A Dbf4 Mutant Contributes to Bypassing the Rad53-mediated Block of Origins of Replication in Response to Genotoxic Stress

Duch, Alba; Palou, Gloria; Jónsson, Zophonı́as O.; Palou, Roger; Wohlschlegel, James A.; Quintana, David G.

doi:10.1074/jbc.m110.190843

Cited by 27 publications

(25 citation statements)

References 30 publications

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“…The BRCT domain of Dbf4 is immediately preceded by an ␣-helix that stabilizes the domain, and thus, it has been previously referred to as HBRCT (12,18). The HBRCT domain of Dbf4 is necessary and sufficient for the interaction with Rad53, yet it does not contain any threonine residue that could serve as the phosphorylation site recognized by Rad53 (7,9,12). A phosphorylated peptide derived from the N-terminal sequence of the HBRCT domain of Dbf4 including the canonical pTXXE FHA-binding motif can interact weakly with the FHA1 domain of Rad53 (19); however, mutation of this threonine in full-length Dbf4 does not disrupt the interaction with the FHA1 domain of Rad53 (12,19).…”

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confidence: 99%

“…One of the binding partners of FHA1 is Dbf4 (4), the regulatory subunit of the initiator kinase Cdc7 (5,6). The association of Rad53 and Dbf4 mediates the Rad53-dependent phosphorylation of Dbf4 and the consequent inhibition of Cdc7, thus preventing late origin firing (7)(8)(9)(10). This interaction involves the N-terminal region of Dbf4 that folds as a modified BRCA1 C-terminal (BRCT) domain (4,(11)(12)(13).…”

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confidence: 99%

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A Novel Non-canonical Forkhead-associated (FHA) Domain-binding Interface Mediates the Interaction between Rad53 and Dbf4 Proteins

Matthews

Selvaratnam

Jones

et al. 2014

Journal of Biological Chemistry

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Background:The interaction between the checkpoint kinase Rad53 and Dbf4 is critical to suppress late origin firing and to stabilize stalled forks during replication stress. Results: The FHA1 domain of Rad53 interacts with the BRCT domain of Dbf4 through a novel non-canonical interface. Conclusion: FHA domains gain specificity by engaging multiple binding interfaces. Significance: Understanding how FHA domains interact with their binding partners is key to elucidate how they relay information along signaling pathways.

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confidence: 99%

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confidence: 99%

A Novel Non-canonical Forkhead-associated (FHA) Domain-binding Interface Mediates the Interaction between Rad53 and Dbf4 Proteins

Matthews

Selvaratnam

Jones

et al. 2014

Journal of Biological Chemistry

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“…In parallel, Rad53 down-regulates the RNR inhibitor Sml1 to increase deoxyribonucleotide levels and facilitate DNA synthesis (Zhao et al 2001). In response to replication fork stalling, Rad53 prevents the activation of late replication origins by phosphorylating two proteins required for the initiation of DNA replication: Dbf4 and Sld3 (Lopez-Mosqueda et al 2010;Duch et al 2011). Dbf4 is the regulatory subunit of Cdc7 kinase, which is required to initiate DNA replication at individual origins by phosphorylating the replicative MCM helicase (Tsuji et al 2006;Francis et al 2009;Randell et al 2010;Sheu and Stillman 2010).…”

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confidence: 99%

DNA Replication Checkpoint Signaling Depends on a Rad53–Dbf4 N-Terminal Interaction in Saccharomyces cerevisiae

