Publications by authors named "Atsuya Nishiyama"

19 Publications

  • Page 1 of 1

Recent evolution of a TET-controlled and DPPA3/STELLA-driven pathway of passive DNA demethylation in mammals.

Nat Commun 2020 11 24;11(1):5972. Epub 2020 Nov 24.

Department of Biology II and Center for Integrated Protein Science Munich (CIPSM), Human Biology and BioImaging, Ludwig-Maximilians-Universität München, Planegg-Martinsried, Germany.

Genome-wide DNA demethylation is a unique feature of mammalian development and naïve pluripotent stem cells. Here, we describe a recently evolved pathway in which global hypomethylation is achieved by the coupling of active and passive demethylation. TET activity is required, albeit indirectly, for global demethylation, which mostly occurs at sites devoid of TET binding. Instead, TET-mediated active demethylation is locus-specific and necessary for activating a subset of genes, including the naïve pluripotency and germline marker Dppa3 (Stella, Pgc7). DPPA3 in turn drives large-scale passive demethylation by directly binding and displacing UHRF1 from chromatin, thereby inhibiting maintenance DNA methylation. Although unique to mammals, we show that DPPA3 alone is capable of inducing global DNA demethylation in non-mammalian species (Xenopus and medaka) despite their evolutionary divergence from mammals more than 300 million years ago. Our findings suggest that the evolution of Dppa3 facilitated the emergence of global DNA demethylation in mammals.
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http://dx.doi.org/10.1038/s41467-020-19603-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7686362PMC
November 2020

Generation of a p16 Reporter Mouse and Its Use to Characterize and Target p16 Cells In Vivo.

Cell Metab 2020 Nov 18;32(5):814-828.e6. Epub 2020 Sep 18.

Division of Cancer Cell Biology, The Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan. Electronic address:

Cell senescence plays a key role in age-associated organ dysfunction, but the in vivo pathogenesis is largely unclear. Here, we generated a p16-Cre-tdTomato mouse model to analyze the in vivo characteristics of p16 cells at a single-cell level. We found tdTomato-positive p16 cells detectable in all organs, which were enriched with age. We also found that these cells failed to proliferate and had half-lives ranging from 2.6 to 4.2 months, depending on the tissue examined. Single-cell transcriptomics in the liver and kidneys revealed that p16 cells were present in various cell types, though most dominant in hepatic endothelium and in renal proximal and distal tubule epithelia, and that these cells exhibited heterogeneous senescence-associated phenotypes. Further, elimination of p16 cells ameliorated nonalcoholic steatohepatitis-related hepatic lipidosis and immune cell infiltration. Our new mouse model and single-cell analysis provide a powerful resource to enable the discovery of previously unidentified senescence functions in vivo.
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http://dx.doi.org/10.1016/j.cmet.2020.09.006DOI Listing
November 2020

Two distinct modes of DNMT1 recruitment ensure stable maintenance DNA methylation.

Nat Commun 2020 03 6;11(1):1222. Epub 2020 Mar 6.

Division of Cancer Cell Biology, The Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo, Japan.

Stable inheritance of DNA methylation is critical for maintaining differentiated phenotypes in multicellular organisms. We have recently identified dual mono-ubiquitylation of histone H3 (H3Ub2) by UHRF1 as an essential mechanism to recruit DNMT1 to chromatin. Here, we show that PCNA-associated factor 15 (PAF15) undergoes UHRF1-dependent dual mono-ubiquitylation (PAF15Ub2) on chromatin in a DNA replication-coupled manner. This event will, in turn, recruit DNMT1. During early S-phase, UHRF1 preferentially ubiquitylates PAF15, whereas H3Ub2 predominates during late S-phase. H3Ub2 is enhanced under PAF15 compromised conditions, suggesting that H3Ub2 serves as a backup for PAF15Ub2. In mouse ES cells, loss of PAF15Ub2 results in DNA hypomethylation at early replicating domains. Together, our results suggest that there are two distinct mechanisms underlying replication timing-dependent recruitment of DNMT1 through PAF15Ub2 and H3Ub2, both of which are prerequisite for high fidelity DNA methylation inheritance.
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http://dx.doi.org/10.1038/s41467-020-15006-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7060239PMC
March 2020

Enhanced processivity of Dnmt1 by monoubiquitinated histone H3.

