Checkpoint Regulation of Replication Dynamics in UV-Irradiated Human Cells

Chastain, Paul D.; Heffernan, Timothy P.; Nevis, Kathleen R.; Li, Lin; Kaufmann, William K.; Kaufman, David G.; Cordeiro‐Stone, Marila

doi:10.4161/cc.5.18.3236

Cited by 51 publications

(66 citation statements)

References 46 publications

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“…Direct assessment of replication fork progression using combed DNA and dual-pulse labeling with halogenated nucleotides has confirmed that the progression of replication forks has slowed down, that is, rates of chain elongation are reduced [Unsal-Kacmaz et al, 2007]. Similar effects have been observed when camptothecin is used to activate the intra-S-phase checkpoint [Chastain et al, 2006;Conti et al, 2007].…”

Section: Degradation Of P12 and The Conversion Of Pol D4 To Pol D3 Inmentioning

confidence: 71%

Regulation of human DNA polymerase delta in the cellular responses to DNA damage

Lee

Zhang

Lin

et al. 2012

Environ and Mol Mutagen

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The p12 subunit of polymerase delta (Pol d) is degraded in response to DNA damage induced by UV, alkylating agents, oxidative, and replication stresses. This leads to the conversion of the Pol d4 holoenzyme to the heterotrimer, Pol d3. We review studies that establish that Pol d3 formation is an event that could have a major impact on cellular processes in genomic surveillance, DNA replication, and DNA repair. p12 degradation is dependent on the apical ataxia telangiectasia and Rad3 related (ATR) kinase and is mediated by the ubiquitin-proteasome system. Pol d3 exhibits properties of an ''antimutator'' polymerase, suggesting that it could contribute to an increased surveillance against mutagenesis, for example, when Pol d carries out bypass synthesis past small base lesions that engage in spurious base pairing. Chromatin immunoprecipitation analysis and examination of the spatiotemporal recruitment of Pol d to sites of DNA damage show that Pol d3 is the primary form of Pol d associated with cyclobutane pyrimidine dimer lesions and therefore should be considered as the operative form of Pol d engaged in DNA repair. We propose a model for the switching of Pol d with translesion polymerases, incorporating the salient features of the recently determined structure of monoubiquitinated proliferating cell nuclear antigen and emphasizing the role of Pol d3. Because of the critical role of Pol d activity in DNA replication and repair, the formation of Pol d3 in response to DNA damage opens the prospect that pleiotropic effects may ensue. This opens the horizons for future exploration of how this novel response to DNA damage contributes to genomic stability. Environ. Mol. Mutagen. 53:683-698, 2012. V V C 2012 Wiley Periodicals, Inc.

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Section: Degradation Of P12 and The Conversion Of Pol D4 To Pol D3 Inmentioning

confidence: 71%

Regulation of human DNA polymerase delta in the cellular responses to DNA damage

Lee

Zhang

Lin

et al. 2012

Environ and Mol Mutagen

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“…A prior study showed that ionizing radiation reduced the fraction of green-only tracks in HeLa cells (44), consistent with a reduction in the rate of replicon initiation as a consequence of ATM-dependent intra-S checkpoint signaling (14,50). We recently reported that a low fluence of UVC produced an ATR-and Chk1-dependent reduction in the frequency of green-only tracks (6), demonstrating that the method is capable of detecting the intra-S checkpoint response to UV. When HeLa cells were transfected with a nontargeting control siRNA, UV treatment caused a 64% reduction in the fraction of green-only tracks, indicative of an effective intra-S checkpoint response that inhibits replicon initiation ( Fig.…”

Section: Resultsmentioning

confidence: 88%

The Human Tim/Tipin Complex Coordinates an Intra-S Checkpoint Response to UV That Slows Replication Fork Displacement

Ünsal-Kaçmaz

Chastain

Qü

et al. 2007

Molecular and Cellular Biology

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222

248

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UV-induced DNA damage stalls DNA replication forks and activates the intra-S checkpoint to inhibit replicon initiation. In response to stalled replication forks, ATR phosphorylates and activates the transducer kinase Chk1 through interactions with the mediator proteins TopBP1, Claspin, and Timeless (Tim). Murine Tim recently was shown to form a complex with Tim-interacting protein (Tipin), and a similar complex was shown to exist in human cells. Knockdown of Tipin using small interfering RNA reduced the expression of Tim and reversed the intra-S checkpoint response to UVC. Tipin interacted with replication protein A (RPA) and RPA-coated DNA, and RPA promoted the loading of Tipin onto RPA-free DNA. Immunofluorescence analysis of spread DNA fibers showed that treating HeLa cells with 2.5 J/m2 UVC not only inhibited the initiation of new replicons but also reduced the rate of chain elongation at active replication forks. The depletion of Tim and Tipin reversed the UV-induced inhibition of replicon initiation but affected the rate of DNA synthesis at replication forks in different ways. In undamaged cells depleted of Tim, the apparent rate of replication fork progression was 52% of the control. In contrast, Tipin depletion had little or no effect on fork progression in unirradiated cells but significantly attenuated the UV-induced inhibition of DNA chain elongation. Together, these findings indicate that the Tim-Tipin complex mediates the UV-induced intra-S checkpoint, Tim is needed to maintain DNA replication fork movement in the absence of damage, Tipin interacts with RPA on DNA and, in UV-damaged cells, Tipin slows DNA chain elongation in active replicons.

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“…Figure 1A illustrates the siRNA-mediated depletion of ATR, which in the irradiated cells was accompanied by a severe reduction in phosphorylation of its substrate CHK1, a known mediator of UV-induced S-phase checkpoint activation. [6][7][8] Velocity sedimentation analyses showed that fibroblasts pre-treated with the NTC-siRNA (Fig. 1B) displayed a stereotypical reduction in the abundance of small molecular weight (MW) nascent DNA when exposed to a low fluence of UVC.…”

Section: Depletion Of Atr or Chk1 Abrogated The Intra-s Phase Checkpomentioning

confidence: 99%

“…4,5 The second main signaling cascade recognizes bulky DNA adducts, such as those induced by UV radiation or benzo [a]pyrene diolepoxide. This checkpoint response to UV was found to require the activity of checkpoint proteins ataxia telangiectasia and Rad3-related (ATR) kinase, and its target CHK1, to act on downstream substrate(s) to inhibit the firing of new origins of replication [6][7][8] and to reduce the rate of DNA chain elongation. [9][10][11] At sub-lethal fluences of UV, the ATR/CHK1 pathway has been shown not to depend on degradation of CDC25A, but rather it has been proposed to act on the DBF4-dependent kinase to prevent CDC45-dependent activation of the MCM helicase, thus inhibiting new replicon initiation.…”

Section: Introductionmentioning

confidence: 99%

Is activation of the intra-S checkpoint in human fibroblasts an important factor in protection against UV-induced mutagenesis?

et al. 2013

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Checkpoint Regulation of Replication Dynamics in UV-Irradiated Human Cells

Cited by 51 publications

References 46 publications

Regulation of human DNA polymerase delta in the cellular responses to DNA damage

Regulation of human DNA polymerase delta in the cellular responses to DNA damage

The Human Tim/Tipin Complex Coordinates an Intra-S Checkpoint Response to UV That Slows Replication Fork Displacement

Is activation of the intra-S checkpoint in human fibroblasts an important factor in protection against UV-induced mutagenesis?

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