An in vivo analysis of the localisation and interactions of human p66 DNA polymerase δ subunit

Pohler, J Richard G; Otterlei, Marit; Warbrick, Emma

doi:10.1186/1471-2199-6-17

Cited by 26 publications

(13 citation statements)

References 49 publications

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“…5f). ATR is known to be activated by a wide variety of genotoxic challenges as well as uncoupling of DNA replication proteins from the replication fork (28).…”

Section: Discussionmentioning

confidence: 99%

DNA Methyltransferase 1 Knockdown Activates a Replication Stress Checkpoint

Unterberger¹,

Andrews²,

Weaver

et al. 2006

Molecular and Cellular Biology

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DNA methyltransferase 1 (DNMT1) is an important component of the epigenetic machinery and is responsible for copying DNA methylation patterns during cell division. Coordination of DNA methylation and DNA replication is critical for maintaining epigenetic programming. Knockdown of DNMT1 leads to inhibition of DNA replication, but the mechanism has been unclear. Here we show that depletion of DNMT1 with either antisense or small interfering RNA (siRNA) specific to DNMT1 activates a cascade of genotoxic stress checkpoint proteins, resulting in phosphorylation of checkpoint kinases 1 and 2 (Chk1 and -2), ␥H2AX focus formation, and cell division control protein 25a (CDC25a) degradation, in an ataxia telangiectasia mutatedRad3-related (ATR)-dependent manner. siRNA knockdown of ATR blocks the response to DNMT1 depletion; DNA synthesis continues in the absence of DNMT1, resulting in global hypomethylation. Similarly, the response to DNMT1 knockdown is significantly attenuated in human mutant ATR fibroblast cells from a Seckel syndrome patient. This response is sensitive to DNMT1 depletion, independent of the catalytic domain of DNMT1, as indicated by abolition of the response with ectopic expression of either DNMT1 or DNMT1 with the catalytic domain deleted. There is no response to short-term treatment with 5-aza-deoxycytidine (5-azaCdR), which causes demethylation by trapping DNMT1 in 5-aza-CdR-containing DNA but does not cause disappearance of DNMT1 from the nucleus. Our data are consistent with the hypothesis that removal of DNMT1 from replication forks is the trigger for this response.Maintenance of the epigenome is crucial for normal gene expression and preservation of cell identity. The "epigenome" is the set of heritable properties encoded by elements other than DNA base sequence. The epigenome includes chromatin, which is fashioned by remodeling complexes and modification enzymes as well as a pattern of covalent modification of DNA by methylation. During cell division replication of both the genetic and epigenetic, information is faithfully conserved. Due to a preference for hemimethylated DNA, DNA methyltransferase 1 (DNMT1) is believed to be responsible for copying the DNA methylation pattern within the mother cell and maintaining the pattern of genomic methylation within the daughter cell (38). Several lines of evidence suggest a role for DNMT1 in cellular transformation (2,22,23,37,44): indeed, DNMT1 is upregulated in multiple human cancers (17, 33), and some colorectal cancers have been observed to bear mutations in DNMT1 (19). DNMT1 has therefore been proposed as a target for anticancer therapy (39). Indeed, preclinical studies using antisense to DNMT1 have already shown reversion of tumor growth both in vitro (9) and in vivo (29). Taken together, these data suggest an important role for DNMT1 in both maintaining the epigenome and controlling cell cycle.A loss of DNMT1 during replication would result in a loss of epigenetic information. It has therefore been proposed that cells have developed multiple ways...

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“…5f). ATR is known to be activated by a wide variety of genotoxic challenges as well as uncoupling of DNA replication proteins from the replication fork (28).…”

Section: Discussionmentioning

confidence: 99%

DNA Methyltransferase 1 Knockdown Activates a Replication Stress Checkpoint

Unterberger¹,

Andrews²,

Weaver

et al. 2006

Molecular and Cellular Biology

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“…Although its main application was by far the textile industry, tremendous quantities of this material are consumed in the manufacture of video and audio tapes, X-ray films, food packaging and especially of soft-drink bottles. The overall world consumption of PET amounts to about 26 million tons a year [1] compared to about 13 million tons, reported in 1997. [2] Since the duration of life of PET wastes (mainly from mineral-water and soft-drink bottles) is very small, there is a vast waste stream that reaches each year to the final recipients creating a serious environmental problem.…”

