標的遺伝子に点突然変異をもつマウスの開発研究

2003 Fiscal Year Annual Research Report

• Project/Area Number: 15200032
• Research Category: Grant-in-Aid for Scientific Research (A)
• Research Institution: The Institute of Physical and Chemical Research
• Principal Investigator: 権藤 洋一 独立行政法人理化学研究所, 個体遺伝情報研究チーム, チームリーダー (40225678)
• Co-Investigator(Kenkyū-buntansha): 古関 明彦 千葉大学, 大学院・医学研究部, 教授 (40225446) ; 西田 圭吾 独立行政法人理化学研究所, サイトカインシステム情報制御研究チーム, 研究員 (80360618) ; 阿部 訓也 独立行政法人理化学研究所, 動物変異動態解析技術開発チーム, チームリーダー (40240915) ; 上野 直人 岡崎国立研究機構基礎生物学研究所, 発生生物学研究系, 教授 (40221105) ; 菅原 稔 東北大学, 大学院・医学系研究科, 助教授 (20311558)
• Keywords: ミュータジェネシス / マウス / モデル動物 / 標的遺伝子変異体 / ゲノム機能 / 遺伝子機能 / 突然変異誘発 / エチルニトロソウレア

Research Abstract

ゲノム上の任意の遺伝子に点突然変異をもつマウスを樹立し,その生物学的機能を解明する。高速汎用化することで、誰でも希望する遺伝子の点突然変異マウスを入手し独自の研究を発展展開できるよう基盤整備することを目的とする。初年度の今年は、まず,基盤となる精子凍結したENU誘発変異をもつG1オスマウス数が8000を越えた。発表されている中では世界最大規模である。また、それぞれの精巣からゲノムDNA抽出も進め、現在、6000匹を96プレート60枚分、変異検索ゲノムDNAアーカイブとして整備した。また、点突然変異を検索する標的として、初期発生関連、老化関連、免疫関連、ポリコーム群、PCP関連遺伝子において、検索するPCR領域の選定と、プライマー設計、構築、条件検討が進み、実際に変異DNAアーカイブのスクリーニングを開始した。精巣関連、DNA修復関連遺伝子については設定計画中であり、変異マウスの形態形成解析法整備も進んでいる。TGCE法を用いて、これまで8遺伝子から17の標的PCR領域についてのべ20Mbスクリーンした。11以上の点突然変異を検出し、ダイレクトシーケンシング法で塩基変化を確認したところ、アミノ酸置換を伴うもの4つ、スプライシングを変えるものひとつが含まれていた。この検出効率はダイレクトシーケンシングのみでスクリーンする方法に比べ感度が半分程度であり、見落としがあることが推定される。そこで、膨大なダイレクトシーケンシングデータを自動解析するプログラム開発も着手した。また、利用を外部にも呼びかけ、標的とする遺伝子総数も内外で120を越えた。遺伝子名や進捗をWEBサイトにおいて公開している(http://www.gsc.riken.jp/Mouse/)

