ゲノム分配を司るセントロメア機能

• Project/Area Number: 20247001
• Research Category: Grant-in-Aid for Scientific Research (A)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Genetics/Genome dynamics
• Research Institution: National Institute of Genetics
• Principal Investigator: 深川 竜郎 National Institute of Genetics, 教授 (60321600)
• Project Period (FY): 2008 – 2010
• Project Status: Completed (Fiscal Year 2011)
• Budget Amount: ¥48,100,000 (Direct Cost: ¥37,000,000、Indirect Cost: ¥11,100,000); Fiscal Year 2011: ¥13,260,000 (Direct Cost: ¥10,200,000、Indirect Cost: ¥3,060,000); Fiscal Year 2010: ¥13,260,000 (Direct Cost: ¥10,200,000、Indirect Cost: ¥3,060,000); Fiscal Year 2009: ¥13,260,000 (Direct Cost: ¥10,200,000、Indirect Cost: ¥3,060,000); Fiscal Year 2008: ¥8,320,000 (Direct Cost: ¥6,400,000、Indirect Cost: ¥1,920,000)
• Keywords: セントロメア / DT40細胞 / 染色体分配 / キネトコア / RNAiマシーナリー / CENPs / プロテオミクス / 細胞分裂

Research Abstract

生物の生命維持には、ゲノム情報が安定に保持・増殖されなければならない。ゲノム複製・分配のような基本的な生体反応に狂いが生じると染色体の異数化、がん化など細胞に対する悪影響が生じる。したがって、ゲノム複製や分配の正確な分子機構を解明することは、遺伝学における本質的な課題の一つである。本研究では、ゲノム分配を司るセントロメアに焦点を当て、本年度は以下の2課題についての研究を行い、成果を挙げた
1)セントロメアのクロマチンを構成する因子の同定および機能解析
21年度までの研究で、セントロメアのクロマチン領域に存在する複数のタンパク質を同定している。本年度は、セントロメアクロマチンの詳細について次世代シーケンサーなどを活用したChIP-seq解析を行い、クロマチン内の分布を明らかにした。また、複数のタンパク質についてDT40細胞を用いてノックアウト細胞を樹立して表現型を解析した。
2)キネトコアタンパク質複合体のin vitro再構成および構造解析
我々は、これまで約20種類程度のセントロメア構成タンパク質を同定し、それらの機能について遺伝学と細胞生物学的手法を併用して解析してきた。ある程度の機能分担が細胞レベルで理解できた後は、タンパク質のin vitroでの再構成および構造学的解析が必要になると考えた。昨年度は、CENP-T/CENP-W複合体の再構成を試み、成功した。本年度は、このin vitroで再構成したCENP-T/CENP-WがDNAあるいは再構成ヌクレオソームと形式するのか、あるいはどのように結合しているのかについて、スーパーコイリングアッセイなどで明らかにした。また、CENP-T/CENP-W複合体の結晶化に成功して、X線構造学的解析も行なった。

