Reconstitution of an ATM-Dependent Checkpoint that Inhibits Chromosomal DNA Replication following DNA Damage

Costanzo, Vincenzo; Robertson, Kirsten; Ying, Carol Y.; Kim, Edward; Avvedimento, Enrico V.; Gottesman, Max E.; Grieco, Domenico; Gautier, Jean

doi:10.1016/s1097-2765(00)00063-0

Cited by 163 publications

(144 citation statements)

References 78 publications

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“…Xenopus cell-free extracts have been shown to recapitulate signalling pathways induced by DNA damage and have also contributed to delineation of the roles of Mre11 and ATM in this process (Costanzo et al, 2000(Costanzo et al, , 2001. Depletion of Mre11 from these extracts abrogated the response to DNA DSB.…”

Section: Viral Infection and In Vitro Models To Investigate The Role mentioning

confidence: 99%

ATM and the Mre11 complex combine to recognize and signal DNA double-strand breaks

2007

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The recognition and repair of DNA double-strand breaks (DSBs) is a complex process that draws upon a multitude of proteins. This is not surprising since this is a lethal lesion if left unrepaired and also contributes to genome instability and the consequential risk of cancer and other pathologies. Some of the key proteins that recognize these breaks in DNA are mutated in distinct genetic disorders that predispose to agent sensitivity, genome instability, cancer predisposition and/or neurodegeneration. These include members of the Mre11 complex (Mre11/Rad50/ Nbs1) and ataxia-telangiectasia (A-T) mutated (ATM), mutated in the human genetic disorder A-T. The mre11 (MRN) complex appears to be the major sensor of the breaks and subsequently recruits ATM where it is activated to phosphorylate in turn members of that complex and a variety of other proteins involved in cellcycle control and DNA repair. The MRN complex is also upstream of ATM and ATR (A-T-mutated and rad3-related) protein in responding to agents that block DNA replication. To date, more than 30 ATM-dependent substrates have been identified in multiple pathways that maintain genome stability and reduce the risk of disease. We focus here on the relationship between ATM and the MRN complex in recognizing and responding to DNA DSBs.

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Section: Viral Infection and In Vitro Models To Investigate The Role mentioning

confidence: 99%

ATM and the Mre11 complex combine to recognize and signal DNA double-strand breaks

2007

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“…Furthermore, active ATM-Chk2/ATR-Chk1 targeting of Cdc25A leads to the rapid destruction of phosphatase activity and consequently the levels of Cdk2 inhibitory phosphorylation increase (Molinari et al, 2000;Falck et al, 2001;Mailand et al, 2000). In combination, the negative regulation of Cdk2 contributes to G1 cell cycle arrest by preventing Cdk2 phosphorylation of Cdc45 and therefore, its loading onto orgins, which is required in the initiation of DNA replication (Costanzo et al, 2000;Broderick and Nasheuer, 2009). …”

Section: Dna Damage Checkpoint Responses Control Cell Cycle Progressimentioning

confidence: 99%

“…Typically, Xenopus egg extract has been applied in developmental and cell cycle control studies, in recent years it is being used increasingly to investigate DNA damage responses Gautier et al, 1990;Costanzo et al, 2004b;Garner and Costanzo, 2009). It has been shown that Xenopus egg extract can reproduce several aspects of DNA damage response and has provided significant insights into checkpoint mechanisms (Costanzo et al, 2000;Costanzo et al, 2003). Importantly, ATM/ATR have been isolated and found to function in regulating DNA damage checkpoint pathways within Xenopus egg extract (Robertson et al, 1999;Costanzo et al, 2000;Hekmat-Nejad et al, 2000;.…”

Section: Xenopus Laevis As a Model Systemmentioning

confidence: 99%

“…It has been shown that Xenopus egg extract can reproduce several aspects of DNA damage response and has provided significant insights into checkpoint mechanisms (Costanzo et al, 2000;Costanzo et al, 2003). Importantly, ATM/ATR have been isolated and found to function in regulating DNA damage checkpoint pathways within Xenopus egg extract (Robertson et al, 1999;Costanzo et al, 2000;Hekmat-Nejad et al, 2000;. In using this system, chemicals can be added to interfere with biochemical processes.…”

Section: Xenopus Laevis As a Model Systemmentioning

confidence: 99%

“…However, these works have mainly furthered the understanding of ATM and ATR checkpoint signalling in the setting of DNA replication. Typically, ATM and ATR activation has been prompted in egg extracts by the addition of DNA linear molecules mimicking DSBs (Costanzo et al, 2000;Guo and Murphy, 2000;Kumagai and Durphy, 2000;Costanzo et al, 2004a). ATM and ATR induction (as well as DNA-PK induction) in egg extract are sensitive to the presence of caffeine Blasina et al, 1999;.…”