Research Products

• [Publications] 権藤洋一: "モデル動物による遺伝子機能解明"Medical Science Digest. 29(7). 277-280 (2003)
• [Publications] 権藤洋一: "大規模マウスミュータジェネシス"Medical Science Digest. 30(2). 49-50 (2004)
• [Publications] Y.Gondo: "Megasatellite DNA"Nature Encyclopedia of the Human Genome. 3. 862-865 (2003)
• [Publications] T.Sagai: "Phylogenetic conservation of a limb-specific, cis-acting regulator of Sonic hedgehog (Shh)"Mammalian Genome. 15(1). 23-34 (2004)
• [Publications] B.Ohawara: "Role of glypican 4 in the regulation of convergent extension movements during gastrulation in Xenopus laevis."Development. 130. 2129-2138 (2003)
• [Publications] M.Tekeuchi: "Prickle-related gene in vertebrates is essential for gastrulation cell mevements."Curr.Biol.. 13. 674-679 (2003)
• [Publications] T.Ishitani: "he TAKI-NLK mitogen-activated protein kinase cascade functions in the Wnt-5a/Ca (2+) pathway to antagonize Wnt/beta-catenin signaling."Mol Cell Biol.. 23. 131-139 (2003)
• [Publications] M.Suzuki: "Hox proteins functionally cooperate with the GC box-binding protein system through distinct domains."J Biol Chem.. 278. 30148-30156 (2003)
• [Publications] N.Kinoshita: "PKCd is essential for Dishevelled function in a non-canonical Wnt pathway that regulates Xenopus convergent extension movements."Genes Dev.. 17. 1663-1676 (2003)
• [Publications] K.Azumi: "Construction of a cDNA microarray derived from the ascidian Ciona intestinalis."Zoolog Sci.. 20. 1223-1229 (2003)
• [Publications] K.Hotta: "A genome-wide survey of the genes for planar polarity signaling or convergent extension-related genes in Ciona intestinalis and phylogenetic comparisons of evolutionary conserved signaling components."Gene. 317. 165-185 (2003)
• [Publications] F.Carreira-Barbosa: "Prickle 1 regulates cell movements during gastrulation and neuronal migration in zebrafish."Development. 130. 4037-4046 (2003)
• [Publications] N.Ueno: "Planar cell polarity genes and neural tube closure."Birth Defects Res. Part C Embryo Today. 69. 318-324 (2004)
• [Publications] H.Iioka: "Essential role of MARCKS in cortical actin dynamics during gastrulation movements."J Cell Biol.. 164. 169-174 (2004)
• [Publications] 鈴木堅太郎: "マウス外生殖器原基(生殖結節)にみられる伸長制御メカニズム:BMP signalillgの機能解析"細胞工学. 23. 208-209 (2004)
• [Publications] 太田将: "ソノポレーション法によるニワトリ胚への遺伝子導入"細胞工学. 23(1). 94-99 (2004)
• [Publications] M.Yamada: "Silicon Compatible with the Height of Human Vertebral Column."Biological Trace Element Research. 95. 113-121 (2003)
• [Publications] K.Suzuki: "Regulation of outgrowth and apoptosis for the terminal appendage, external genitalia, development by concerted functions of Bmp signaling."Development. 131. 6209-6220 (2003)
• [Publications] S.Ohta: "Microbubble-Enhanced Sonoporation ; Efficient Gene Transduction Technique for Chick Embryos."Genesis. 37. 91-101 (2003)
• [Publications] G.Yamada: "Cellular and molecular mechanisms of development of the external Genitalia."Differentiation. 71. 445-460 (2003)
• [Publications] K.Suzuki: "Developmental biology meets with reproductive engineering ; interdiscipinar science area as a break through."Cell.Mol. Biol.. 49. 653-660 (2003)
• [Publications] I.Oishi: "The receptor tyrosine kinase Ror2 is involved in non-canonical Wnt5a/JNK signalling pathway."Genes Cells. 8. 645-654 (2003)
• [Publications] S.Yamazaki: "Mice with defects in HB-EGF ectodomain shedding show severe developmental abnormalities."J Cell Biol.. 163. 469-475 (2003)
• [Publications] H.Ozaki: "Six1 controls patterning of the mouse otic vesicle."Development. 131. 551-562 (2003)
• [Publications] Y.Nakaoka: "Activation of gp130 Transduces Hypertrophic Signal through Interaction of Scaffolding/Docking Protein Gab1 with Tyrosine Phosphatase SHP2 in Cardiomyocytes"Circulation Research. 93. 221-229 (2003)
• [Publications] S.Yamasaki: "Gads/Grb2-mediated association with LAT is critical for the inhibitory function of Gab2 in T cells."Mol Cell Biol.. 23. 2515-2529 (2003)
• [Publications] S.Yamasaki: "Gab1 is required for EGF receptor signaling and the transformation by activated ErbB2."Oncogene. 13. 1546-1556 (2003)
• [Publications] K.Nishida: "The role of Gab family scaffolding adapter proteins in the signal transduction of cytokine and growth factor receptors."Cancer Science. 94. 1029-1033 (2003)