Research Products

• [Journal Article] Aurora B kinase controls the targeting of the Astrin/SKAP complex to bi-oriented kinetochores. (2010)
  • Author(s): Jens C.Schmidt
  • Journal Title: Journal of Cell Biology Volume: 119 Pages: 269-280
  • Peer Reviewed
• [Journal Article] The protein composition of mitotic chromosomes determined using multi-classifier combinatorial proteomics. (2010)
  • Author(s): Shinya Ohta
  • Journal Title: Cell Volume: 142 Pages: 810-821
  • Peer Reviewed
• [Journal Article] Chickens possess centromeres with both extended tandem repeats and short non-tandem-repetitive sequences. (2010)
  • Author(s): Wei-Hao Shang
  • Journal Title: Genome Research Volume: 20 Pages: 1219-1228
  • Peer Reviewed
• [Journal Article] A super-resolution map of the vertebrate kinetochore. (2010)
  • Author(s): Susana Abreu Ribeiro
  • Journal Title: Proceeding of National Academy of Science USA Volume: 107 Pages: 10484-10489
  • Peer Reviewed
• [Journal Article] Vertebrate Kinetochore Protein Architecture : Protein Copy Number. (2010)
  • Author(s): Katherine Johnston
  • Journal Title: Journal of Cell Biology Volume: 189 Pages: 937-943
  • Peer Reviewed
• [Journal Article] The KRAB zinc finger protein ZNF382 is a general, proapoptotic tumor suppressor repressing multiple oncogenes and frequently silenced in multiple carcinomas. (2010)
  • Author(s): Yingduan Cheng, Hua Geng
  • Journal Title: Cancer Research Volume: 70 Pages: 6516-6526
  • Peer Reviewed
• [Journal Article] Aurora B phosphorylates spatially distinct targets to differentially regulate the kinetochore-microtubule interface. (2010)
  • Author(s): Julie P.I.Welburn
  • Journal Title: Molecular Cell Volume: 38 Pages: 383-392
  • Peer Reviewed
• [Journal Article] Identification and characterization of two isoforms of the vertebrate H2A.Z histone variant. (2010)
  • Author(s): Ryo Matsuda
  • Journal Title: Nucleic Acids Research Volume: 38 Pages: 4263-4273
  • Peer Reviewed
• [Journal Article] Regulated targeting of protein phosphatase 1 to the outer kinetochore by KNL1 opposes Aurora B kinase. (2010)
  • Author(s): Dan Liu
  • Journal Title: Journal of Cell Biology Volume: 188 Pages: 809-820
  • Peer Reviewed
• [Journal Article] INCENP-aurora B interactions modulate kinase activity and chromosomal passenger complex localization (2009)
  • Author(s): Zhenjie Xu
  • Journal Title: Journal of Cell Biology 187 Pages: 637-653
  • Peer Reviewed
• [Journal Article] An auxin-based degron system for the rapid deletion of proteins in nonplant cells (2009)
  • Author(s): Kohei Nishimura
  • Journal Title: Nature Methods 6 Pages: 917-922
  • Peer Reviewed
• [Journal Article] CENP-H-containing complex facitates centromere deposition of CENP-A in cooperation with FACT and CHD1 (2009)
  • Author(s): Masahiro Okada
  • Journal Title: Molecular Biology of the Cell 20 Pages: 3986-3995
  • Peer Reviewed
• [Journal Article] The CENP-S complex is essential for the stable assembly of outer kineto chore structure (2009)
  • Author(s): Miho Amano
  • Journal Title: Journal of Cell Biology 186 Pages: 173-182
  • Peer Reviewed
• [Journal Article] Two Distinct Repeat Sequences of Nup98 Nucleoporins Characterize Dual Nuclei in the Binucleated Ciliate Tetrahymena (2009)
  • Author(s): Masaaki Iwamoto
  • Journal Title: Current Biology 19 Pages: 843-847
  • Peer Reviewed
• [Journal Article] A Rac GTPase activating protein MgcRacGAP is an NLS-containing nuclear chaperone in the activation of STAT transcription factors (2009)
  • Author(s): Toshiyuki Kawashima
  • Journal Title: Molecular and Cellular Biology 29 Pages: 1796-1813
  • Peer Reviewed
• [Journal Article] Cohesin associates with spindle poles in a mitosis-specific manner and functions in spindle assemble in vertebrate cells (2009)
  • Author(s): Xiangduo Kong
  • Journal Title: Molecular Biology of the Cell 20 Pages: 1289-1301
  • Peer Reviewed
• [Journal Article] Role of glucocorticoid receptor in the regulation of cellular sensitivity to irinotec an hydrochloride (2009)
  • Author(s): Takanori Akagi