et al. 2013

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Dbf4-dependent kinase (DDK) and cyclin-dependent kinase (CDK) are essential to initiate DNA replication at individual origins. During replication stress, the S-phase checkpoint inhibits the DDK-and CDK-dependent activation of late replication origins. Rad53 kinase is a central effector of the replication checkpoint and both binds to and phosphorylates Dbf4 to prevent late-origin firing. The molecular basis for the Rad53-Dbf4 physical interaction is not clear but occurs through the Dbf4 N terminus. Here we found that both Rad53 FHA1 and FHA2 domains, which specifically recognize phospho-threonine (pT), interacted with Dbf4 through an N-terminal sequence and an adjacent BRCT domain. Purified Rad53 FHA1 domain (but not FHA2) bound to a pT Dbf4 peptide in vitro, suggesting a possible phospho-threonine-dependent interaction between FHA1 and Dbf4. The Dbf4-Rad53 interaction is governed by multiple contacts that are separable from the Cdc5-and Msa1-binding sites in the Dbf4 N terminus. Importantly, abrogation of the Rad53-Dbf4 physical interaction blocked Dbf4 phosphorylation and allowed late-origin firing during replication checkpoint activation. This indicated that Rad53 must stably bind to Dbf4 to regulate its activity.T HE fidelity of chromosome replication depends on checkpoint mechanisms to stabilize stalled forks, regulate origin activation, and repair DNA damage (Hartwell and Weinert 1989;Bartek et al. 2004;Segurado and Tercero 2009). In response to replication stress, the replication checkpoint maintains replisome stability and prevents late origins from firing, which allows time for DNA repair and the completion of DNA replication prior to chromosome segregation. Incomplete DNA replication or uncoordinated origin firing following DNA damage can result in genomic instability, cancer predisposition, and premature aging (Branzei and Foiani 2010).In the budding yeast Saccharomyces cerevisiae, activation of the checkpoint sensor kinase Mec1 (vertebrate ATR, Ataxia Telangiectasia and Rad3-related) is triggered at stalled forks or sites of DNA damage (Majka et al. 2006;Labib and De Piccoli 2011). Subsequent signal amplification through the Mrc1 or Rad9 adaptors leads to activation of the checkpoint kinase Rad53 (the ortholog of the human tumor suppressor Chk2) (Branzei and Foiani 2009). Rad53 is an integral transducer of various cellular responses to replication stress or DNA damage. Rad53 induces a series of transcriptional responses through MBF-regulated genes (Bastos de Oliveira et al. 2012;Travesa et al. 2012) and also activates the Dun1 kinase, which promotes the expression of ribonucleotide reductase (RNR) subunits and additional DNA repair genes (Huang et al. 1998). In parallel, Rad53 down-regulates the RNR inhibitor Sml1 to increase deoxyribonucleotide levels and facilitate DNA synthesis (Zhao et al. 2001). In response to replication fork stalling, Rad53 prevents the activation of late replication origins by phosphorylating two proteins required for the initiation of DNA replication: Dbf4 and Sld3 (...

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“…Very recent work with budding yeast has identified the main features of the mechanism by which the Rad53 kinase blocks the initiation step of replication [39][40][41]. Rad53 phosphorylates two factors that play a key role during the initiation of chromosome replication at each origin: the Dbf4 subunit of the Cdc7 kinase, and the Sld3 protein.…”

Section: Checkpoint Activation At Defective Replication Forks Inhibitmentioning

confidence: 99%

Surviving chromosome replication: the many roles of the S-phase checkpoint pathway

Labib

Piccoli

2011

Phil. Trans. R. Soc. B

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Checkpoints were originally identified as signalling pathways that delay mitosis in response to DNA damage or defects in chromosome replication, allowing time for DNA repair to occur. The ATR (ataxia-and rad-related) and ATM (ataxia-mutated) protein kinases are recruited to defective replication forks or to sites of DNA damage, and are thought to initiate the DNA damage response in all eukaryotes. In addition to delaying cell cycle progression, however, the S-phase checkpoint pathway also controls chromosome replication and DNA repair pathways in a highly complex fashion, in order to preserve genome integrity. Much of our understanding of this regulation has come from studies of yeasts, in which the best-characterized targets are the stimulation of ribonucleotide reductase activity by multiple mechanisms, and the inhibition of new initiation events at later origins of DNA replication. In addition, however, the S-phase checkpoint also plays a more enigmatic and apparently critical role in preserving the functional integrity of defective replication forks, by mechanisms that are still understood poorly. This review considers some of the key experiments that have led to our current understanding of this highly complex pathway.

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A Dbf4 Mutant Contributes to Bypassing the Rad53-mediated Block of Origins of Replication in Response to Genotoxic Stress

Cited by 27 publications

References 30 publications

A Novel Non-canonical Forkhead-associated (FHA) Domain-binding Interface Mediates the Interaction between Rad53 and Dbf4 Proteins

A Novel Non-canonical Forkhead-associated (FHA) Domain-binding Interface Mediates the Interaction between Rad53 and Dbf4 Proteins

DNA Replication Checkpoint Signaling Depends on a Rad53–Dbf4 N-Terminal Interaction in Saccharomyces cerevisiae

Surviving chromosome replication: the many roles of the S-phase checkpoint pathway

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