Genes Cells 2020 Jan 3;25(1):22-32. Epub 2019 Dec 3.

Laboratory of Epigenetics, Institute for Protein Research, Osaka University, Suita, Japan.

DNA methylation controls gene expression, and once established, DNA methylation patterns are faithfully copied during DNA replication by the maintenance DNA methyltransferase Dnmt1. In vivo, Dnmt1 interacts with Uhrf1, which recognizes hemimethylated CpGs. Recently, we reported that Uhrf1-catalyzed K18- and K23-ubiquitinated histone H3 binds to the N-terminal region (the replication focus targeting sequence, RFTS) of Dnmt1 to stimulate its methyltransferase activity. However, it is not yet fully understood how ubiquitinated histone H3 stimulates Dnmt1 activity. Here, we show that monoubiquitinated histone H3 stimulates Dnmt1 activity toward DNA with multiple hemimethylated CpGs but not toward DNA with only a single hemimethylated CpG, suggesting an influence of ubiquitination on the processivity of Dnmt1. The Dnmt1 activity stimulated by monoubiquitinated histone H3 was additively enhanced by the Uhrf1 SRA domain, which also binds to RFTS. Thus, Dnmt1 activity is regulated by catalysis (ubiquitination)-dependent and -independent functions of Uhrf1.
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http://dx.doi.org/10.1111/gtc.12732DOI Listing
January 2020

Structure of the Dnmt1 Reader Module Complexed with a Unique Two-Mono-Ubiquitin Mark on Histone H3 Reveals the Basis for DNA Methylation Maintenance.

Mol Cell 2017 Oct;68(2):350-360.e7

Division of Cancer Cell Biology, The Institute of Medical Science, The University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan; Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601, Japan. Electronic address:

The proper location and timing of Dnmt1 activation are essential for DNA methylation maintenance. We demonstrate here that Dnmt1 utilizes two-mono-ubiquitylated histone H3 as a unique ubiquitin mark for its recruitment to and activation at DNA methylation sites. The crystal structure of the replication foci targeting sequence (RFTS) of Dnmt1 in complex with H3-K18Ub/23Ub reveals striking differences to the known ubiquitin-recognition structures. The two ubiquitins are simultaneously bound to the RFTS with a combination of canonical hydrophobic and atypical hydrophilic interactions. The C-lobe of RFTS, together with the K23Ub surface, also recognizes the N-terminal tail of H3. The binding of H3-K18Ub/23Ub results in spatial rearrangement of two lobes in the RFTS, suggesting the opening of its active site. Actually, incubation of Dnmt1 with H3-K18Ub/23Ub increases its catalytic activity in vitro. Our results therefore shed light on the essential role of a unique ubiquitin-binding module in DNA methylation maintenance.
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http://dx.doi.org/10.1016/j.molcel.2017.09.037DOI Listing
October 2017

Usp7-dependent histone H3 deubiquitylation regulates maintenance of DNA methylation.

Sci Rep 2017 03 3;7(1):55. Epub 2017 Mar 3.

Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1, Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya, 467-8601, Japan.

Uhrf1-dependent histone H3 ubiquitylation plays a crucial role in the maintenance of DNA methylation via the recruitment of the DNA methyltransferase Dnmt1 to DNA methylation sites. However, the involvement of deubiquitylating enzymes (DUBs) targeting ubiquitylated histone H3 in the maintenance of DNA methylation is largely unknown. With the use of Xenopus egg extracts, we demonstrate here that Usp7, a ubiquitin carboxyl-terminal hydrolase, forms a stable complex with Dnmt1 and is recruited to DNA methylation sites during DNA replication. Usp7 deubiquitylates ubiquitylated histone H3 in vitro. Inhibition of Usp7 activity or its depletion in egg extracts results in enhanced and extended binding of Dnmt1 to chromatin, suppressing DNA methylation. Depletion of Usp7 in HeLa cells causes enhanced histone H3 ubiquitylation and enlargement of Dnmt1 nuclear foci during DNA replication. Our results thus suggest that Usp7 is a key factor that regulates maintenance of DNA methylation.
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http://dx.doi.org/10.1038/s41598-017-00136-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5427934PMC
March 2017

The replication foci targeting sequence (RFTS) of DNMT1 functions as a potent histone H3 binding domain regulated by autoinhibition.

