Postreplication Repair and PCNA Modification in Schizosaccharomyces pombe

Ubiquitination of proliferating cell nuclear antigen (PCNA) plays a crucial role in regulating replication past DNA damage in eukaryotes, but the detailed mechanisms appear to vary in different organisms. We have examined the modification of PCNA in Schizosaccharomyces pombe. We find that, in response to UV irradiation, PCNA is mono- and poly-ubiquitinated in a manner similar to that in Saccharomyces cerevisiae. However in undamaged Schizosaccharomyces pombe cells, PCNA is ubiquitinated in S phase, whereas in S. cerevisiae it is sumoylated. Furthermore we find that, unlike in S. cerevisiae, mutants defective in ubiquitination of PCNA are also sensitive to ionizing radiation, and PCNA is ubiquitinated after exposure of cells to ionizing radiation, in a manner similar to the response to UV-irradiation. We show that PCNA modification and cell cycle checkpoints represent two independent signals in response to DNA damage. Finally, we unexpectedly find that PCNA is ubiquitinated in response to DNA damage when cells are arrested in G2.

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p53-Mediated Regulation of Proliferating Cell Nuclear Antigen Expression in Cells Exposed to Ionizing Radiation

Molecular and Cellular Biology ◽

10.1128/mcb.19.1.12 ◽

1999 ◽

Vol 19 (1) ◽

pp. 12-20 ◽

Cited By ~ 63

Author(s):

Jin Xu ◽

Gilbert F. Morris

Keyword(s):

Dna Damage ◽

Dna Repair ◽

Ionizing Radiation ◽

Proliferating Cell Nuclear Antigen ◽

Cellular Response ◽

Nuclear Antigen ◽

Dominant Negative Mutant ◽

Mrna Levels ◽

Mobility Shift ◽

Proliferating Cell

ABSTRACT The proliferating cell nuclear antigen (PCNA) is a highly conserved cellular protein that functions both in DNA replication and in DNA repair. Exposure of a rat embryo fibroblast cell line (CREF cells) to γ radiation induced simultaneous expression of PCNA with the p53 tumor suppressor protein and the cyclin-dependent kinase inhibitor p21WAF1/Cip1. PCNA mRNA levels transiently increased in serum-starved cells exposed to ionizing radiation, an observation suggesting that the radiation-associated increase in PCNA expression could be dissociated from cell cycle progression. Irradiation of CREF cells activated a transiently expressed PCNA promoter chloramphenicol acetyltransferase construct through p53 binding sequences via a mechanism blocked by a dominant negative mutant p53. Electrophoretic mobility shift assays with nuclear extracts prepared from irradiated CREF cells produced four p53-specific DNA-protein complexes with the PCNA p53 binding site. Addition of monoclonal antibody PAb421 (p53-specific) or AC238 (specific to the transcriptional coactivator p300/CREB binding protein) to the mobility shift assay distinguished different forms of p53 that changed in relative abundance with time after irradiation. These findings suggest a complex cellular response to DNA damage in which p53 transiently activates expression of PCNA for the purpose of limited DNA repair. In a population of nongrowing cells with diminished PCNA levels, this pathway may be crucial to survival following DNA damage.

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CDK Inhibitor p21 Is Degraded by a Proliferating Cell Nuclear Antigen-coupled Cul4-DDB1Cdt2Pathway during S Phase and after UV Irradiation

Journal of Biological Chemistry ◽

10.1074/jbc.m806045200 ◽

2008 ◽

Vol 283 (43) ◽

pp. 29045-29052 ◽

Cited By ~ 156

Author(s):

Hideo Nishitani ◽

Yasushi Shiomi ◽

Hiroka Iida ◽

Masato Michishita ◽

Toshihiro Takami ◽

...

Keyword(s):

Uv Irradiation ◽

Proliferating Cell Nuclear Antigen ◽

S Phase ◽

Nuclear Antigen ◽

Cdk Inhibitor ◽

Proliferating Cell

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ATM Protein Physically and Functionally Interacts with Proliferating Cell Nuclear Antigen to Regulate DNA Synthesis

Journal of Biological Chemistry ◽

10.1074/jbc.m112.352310 ◽

2012 ◽

Vol 287 (15) ◽

pp. 12445-12454 ◽

Cited By ~ 10

Author(s):

Armin M. Gamper ◽

Serah Choi ◽

Yoshihiro Matsumoto ◽

Dibyendu Banerjee ◽

Alan E. Tomkinson ◽

...

Keyword(s):

Dna Damage ◽

Dna Replication ◽

Ionizing Radiation ◽

Dna Synthesis ◽

Dna Polymerase ◽

Proliferating Cell Nuclear Antigen ◽

Nuclear Antigen ◽

Atm Kinase ◽

Atm Protein ◽

Proliferating Cell

Ataxia telangiectasia (A-T) is a pleiotropic disease, with a characteristic hypersensitivity to ionizing radiation that is caused by biallelic mutations in A-T mutated (ATM), a gene encoding a protein kinase critical for the induction of cellular responses to DNA damage, particularly to DNA double strand breaks. A long known characteristic of A-T cells is their ability to synthesize DNA even in the presence of ionizing radiation-induced DNA damage, a phenomenon termed radioresistant DNA synthesis. We previously reported that ATM kinase inhibition, but not ATM protein disruption, blocks sister chromatid exchange following DNA damage. We now show that ATM kinase inhibition, but not ATM protein disruption, also inhibits DNA synthesis. Investigating a potential physical interaction of ATM with the DNA replication machinery, we found that ATM co-precipitates with proliferating cell nuclear antigen (PCNA) from cellular extracts. Using bacterially purified ATM truncation mutants and in vitro translated PCNA, we showed that the interaction is direct and mediated by the C terminus of ATM. Indeed, a 20-amino acid region close to the kinase domain is sufficient for strong binding to PCNA. This binding is specific to ATM, because the homologous regions of other PIKK members, including the closely related kinase A-T and Rad3-related (ATR), did not bind PCNA. ATM was found to bind two regions in PCNA. To examine the functional significance of the interaction between ATM and PCNA, we tested the ability of ATM to stimulate DNA synthesis by DNA polymerase δ, which is implicated in both DNA replication and DNA repair processes. ATM was observed to stimulate DNA polymerase activity in a PCNA-dependent manner.