Section: Atm and Atr Activation In Mitotic Xenopus Egg Extractsmentioning

confidence: 99%

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An ATM- and ATR-dependent checkpoint inactivates spindle assembly by targeting CEP63

Smith¹,

Dejsuphong

Balestrini³

et al. 2009

Nat Cell Biol

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The effects of ATM and ATR signalling induced by chromosomal breakage have been described extensively in modulating cell cycle progression up to the onset of mitosis.However, DNA damage checkpoint responses in mitotic cells are not well understood.This thesis reports on the effects of double strand breaks on the progression of mitosis.We found ATM and ATR activation can occur in mitotic Xenopus laevis egg extract and the induction of ATM and ATR by chromosomal breakages inhibits spindle assembly in both Xenopus egg extract and somatic cells. The delay in mitotic progression induced by ATM and ATR was found not to involve major spindle assembly factors activities such as, Cdk1, Plx1 and RCC1/Ran-GTP. However, normal anastral spindles formation around linear DNA coated beads, which can activate ATM and ATR, linked centrosome-driven spindle assembly to ATM and ATR dependent spindle defects. cDNA expression library screening was undertaken to identify novel ATM and ATR targets in this mitotic checkpoint pathway, through which the novel centrosomal protein XCEP63 was identified as a likely candidate. Data obtained from depletion and reconstitution of XCEP63 in Xenopus egg extract established that normal centrosome-driven spindle assembly requires XCEP63. Moreover, ATM and ATR phosphorylates XCEP63 on serine 560 and promotes delocalisation from the centrosome. ATM and ATR inhibition or addition of non-phosphorylable XCEP63 recombinant protein mutated at serine 560 prevents spindle assembly abnormalities.These findings suggest that ATM and ATR regulate mitotic events by targeting XCEP63 and centrosome-dependent spindle assembly. This pathway may provide support for DNA repair processes or regulate cell survival in the presence of mitotic DNA damage. 3

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Eukaryotic Replication Origins and Initiation of DNA Replication

DePamphilis¹,

Aladjem²

2010

Encyclopedia of Life Sciences

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Eukaryotic deoxyribonucleic acid (DNA) replication begins at specific genomic sites called replication origins that serve as assembly sites for prereplication complexes (preRCs). PreRCs include proteins that recognise origin sequences, helicases that separate the two strands of DNA and accessory proteins that facilitate helicase binding and interaction with cell cycle regulatory pathways. Assembly of preRCs is required for initiation of DNA replication which occurs after those complexes recruit additional proteins including DNA polymerases . The sequence requirements for replication origins vary but they all include several distinct DNA elements that act synergistically to facilitate preRC assembly. The number of potential replication origins is higher than the number of actual replication initiation sites. Epigenetic processes and metabolic conditions dynamically select the location of replication initiation events of each cell cycle to insure complete and accurate replication of the entire genome in coordination with gene expression and chromatin condensation. Key Concepts: Replication initiates at distinct chromosomal locations that recruit prereplication (preRC) complexes. PreRCs are recruited sequentially, each step subject to strict regulation to prevent rereplication. Cyclin‐dependent kinases (CDK) levels change during the cell cycle. Low CDK activity is required for preRC assembly as cells exit mitosis. High CDK activity is required for activation of existing preRCs and initiation of DNA synthesis, while simultaneously suppressing assembly of new preRCs. Replication origin sequences vary among metazoans, but the functions of proteins that bind replication origins are conserved. Eukaryotic origins exhibit a modular structure. Modularity is detectable in single‐cell eukaryotes such as yeast and is more pronounced in metazoans, in which replication origins often cluster. Not all potential replication origins are activated in each cell cycle. Utilisation of replication origins is regulated dynamically to facilitate coordination with other metabolic processes occurring on chromatin. Either hyperactivation or suppression of replication origins can damage DNA and cause chromosomal rearrangements, suggesting that spacing replication initiation events at defined intervals facilitates genomic stability.

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Reconstitution of an ATM-Dependent Checkpoint that Inhibits Chromosomal DNA Replication following DNA Damage

Cited by 163 publications

References 78 publications

ATM and the Mre11 complex combine to recognize and signal DNA double-strand breaks

ATM and the Mre11 complex combine to recognize and signal DNA double-strand breaks

An ATM- and ATR-dependent checkpoint inactivates spindle assembly by targeting CEP63

Eukaryotic Replication Origins and Initiation of DNA Replication

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