Biochem Biophys Res Commun 2016 Feb 14;470(3):741-747. Epub 2016 Jan 14.

Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1, Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya, Aichi, 467-8601, Japan. Electronic address:

DNA methyltransferase 1 (DNMT1) plays an essential role in propagation of the DNA methylation pattern to daughter cells. The replication foci targeting sequence (RFTS) of DNMT1 is required for the recruitment of DNMT1 to DNA methylation sites through direct binding to ubiquitylated histone H3 mediated by UHRF1 (Ubiquitin-like containing PHD and RING finger domains 1). Recently, it has been reported that the RFTS plugs the catalytic pocket of DNMT1 in an intermediated manner and inhibits its DNA methyltransferase activity. However, it is unclear whether this binding affects RFTS function in terms of recruitment to DNA methylation sites. Using Xenopus egg extracts, we demonstrate here that abrogation of the interaction between the RFTS and the catalytic center of DNMT1, by deletion of the C-terminal portion or disruption of the hydrogen bond, results in non-ubiquitylated histone H3 binding and abnormal accumulation of DNMT1 on the chromatin. Interestingly, DNMT1 mutants identified in patients with a neurodegenerative disease, ADCA-DN, bound to non-ubiquitylated histone H3 and accumulated on chromatin during S phase in Xenopus egg extracts. These results suggest that the interaction between the RFTS and the catalytic center of DNMT1 serves as an autoinhibitory mechanism for suppressing the histone H3 binding of DNMT1 and ensuring the accurate recruitment of DNMT1 to sites of DNA methylation. The autoinhibitory mechanism may play an important role in the regulation of gene expression in neurogenesis.
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http://dx.doi.org/10.1016/j.bbrc.2016.01.029DOI Listing
February 2016

Loss of maintenance DNA methylation results in abnormal DNA origin firing during DNA replication.

Biochem Biophys Res Commun 2016 Jan 22;469(4):960-6. Epub 2015 Dec 22.

Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601, Japan. Electronic address:

The mammalian maintenance methyltransferase DNMT1 [DNA (cytosine-5-)-methyltransferase 1] mediates the inheritance of the DNA methylation pattern during replication. Previous studies have shown that depletion of DNMT1 causes a severe growth defect and apoptosis in differentiated cells. However, the detailed mechanisms behind this phenomenon remain poorly understood. Here we show that conditional ablation of Dnmt1 in murine embryonic fibroblasts (MEFs) resulted in an aberrant DNA replication program showing an accumulation of late-S phase replication and causing severely defective growth. Furthermore, we found that the catalytic activity and replication focus targeting sequence of DNMT1 are required for a proper DNA replication program. Taken together, our findings suggest that the maintenance of DNA methylation by DNMT1 plays a critical role in proper regulation of DNA replication in mammalian cells.
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http://dx.doi.org/10.1016/j.bbrc.2015.12.090DOI Listing
January 2016

Regulation of maintenance DNA methylation via histone ubiquitylation.

J Biochem 2016 Jan 20;159(1):9-15. Epub 2015 Nov 20.

Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601, Japan

DNA methylation is one of the most stable but dynamically regulated epigenetic marks that act as determinants of cell fates during embryonic development through regulation of various forms of gene expression. DNA methylation patterns must be faithfully propagated throughout successive cell divisions in order to maintain cell-specific function. We have recently demonstrated that Uhrf1-dependent ubiquitylation of histone H3 at lysine 23 is critical for Dnmt1 recruitment to DNA replication sites, which catalyzes the conversion of hemi-methylated DNA to fully methylated DNA. In this review, we provide an overview of recent progress in understanding the mechanism underlying maintenance DNA methylation.
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http://dx.doi.org/10.1093/jb/mvv113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4882649PMC
January 2016

Uhrf1-dependent H3K23 ubiquitylation couples maintenance DNA methylation and replication.

Nature 2013 Oct 8;502(7470):249-53. Epub 2013 Sep 8.

Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, Nagoya 467-8601, Japan.