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Saccharomyces cerevisiae replication factor C. II. Formation and activity of complexes with the proliferating cell nuclear antigen and with DNA polymerases delta and epsilon.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)54625-1 ◽

1991 ◽

Vol 266 (33) ◽

pp. 22698-22706 ◽

Cited By ~ 11

Author(s):

P.M. Burgers

Keyword(s):

Saccharomyces Cerevisiae ◽

Proliferating Cell Nuclear Antigen ◽

Dna Polymerases ◽

Nuclear Antigen ◽

Replication Factor C ◽

Replication Factor ◽

Factor C ◽

Proliferating Cell

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Oxidative DNA Damage Bypass in Arabidopsis thaliana Requires DNA Polymerase λ and Proliferating Cell Nuclear Antigen 2

The Plant Cell ◽

10.1105/tpc.110.081455 ◽

2011 ◽

Vol 23 (2) ◽

pp. 806-822 ◽

Cited By ~ 36

Author(s):

Alessandra Amoroso ◽

Lorenzo Concia ◽

Caterina Maggio ◽

Cécile Raynaud ◽

Catherine Bergounioux ◽

...

Keyword(s):

Dna Damage ◽

Arabidopsis Thaliana ◽

Dna Polymerase ◽

Proliferating Cell Nuclear Antigen ◽

Oxidative Dna Damage ◽

Nuclear Antigen ◽

Dna Polymerase Λ ◽

Proliferating Cell

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Proliferating Cell Nuclear Antigen-dependent Rapid Recruitment of Cdt1 and CRL4Cdt2at DNA-damaged Sites after UV Irradiation in HeLa Cells

Journal of Biological Chemistry ◽

10.1074/jbc.m110.161661 ◽

2010 ◽

Vol 285 (53) ◽

pp. 41993-42000 ◽

Cited By ~ 21

Author(s):

Takashi Ishii ◽

Yasushi Shiomi ◽

Toshihiro Takami ◽

Yusuke Murakami ◽

Naho Ohnishi ◽

...

Keyword(s):

Uv Irradiation ◽

Hela Cells ◽

Proliferating Cell Nuclear Antigen ◽

Nuclear Antigen ◽

Proliferating Cell

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Proliferating cell nuclear antigen destabilizes c-Abl tyrosine kinase and regulates cell apoptosis in response to DNA damage

APOPTOSIS ◽

10.1007/s10495-009-0313-2 ◽

2009 ◽

Vol 14 (3) ◽

pp. 268-275 ◽

Cited By ~ 9

Author(s):

Xiang He ◽

Congwen Wei ◽

Ting Song ◽

Jing Yuan ◽

Yanhong Zhang ◽

...

Keyword(s):

Dna Damage ◽

Tyrosine Kinase ◽

Cell Apoptosis ◽

Proliferating Cell Nuclear Antigen ◽

Nuclear Antigen ◽

Abl Tyrosine Kinase ◽

Proliferating Cell

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Effect of Proliferating Cell Nuclear Antigen Ubiquitination and Chromatin Structure on the Dynamic Properties of the Y-family DNA Polymerases

Molecular Biology of the Cell ◽

10.1091/mbc.e08-07-0724 ◽

2008 ◽

Vol 19 (12) ◽

pp. 5193-5202 ◽

Cited By ~ 64

Author(s):

Simone Sabbioneda ◽

Audrey M. Gourdin ◽

Catherine M. Green ◽

Angelika Zotter ◽

Giuseppina Giglia-Mari ◽

...

Keyword(s):

Chromatin Structure ◽

Proliferating Cell Nuclear Antigen ◽

Dynamic Properties ◽

Human Fibroblasts ◽

Dna Polymerases ◽

S Phase ◽

Nuclear Antigen ◽

Replication Foci ◽

Y Family ◽

Proliferating Cell

Y-family DNA polymerases carry out translesion synthesis past damaged DNA. DNA polymerases (pol) η and ι are usually uniformly distributed through the nucleus but accumulate in replication foci during S phase. DNA-damaging treatments result in an increase in S phase cells containing polymerase foci. Using photobleaching techniques, we show that polη is highly mobile in human fibroblasts. Even when localized in replication foci, it is only transiently immobilized. Although ubiquitination of proliferating cell nuclear antigen (PCNA) is not required for the localization of polη in foci, it results in an increased residence time in foci. polι is even more mobile than polη, both when uniformly distributed and when localized in foci. Kinetic modeling suggests that both polη and polι diffuse through the cell but that they are transiently immobilized for ∼150 ms, with a larger proportion of polη than polι immobilized at any time. Treatment of cells with DRAQ5, which results in temporary opening of the chromatin structure, causes a dramatic immobilization of polη but not polι. Our data are consistent with a model in which the polymerases are transiently probing the DNA/chromatin. When DNA is exposed at replication forks, the polymerase residence times increase, and this is further facilitated by the ubiquitination of PCNA.

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