Section: Introductionmentioning

confidence: 99%

Hydrolytic Depolymerization of PET in a Microwave Reactor

Siddiqui

Achilias

Redhwi

et al. 2010

Macro Materials & Eng

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Recycling of PET was examined using hydrolytic depolymerization in an alkaline solution under microwave irradiation. The reaction was carried out in a sealed microwave reactor in which the pressure and temperature were controlled and recorded. The main products were the monomers TPA and EG. The effect of reaction temperature, time, amount of PET and alkaline concentration on the degree of PET depolymerization and TPA recovery was investigated. Microwave irradiation was found to reduce the time needed to achieve a specific degradation of PET significantly, with almost complete depolymerization occurring in 30 min at 180 °C and only 46 W of microwave power. Using a phase transfer catalyst (TOMAB) resulted in the same amount of unreacted PET but at significantly lower depolymerization temperatures.magnified image

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“…Human Pol ␦ consists of four subunits, p125, p50, p68, and p12. Subunit p125 harbors the polymerase and exonuclease active sites, p50 is tightly associated with the p125 subunit, p68 is associated with p50, and p12 binds to both p125 and p50 (23)(24)(25)(26).…”

mentioning

confidence: 99%

Coronavirus Infection Induces DNA Replication Stress Partly through Interaction of Its Nonstructural Protein 13 with the p125 Subunit of DNA Polymerase δ

Huang

Fang

et al. 2011

Journal of Biological Chemistry

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Perturbation of cell cycle regulation is a characteristic feature of infection by many DNA and RNA viruses, including Coronavirus infectious bronchitis virus (IBV). IBV infection was shown to induce cell cycle arrest at both S and G 2 /M phases for the enhancement of viral replication and progeny production. However, the underlying mechanisms are not well explored. In this study we show that activation of cellular DNA damage response is one of the mechanisms exploited by Coronavirus to induce cell cycle arrest. An ATR-dependent cellular DNA damage response was shown to be activated by IBV infection. Suppression of the ATR kinase activity by chemical inhibitors and siRNA-mediated knockdown of ATR reduced the IBV-induced ATR signaling and inhibited the replication of IBV. Furthermore, yeast two-hybrid screens and subsequent biochemical and functional studies demonstrated that interaction between Coronavirus nsp13 and DNA polymerase ␦ induced DNA replication stress in IBV-infected cells. These findings indicate that the ATR signaling activated by IBV replication contributes to the IBV-induced S-phase arrest and is required for efficient IBV replication and progeny production.DNA damage response is a signal transduction pathway that coordinates cell cycle transition, DNA replication, and repair in response to DNA damage or replication stress (1, 2). It is essential for maintenance of genome integrity and cell survival. DNA damage response is primarily mediated by two related protein kinases, the ataxia-telangiectasia mutated (ATM) 2 and ATM/ Rad3-related (ATR). ATM is activated as a result of doublestranded breaks (DSBs) and is recruited to DSBs by Mre11-Rad50-NBS1 complex (3, 4). ATR, on the other hand, is activated by a wide range of DNA damage, including stalled DNA replication forks and the subsequent single-stranded lesion (ssDNA), base adducts, ultraviolet (UV)-induced nucleotide damage, and double-stranded breaks during S phase (1, 5). ATR is recruited to replication factor A (RPA)-coated ssDNA by ATR-interacting protein (ATRIP) (6, 7). When ATM or ATR is recruited to sites of damage, they phosphorylate and activate different substrates, including checkpoint kinase-2 (CHK2) and CHK1, respectively (1,8,9). A large number of overlapping substrates of ATM and ATR that might be involved in DNA damage response has been presented (e.g. H2AX, RPA32, p53, BRCA1) (10, 11). Those signaling modules finally lead to cell cycle arrest to allow for DNA repair or apoptosis in cases of severe DNA damage. In contrast to ATM, ATR prevents replication fork collapse at stalled replication forks and is essential for cell viability (7,(12)(13)(14)(15)(16)(17)(18).Many DNA viruses and retroviruses, including Epstein-Barr virus, herpes simplex virus 1, human Papillomavirus 16, human immunodeficiency virus (HIV), adenovirus, simian virus 40 (SV40), and Polyomavirus, are known to eliminate, circumvent, or exploit various aspects of cellular DNA damage response machinery to maximize their own replication. In RNA virus families, however,...

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An in vivo analysis of the localisation and interactions of human p66 DNA polymerase δ subunit

Cited by 26 publications

References 49 publications

DNA Methyltransferase 1 Knockdown Activates a Replication Stress Checkpoint

DNA Methyltransferase 1 Knockdown Activates a Replication Stress Checkpoint

Hydrolytic Depolymerization of PET in a Microwave Reactor

Coronavirus Infection Induces DNA Replication Stress Partly through Interaction of Its Nonstructural Protein 13 with the p125 Subunit of DNA Polymerase δ

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