DNA damage checkpoint triggers autophagy to regulate the initiation of anaphase (original) (raw)
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Molecular Cell, 2001
In budding yeast, a single unrepaired DSB, for example, one created by expression of the site-specific HO endonuclease, is sufficient to cause G2/M arrest (Sandell and Zakian, 1993). However, these cells do not remain permanently arrested; rather, after 8-12 hr they resume progression through the cell cycle (Sandell and Waltham, Massachusetts 02454 Zakian, 1993; Toczyski et al., 1997; Lee et al., 1998). This † Istituto F.I.R.C. di Oncologia Molecolare and escape from G2/M arrest occurs despite the continued Dipartimento di Genetica e di Biologia dei presence of the broken chromosome and hence is Microrganismi termed adaptation (Sandell and Zakian, 1993; Toczyski Universita' degli Studi di Milano et al., 1997; Lee et al., 1998).
The yeast DNA damage checkpoint proteins control a cytoplasmic response to DNA damage
Proceedings of the National Academy of Sciences, 2007
A single HO endonuclease-induced double-strand break (DSB) is sufficient to activate the DNA damage checkpoint and cause Saccharomyces cells to arrest at G 2 /M for 12–14 h, after which cells adapt to the presence of the DSB and resume cell cycle progression. The checkpoint signal leading to G 2 /M arrest was previously shown to be nuclear-limited. Cells lacking ATR-like Mec1 exhibit no DSB-induced cell cycle delay; however, cells lacking Mec1's downstream protein kinase targets, Rad53 or Chk1, still have substantial G 2 /M delay, as do cells lacking securin, Pds1. This delay is eliminated only in the triple mutant chk1 Δ rad53 Δ pds1 Δ, suggesting that Rad53 and Chk1 control targets other than the stability of securin in enforcing checkpoint-mediated cell cycle arrest. The G 2 /M arrest in rad53 Δ and chk1 Δ revealed a unique cytoplasmic phenotype in which there are frequent dynein-dependent excursions of the nucleus through the bud neck, without entering anaphase. Such excursi...
The DNA Damage Checkpoint Signal in Budding Yeast Is Nuclear Limited
Molecular Cell, 2000
and of several key components is regulated; for example, in Department of Genetics mammalian cells Cdk1 is prevented from entering the Stanford University nucleus in response to DNA damage by a 14-3-3 protein Stanford, California 94305 (Jin et al., 1998; Chan et al., 1999). In fission yeast, † Rosenstiel Center and although Cdk1 seems to be predominantly nuclear at Department of Biology all times, the localization of its activating phosphatase, Brandeis University Cdc25, changes in response to DNA damage, becoming Waltham, Massachusetts 02454 cytoplasmic (Furnari et al., 1999). These observations suggest that control of the entry into mitosis after DNA damage might require transmission of the DNA damage Summary signal to the cytoplasmic compartment, although they could also be explained by a nuclear-limited active ex-The nature of the DNA damage-induced checkpoint port of a key mitotic component from the nucleus. signal that causes the arrest of cells prior to mitosis In the budding yeast Saccharomyces cerevisiae, inis unknown. To determine if this signal is transmitted hibitory phosphorylation of Cdc28p, the homolog of Cdk1, does not play a role in cell cycle arrest after DNA through the cytoplasm or is confined to the nucleus, damage (Amon et al., 1992; Sorger and Murray, 1992).
PLoS Genetics, 2011
Checkpoint pathways regulate genomic integrity in part by blocking anaphase until all chromosomes have been completely replicated, repaired, and correctly aligned on the spindle. In Saccharomyces cerevisiae, DNA damage and mono-oriented or unattached kinetochores trigger checkpoint pathways that bifurcate to regulate both the metaphase to anaphase transition and mitotic exit. The sensor-associated kinase, Mec1, phosphorylates two downstream kinases, Chk1 and Rad53. Activation of Chk1 and Rad53 prevents anaphase and causes inhibition of the mitotic exit network. We have previously shown that the PKA pathway plays a role in blocking securin and Clb2 destruction following DNA damage. Here we show that the Mec1 DNA damage checkpoint regulates phosphorylation of the regulatory (R) subunit of PKA following DNA damage and that the phosphorylated R subunit has a role in restraining mitosis following DNA damage. In addition we found that proteins known to regulate PKA in response to nutrients and stress either by phosphorylation of the R subunit or regulating levels of cAMP are required for the role of PKA in the DNA damage checkpoint. Our data indicate that there is cross-talk between the DNA damage checkpoint and the proteins that integrate nutrient and stress signals to regulate PKA.
Molecular and cellular biology, 2007
In Saccharomyces cerevisiae, double-strand breaks (DSBs) activate DNA checkpoint pathways that trigger several responses including a strong G(2)/M arrest. We have previously provided evidence that the phosphatases Ptc2 and Ptc3 of the protein phosphatase 2C type are required for DNA checkpoint inactivation after a DSB and probably dephosphorylate the checkpoint kinase Rad53. In this article we have investigated further the interactions between Ptc2 and Rad53. We showed that forkhead-associated domain 1 (FHA1) of Rad53 interacts with a specific threonine of Ptc2, T376, located outside its catalytic domain in a TXXD motif which constitutes an optimal FHA1 binding sequence in vitro. Mutating T376 abolishes Ptc2 interaction with the Rad53 FHA1 domain and results in adaptation and recovery defects following a DSB. We found that Ckb1 and Ckb2, the regulatory subunits of the protein kinase CK2, are necessary for the in vivo interaction between Ptc2 and the Rad53 FHA1 domain, that Ckb1 bind...
Molecular and Cellular Biology, 2004
DNA damage checkpoint pathways sense DNA lesions and transduce the signals into appropriate biological responses, including cell cycle arrest, induction of transcriptional programs, and modification or activation of repair factors. Here we show that the Saccharomyces cerevisiae Sae2 protein, known to be involved in processing meiotic and mitotic double-strand breaks, is required for proper recovery from checkpoint-mediated cell cycle arrest after DNA damage and is phosphorylated periodically during the unperturbed cell cycle and in response to DNA damage. Both cell cycle-and DNA damage-dependent Sae2 phosphorylation requires the main checkpoint kinase, Mec1, and the upstream components of its pathway, Ddc1, Rad17, Rad24, and Mec3. Another pathway, involving Tel1 and the MRX complex, is also required for full DNA damage-induced Sae2 phosphorylation, that is instead independent of the downstream checkpoint transducers Rad53 and Chk1, as well as of their mediators Rad9 and Mrc1. Mutations altering all the favored ATM/ATR phosphorylation sites of Sae2 not only abolish its in vivo phosphorylation after DNA damage but also cause hypersensitivity to methyl methanesulfonate treatment, synthetic lethality with RAD27 deletion, and decreased rates of mitotic recombination between inverted Alu repeats, suggesting that checkpoint-mediated phosphorylation of Sae2 is important to support its repair and recombination functions.