 2015 Platel et al, 2015 Forey et al, 2020 ). DNA replication stress, through the activation of the S-phase checkpoint kinase Rad53, can inhibit origin firing by phosphorylating and inhibiting Sld3 and Dbf4 ( Zegerman and Diley, 2010 ).…”

Section: Introductionmentioning

confidence: 99%

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Organization of DNA replication origin firing inXenopusegg extracts : the role of intra-S checkpoint

Ciardo

Haccard

Narassimprakash

et al. 2020

Preprint

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During cell division, the duplication of the genome starts at multiple positions called 14 replication origins. Origin firing requires the interaction of rate-limiting factors with potential 15 origins during the S(ynthesis)-phase of the cell cycle. Origins fire as synchronous clusters is 16 proposed to be regulated by the intra-S checkpoint. By modelling either the unchallenged or the 17 checkpoint-inhibited replication pattern of single DNA molecules from Xenopus sperm chromatin 18 replicated in egg extracts, we demonstrate that the quantitative modelling of data require: 1) a 19 segmentation of the genome into regions of low and high probability of origin firing; 2) that regions 20 with high probability of origin firing escape intra-S checkpoint regulation; 3) that the intra-S 21 checkpoint controls the firing of replication origins in regions with low probability of firing. This 22 model implies that the intra-S checkpoint is not the main regulator of origin clustering. The minimal 23 nature of the proposed model foresees its use to analyse data from other eukaryotic organisms. 24 25 2013; Rhind and Gilbert, 2013). The core motor component of the replicative helicase, the MCM2-33 7 complex, is loaded on chromatin from late mitosis until the end of G1 phase as an inactive 34 head-to-head double hexamer (DH) to form a large excess of potential origins (DePamphilis et al., 35 2006; Ticau et al., 2015). During S phase, only a fraction of the MCM2-7 DHs are activated to 36 form a pair of active Cdc45-MCM2-7-GINS (CMG) helicases and establish bidirectional replisomes 37 (DePamphilis and Bell, 2010). MCM2-7 DHs that fail to fire are inactivated by forks emanating from 38 neighboring fired origins (Blow et al., 2011). Origin firing requires S-phase cyclin-dependent kinase 39 (CDK) and Dbf4-dependent kinase (DDK) activities as well as the CDK targets Sld2 and Sld3 and the 40 1 of 29 Manuscript submitted to eLife replisome-maturation scaffolds Dpb11 and Sld7 in S. cerevisiae. The six initiation factors Sld2, Sld3, 41 Dpb11, Dbf4, Sld7 and Cdc45 are expressed at concentrations significantly lower than the MCM 42 complex and core replisome components, suggesting that they may be rate-limiting for origin firing 43 (Mantiero et al., 2011; Tanaka et al., 2011). Among these six factors, Cdc45 is the only one to travel 44 with the replication fork. 45 DNA replication initiates without sequence specificity in Xenopus eggs (Harland and Laskey, 1980; 46 Méchali and Kearsey, 1984), egg extracts (Mahbubani et al., 1992; Hyrien and Méchali, 1992; Carli 47 et al., 2016, 2018) and early embryos (Hyrien and Méchali, 1993; Hyrien et al., 1995) (for review see 48 Hyrien et al. (2003)). To understand how a lack of preferred sequences for replication initiation 49 is compatible with a precise S-phase completion time, investigators have studied replication at 50 the single DNA molecule level using the DNA combing technique (Lucas et al., 2000; Herrick et al., 51 2000; Blow et al., 2001; Marheineke and Hyrien, 2001, 2004). In contr...

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Rad53 arrests leading and lagging strand DNA synthesis via distinct mechanisms in response to DNA replication stress

Zhang

2022

BioEssays

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DNA replication stress threatens ordinary DNA synthesis. The evolutionarily conserved DNA replication stress response pathway involves sensor kinase Mec1/ATR, adaptor protein Mrc1/Claspin, and effector kinase Rad53/Chk1, which spurs a host of changes to stabilize replication forks and maintain genome integrity. DNA replication forks consist of largely distinct sets of proteins at leading and lagging strands that function autonomously in DNA synthesis in vitro. In this article, we discuss eSPAN and BrdU‐IP‐ssSeq, strand‐specific sequencing technologies that permit analysis of protein localization and DNA synthesis at individual strands in budding yeast. Using these approaches, we show that under replication stress Rad53 stalls DNA synthesis on both leading and lagging strands. On lagging strands, it stimulates PCNA unloading, and on leading strands, it attenuates the replication function of Mrc1‐Tof1. We propose that in doing so, Rad53 couples leading and lagging strand DNA synthesis during replication stress, thereby preventing the emergence of harmful ssDNA.

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Mec1 Is Activated at the Onset of Normal S Phase by Low-dNTP Pools Impeding DNA Replication

Cited by 51 publications

References 80 publications

DNA Replication Stress and Chromosomal Instability: Dangerous Liaisons

DNA Replication Stress and Chromosomal Instability: Dangerous Liaisons

Organization of DNA replication origin firing inXenopusegg extracts : the role of intra-S checkpoint

Rad53 arrests leading and lagging strand DNA synthesis via distinct mechanisms in response to DNA replication stress

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