Faithful propagation of DNA methylation patterns during DNA replication is critical for maintaining cellular phenotypes of individual differentiated cells. Although it is well established that Uhrf1 (ubiquitin-like with PHD and ring finger domains 1; also known as Np95 and ICBP90) specifically binds to hemi-methylated DNA through its SRA (SET and RING finger associated) domain and has an essential role in maintenance of DNA methylation by recruiting Dnmt1 to hemi-methylated DNA sites, the mechanism by which Uhrf1 coordinates the maintenance of DNA methylation and DNA replication is largely unknown. Here we show that Uhrf1-dependent histone H3 ubiquitylation has a prerequisite role in the maintenance DNA methylation. Using Xenopus egg extracts, we successfully reproduce maintenance DNA methylation in vitro. Dnmt1 depletion results in a marked accumulation of Uhrf1-dependent ubiquitylation of histone H3 at lysine 23. Dnmt1 preferentially associates with ubiquitylated H3 in vitro though a region previously identified as a replication foci targeting sequence. The RING finger mutant of Uhrf1 fails to recruit Dnmt1 to DNA replication sites and maintain DNA methylation in mammalian cultured cells. Our findings represent the first evidence, to our knowledge, of the mechanistic link between DNA methylation and DNA replication through histone H3 ubiquitylation.
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http://dx.doi.org/10.1038/nature12488DOI Listing
October 2013

MCM8- and MCM9-deficient mice reveal gametogenesis defects and genome instability due to impaired homologous recombination.

Mol Cell 2012 Aug 5;47(4):523-34. Epub 2012 Jul 5.

DNA Replication and Genome Dynamics, Institute of Human Genetics, CNRS, 141 Rue de la Cardonille, 34396 Montpellier, Cedex 5, France.

We generated knockout mice for MCM8 and MCM9 and show that deficiency for these genes impairs homologous recombination (HR)-mediated DNA repair during gametogenesis and somatic cells cycles. MCM8(-/-) mice are sterile because spermatocytes are blocked in meiotic prophase I, and females have only arrested primary follicles and frequently develop ovarian tumors. MCM9(-/-) females also are sterile as ovaries are completely devoid of oocytes. In contrast, MCM9(-/-) testes produce spermatozoa, albeit in much reduced quantity. Mcm8(-/-) and Mcm9(-/-) embryonic fibroblasts show growth defects and chromosomal damage and cannot overcome a transient inhibition of replication fork progression. In these cells, chromatin recruitment of HR factors like Rad51 and RPA is impaired and HR strongly reduced. We further demonstrate that MCM8 and MCM9 form a complex and that they coregulate their stability. Our work uncovers essential functions of MCM8 and MCM9 in HR-mediated DSB repair during gametogenesis, replication fork maintenance, and DNA repair.
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http://dx.doi.org/10.1016/j.molcel.2012.05.048DOI Listing
August 2012

Xenopus laevis Ctc1-Stn1-Ten1 (xCST) protein complex is involved in priming DNA synthesis on single-stranded DNA template in Xenopus egg extract.

J Biol Chem 2012 Jan 14;287(1):619-627. Epub 2011 Nov 14.

Laboratory of Cell Cycle Regulation, Department of Gene Mechanisms, Graduate School of Biostudies, Kyoto University, Yoshida-Konoe-cho, Sakyo-ku, Kyoto 606-8501, Japan. Electronic address:

The Ctc1-Stn1-Ten1 (CST) complex is an RPA (replication protein A)-like protein complex that binds to single-stranded (ss) DNA. It localizes at telomeres and is involved in telomere end protection in mammals and plants. It is also known to stimulate DNA polymerase α-primase in vitro. However, it is not known how CST accomplishes these functions in vivo. Here, we report the identification and characterization of Xenopus laevis CST complex (xCST). xCST showed ssDNA binding activity with moderate preference for G (guanine)-rich sequences. xStn1-immunodepleted Xenopus egg extracts supported chromosomal DNA replication in in vitro reconstituted sperm nuclei, suggesting that xCST is not a general replication factor. However, the immunodepletion or neutralization of xStn1 compromised DNA synthesis on ssDNA template. Because primed ssDNA template was replicated in xStn1-immunodepleted extracts as efficiently as in control ones, we conclude that xCST is involved in the priming step on ssDNA template. These results are consistent with the current model that CST is involved in telomeric C-strand synthesis through the regulation of DNA polymerase α-primase.
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http://dx.doi.org/10.1074/jbc.M111.263723DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3249116PMC
January 2012

Essential roles of Xenopus TRF2 in telomere end protection and replication.

Genes Cells 2011 Jun 10;16(6):728-39. Epub 2011 May 10.