  • Journal Title: Journal of Pharmacological Sciences 109 Pages: 265-274
  • Peer Reviewed
• [Journal Article] Cohesin Associates with Spindle Poles in a Mitosis-specific Manner and Functions in Spindle Assembly in Vertebrate Cells (2009)
  • Author(s): Xiangduo Kong
  • Journal Title: Molecular Biology of the Cell 20 Pages: 1289-1301
  • Peer Reviewed
• [Journal Article] A Rac GTPase-Activating Protein, MgcRacGAP, Is a Nuclear Localizing Signal-Containing Nuclear Chaperone in the Activation of STAT Transcription Factors (2009)
  • Author(s): Toshiyuki Kawashima
  • Journal Title: Molecular and Cellular Biology 29 Pages: 1796-1813
  • Peer Reviewed
• [Journal Article] CCAN makes multiple contacts with centromeric DNA to provide distinct pathways to the outer kinetochore (2008)
  • Author(s): Tetsuya Hori
  • Journal Title: Cell 135 Pages: 1039-1052
  • Peer Reviewed
• [Presentation] Structural dynamics of inner-kinetochore for faithful chromosome segregation (2010)
  • Author(s): Tatsuo Fukagawa
  • Organizer: BMB2010第33回日本分子生物学会年会・第83回日本生化学会大会合同大会
  • Place of Presentation: 神戸ポートアイランド(神戸)
  • Year and Date: 2010-12-09
• [Presentation] Structural dynamics of inner-kinetochore for faithful chromosome segregation (2010)
  • Author(s): Tatsuo Fukagawa
  • Organizer: EMBO workshop on chromosome segregation and aneuploidy
  • Place of Presentation: Royal College of Surgeons, Edinburgh, UK
  • Year and Date: 2010-06-19
• [Presentation] 染色体分配におけるインナーキネトコアの構造変化 (2010)
  • Author(s): 深川竜郎
  • Organizer: 第62回日本細胞生物学会シンポジウム
  • Place of Presentation: 大阪国際会議場(大阪)
  • Year and Date: 2010-05-19
• [Presentation] 動物細胞のキネトコア構造 (2010)
  • Author(s): 深川竜郎
  • Organizer: 構造エピゲノム研究会 発足記念シンポジウム
  • Place of Presentation: 理化学研究所(横浜)
  • Year and Date: 2010-04-30
• [Presentation] Molecular structure of the vertebrate kinetochore (2009)
  • Author(s): 深川竜郎
  • Organizer: 日本分子生物学会
  • Place of Presentation: パシフィコ横浜(横浜)
  • Year and Date: 2009-12-11
• [Presentation] セントロメアを形成するために必要なヌクレオソーム構造 (2009)
  • Author(s): 深川竜郎
  • Organizer: 日本生化学会
  • Place of Presentation: 神戸国際会議場(神戸)
  • Year and Date: 2009-10-23
• [Presentation] 核内ダイナミクスの基盤となるセントロメア構造 (2009)
  • Author(s): 深川竜郎
  • Organizer: 日本遺伝学会
  • Place of Presentation: 信州大学(松本)
  • Year and Date: 2009-09-17
• [Presentation] Molecular architecture of the vertebrate constitutive centromere associated network (2009)
  • Author(s): 深川竜郎
  • Organizer: The 17th International Chromosome Conference
  • Place of Presentation: Boone, NC, USA
  • Year and Date: 2009-06-25
• [Presentation] CENP-Aのセントロメア局在化の関与する因子の機能解析 (2008)
  • Author(s): 岡田聖裕
  • Organizer: BMB2008(第31会日本分子生物学会シンポジウム)
  • Place of Presentation: 神戸
  • Year and Date: 2008-12-11
• [Presentation] セントロメアDNAに結合するCENP-T複合体の機能解析 (2008)
  • Author(s): 堀哲也
  • Organizer: MB2008(第31会日本分子生物学会シンポジウム)
  • Place of Presentation: 神戸
  • Year and Date: 2008-12-11
• [Presentation] セントロメア特異的クロマチン構造の構築 (2008)
  • Author(s): 深川竜郎
  • Organizer: BMB2008(第31会日本分子生物学会シンポジウム)
  • Place of Presentation: 神戸
  • Year and Date: 2008-12-10
• [Presentation] Molecular architecture of the vertebrate constitutive centromere associated network (2008)
  • Author(s): Tatsuo Fukagawa
  • Organizer: EMBO workshop on Chromosome Segregation : Centromers and Kinetochores
  • Place of Presentation: Bordeaux
  • Year and Date: 2008-09-28
• [Presentation] インナーキネトコアの分子構築 (2008)
  • Author(s): 深川竜郎
  • Organizer: 第60回日本細胞生物学会シンポジウム
  • Place of Presentation: 横浜
  • Year and Date: 2008-06-29
• [Presentation] Molecular architecture of the vertebrate constitutive centromere associated network (2008)
  • Author(s): Tatsuo Fukagawa
  • Organizer: International Symposium on chromosome dynamics in Ise
  • Place of Presentation: Ise
  • Year and Date: 2008-05-28
• [Presentation] Functional analysis of CENP-O class kinetochore proteins (2008)
  • Author(s): Tetsuya Hori
  • Organizer: The 3rd International Symposium on Chromo some Dynamics
  • Place of Presentation: Mishima
  • Year and Date: 2008-05-26
• [Book] 細胞工学「動原体の分子構築」 (2010)
  • Author(s): 竹内康造, 深川竜郎
  • Total Pages: 7
  • Publisher: 秀潤社