Laboratory of Cell Cycle Regulation, Department of Gene Mechanisms, Graduate School of Biostudies, Kyoto University, Yoshida-Konoe-cho, Sakyo-ku, Kyoto 606-8501, Japan.

TRF1 and TRF2 are double-stranded (ds) telomere DNA-binding proteins and the core members of shelterin, a complex that provides the structural and functional basis of telomere functions. We have reported that unlike mammalian TRF1 that constitutively binds to chromatin, Xenopus TRF1 (xTRF1) associates with mitotic chromatin but dissociates from interphase chromatin reconstituted in Xenopus egg extracts. This finding raised the possibility that xTRF1 and Xenopus TRF2 (xTRF2) contribute to telomere functions in a manner different from mammalian TRF1 and TRF2. Here, we focused on the role of xTRF2. We prepared chromatin reconstituted in egg extracts immunodepleted for xTRF2. Compared to mock-depleted nuclei, DNA damage response at telomeres was activated, and bulk DNAs were poorly replicated in xTRF2-depleted nuclei. The replication defect was rescued by inactivating ATR through the addition of anti-ATR neutralizing antibody, suggesting that ATR plays a role in the defect. Interestingly, the bulk DNA replication defect, but not the DNA damage response at telomeres, was rescued by supplementing the xTRF2-depleted extracts with recombinant xTRF2 (rTRF2). We propose that xTRF2 is required for both efficient replication of bulk DNA and protection from the activation of the DNA damage checkpoints pathway, and that those two functions are mechanistically separable.
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http://dx.doi.org/10.1111/j.1365-2443.2011.01520.xDOI Listing
June 2011

MCM-BP regulates unloading of the MCM2-7 helicase in late S phase.

Genes Dev 2011 Jan 31;25(2):165-75. Epub 2010 Dec 31.

Institute of Human Genetics, CNRS, 34396 Montpellier, France.

Origins of DNA replication are licensed by recruiting MCM2-7 to assemble the prereplicative complex (pre-RC). How MCM2-7 is inactivated or removed from chromatin at the end of S phase is still unclear. Here, we show that MCM-BP can disassemble the MCM2-7 complex and might function as an unloader of MCM2-7 from chromatin. In Xenopus egg extracts, MCM-BP exists in a stable complex with MCM7, but is not associated with the MCM2-7 hexameric complex. MCM-BP accumulates in nuclei in late S phase, well after the loading of MCM2-7 onto chromatin. MCM-BP immunodepletion in Xenopus egg extracts inhibits replication-dependent MCM dissociation without affecting pre-RC formation and DNA replication. When excess MCM-BP is incubated with Xenopus egg extracts or immunopurified MCM2-7, it binds to MCM proteins and promotes disassembly of the MCM2-7 complex. Recombinant MCM-BP also releases MCM2-7 from isolated late-S-phase chromatin, but this activity is abolished when DNA replication is blocked. MCM-BP silencing in human cells also delays MCM dissociation in late S phase. We propose that MCM-BP plays a key role in the mechanism by which pre-RC is cleared from replicated DNA in vertebrate cells.
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http://dx.doi.org/10.1101/gad.614411DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3022262PMC
January 2011

Initiation of DNA replication after fertilization is regulated by p90Rsk at pre-RC/pre-IC transition in starfish eggs.

Proc Natl Acad Sci U S A 2010 Mar 25;107(11):5006-11. Epub 2010 Feb 25.

Laboratory of Cell and Developmental Biology, Graduate School of Bioscience, Tokyo Institute of Technology, Yokohama 226-8501, Japan.

Initiation of DNA replication in eukaryotic cells is controlled through an ordered assembly of protein complexes at replication origins. The molecules involved in this process are well conserved but diversely regulated. Typically, initiation of DNA replication is regulated in response to developmental events in multicellular organisms. Here, we elucidate the regulation of the first S phase of the embryonic cell cycle after fertilization. Unless fertilization occurs, the Mos-MAPK-p90Rsk pathway causes the G1-phase arrest after completion of meiosis in starfish eggs. Fertilization shuts down this pathway, leading to the first S phase with no requirement of new protein synthesis. However, how and in which stage the initiation complex for DNA replication is arrested by p90Rsk remains unclear. We find that in G1-arrested eggs, chromatin is loaded with the Mcm complex to form the prereplicative complex (pre-RC). Inactivation of p90Rsk is necessary and sufficient for further loading of Cdc45 onto chromatin to form the preinitiation complex (pre-IC) and the subsequent initiation of DNA replication. However, cyclin A-, B-, and E-Cdk's activity and Cdc7 accumulation are dispensable for these processes. These observations define the stage of G1 arrest in unfertilized eggs at transition point from pre-RC to pre-IC, and reveal a unique role of p90Rsk for a negative regulator of this transition. Thus, initiation of DNA replication in the meiosis-to-mitosis transition is regulated at the pre-RC stage as like in the G1 checkpoint, but in a manner different from the checkpoint.
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http://dx.doi.org/10.1073/pnas.1000587107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2841915PMC
March 2010

[Replicative stress in telomeres].

Tanpakushitsu Kakusan Koso 2006 Nov;51(14 Suppl):2245-9

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November 2006

Cell-cycle-dependent regulation of telomere binding proteins: roles of Polo-like kinase.

Cell Cycle 2006 Jul 1;5(13):1403-6. Epub 2006 Jul 1.

Laboratory of Cell Cycle Regulation, Department of Gene Mechanisms, Graduate School of Biostudies, Kyoto University, Kyoto, Japan.

The telomere is a functional complex at chromosomal termini consisting of repetitive DNA and associated proteins, and protects the ends against degradation and fusion. Telomeric repeat binding factors TRF1 and TRF2 bind directly to double-stranded telomeric DNA. Although structurally related, TRF1 and TRF2 contribute to telomere maintenance in distinct ways: TRF1 negatively regulates telomerase-dependent telomere lengthening, whereas TRF2 plays an important role in protecting chromosomal ends. It is not known how the proteinaceous complex manages DNA metabolism such as DNA replication, which requires the recruitment of numerous trans-acting factors. We have found that Xenopus TRF1 (xTRF1) specifically associates with mitotic chromatin and dissociates from interphase replicating chromatin. In contrast, Xenopus TRF2 (xTRF2) binds to telomeric DNA throughout the cell cycle. Interestingly, telomerase activity is associated with the interphase chromatin, but not with the mitotic chromatin. These results support a model in which telomeres form a semi-open configuration that allows access of telomerase and replication machineries, yet protects the chromosomal ends in S phase. Interestingly, M phase specific telomere binding of xTRF1 requires Polo-like kinase, a key regulator of mitosis. We discuss the relevance of our studies and recent findings of other groups to indicate the possible role of Polo-like kinase in telomere regulation.
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http://dx.doi.org/10.4161/cc.5.13.2923DOI Listing
July 2006

Cell-cycle-dependent Xenopus TRF1 recruitment to telomere chromatin regulated by Polo-like kinase.

EMBO J 2006 Feb 19;25(3):575-84. Epub 2006 Jan 19.

Laboratory of Cell Cycle Regulation, Department of Gene Mechanisms, Graduate School of Biostudies, Kyoto University, Kitashirakawa-Oiwake-cho, Sakyo-ku, Kyoto, Japan.

Telomeres are regulated by a homeostatic mechanism that includes telomerase and telomeric repeat binding proteins, TRF1 and TRF2. Recently, it has been hypothesized that telomeres assume distinct configurations in a cell-cycle-dependent manner, although direct biochemical evidence is lacking. Here we demonstrated that Xenopus TRF1 (xTRF1) associates with telomere chromatin specifically in mitotic Xenopus egg extracts, and dissociates from it upon mitotic exit. Both the N-terminal TRF-homology (TRFH) domain and the linker region connecting the TRFH domain and the C-terminal Myb domain are required for this cell-cycle-dependent association of xTRF1 with chromatin. In contrast, Xenopus TRF2 (xTRF2) associates with chromatin throughout the cell cycle. We showed that Polo-like kinase (Plx1) phosphorylates xTRF1 in vitro. Moreover, the mitotic xTRF1-chromatin association was significantly impaired when Plx1 was immunodepleted from the extracts. Finally, high telomerase activities were detected in association with replicating interphase chromatin compared with mitotic chromatin. These results indicate that telomere chromatin is actively regulated by cell-cycle-dependent processes, and provide an insight for understanding how telomeres undergo DNA metabolisms during the cell cycle.
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http://dx.doi.org/10.1038/sj.emboj.7600964DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1383544PMC
February 2006