• [Publications] T.Shirasawa: "Oxygen affinity of hemoglobin regulates O2 consumption, metabolism, and physical activity."J Biol Chem. 278. 5035-5043 (2003)
• [Publications] K.Fukuda: "Mesenchymal expression of Foxl1, a winged helix transcriptional factor, regulates generation and maintenance of gut associated lymphoid organs."Dev.Biol.. 255. 278-289 (2003)
• [Publications] T.Wang: "Forkhead transcription factor Foxf2 (LUN) - deficient mice exhibit abnormal development of secondaroy palate."Dev.Biol.. 259. 83-94 (2003)
• [Publications] R.Yamada: "Cell-autonomous involvement of Mab21/1 is essential for lens placode development."Development. 130. 1759-1770 (2003)
• [Publications] H.Miyagishima: "Dissociation of mammalian Polycomb-group proteins, Ring1B and Rae28/Ph1, from the chromatin correlates with configuration changes of the chromatin in mitotic and meiotic prophase."Histochem Cell Biol.. (in press). (2004)
• [Publications] M.Nakajima: "Abnormal blood Vessel development om moce lacking presenilin-1."Mech.Dev.. 120. 657-667 (2003)
• [Publications] Y.Nakamura: "Accumulation of p27^<KIPI> is associated with BMP2-induced grouth arrest and neuronal differentiation of human neuroblastoma-derived cell lines."BBRC.. 307. 206-213 (2003)
• [Publications] K.Murata: "CD69-null mice protected from arthritis induced with anti-typeII collagen antibodies."International Immunology. 15. 987-992 (2003)
• [Publications] Y.Hoki: "Growth retardation and skin abnormalities of the Recql4-deficient mouse."Human Mol.Genet.. 12. 2293-2299 (2003)
• [Publications] N.Okazaki: "Prediction of the coding sequences of mouse homologues of KIAA gene : III. the complete nucleotide sequences of 500 mouse KIAA-homologous cDNAs identified by screening of terminal sequences of cDNA clones randomly sampled from size-fractionated libraries."DNA Res.. 10(4). 167-180 (2003)
• [Publications] Y.Iwata: "Decomposition of hydroxylamine/water solution with added iron ion."J Hazard Mater.. 104(1-3). 39-49 (2003)
• [Publications] W.de Graaff: "Randomly inserted and targeted Hox/reporter fusions transcriptionally silenced in Polycomb mutants."Proc Natl Acad Sci U S A. 100(23). 13362-13367 (2003)
• [Publications] T.Kaneko: "The mouse YAF2 gene generates two distinct transcripts and is expressed in pre-and postimplantation embryos."Gene. 315. 183-192 (2003)
• [Publications] T.Shin: "Cooperative B7-1/2 (CD80/CD86) and B7-DC costimulation of CD4+ T cells independent of the PD-1 receptor."J Exp Med.. 198(1). 31-38 (2003)
• [Publications] A.Horie: "Generation of a monoclonal antibody against the mouse Sf3b1 (SAP155) gene product for U2 snRNP component of spliceosome."Hybrid Hybridomics.. 22(2). 117-119 (2003)
• [Publications] K.Yamamoto: "The KDEL receptor modulates the endoplasmic reticulum stress response through mitogen-activated protein kinase signaling cascades."J Biol Chem.. 278(36). 34525-34532 (2003)
• [Publications] M.Okazaki: "Increase in plasma fibrinogen and plasminogen activator inhibitor level in diabetic KK and KK-Ay mice."In Vivo.. 17(1). 5-11 (2003)
• [Publications] E.Kondo: "Expression of recombination-activating gene in mature peripheral T cells in Peyer's patch."Int Immunol.. 15(3). 393-402 (2003)
• [Publications] M.Kuwamura: "Rat mutations cvd and hob with cerebellar malformations map to Chromosome 2."Exp.Anim.. (in press). (2004)
• [Publications] E.Nakatsukasa: "Cryopreservation of spermatozoa from closed colonies, and inbred, spontaneous mutant, and transgenic strains of rats."Comp Med.. 53(6). 639-641 (2003)
• [Publications] M.Mori: "A new beige mutant rat ACI/N-Lystbg-Kyo"Exp Anim.. 52(1). 31-36 (2003)
• [Publications] A.S.Cachaco: "Knock-in of integrin β1D affects primmary but not secondary myogenesis in mice."Development. 130. 1659-1671 (2003)
• [Publications] T.Ara: "Impaired colonization of the gonads by primordial germ cells in mice lacking a chemokine, stromal cell-derived factor-1 (SDF-1)."Proceedings of the National Academy of Science of the USA. 100. 5319-5323 (2003)
• [Publications] K.Ohbo: "Identification and characterization of stem cells in prepubertal spermatogenesis in mice."Developmental Biology. 258. 209-225 (2003)
• [Publications] F.Santagati: "Identification of cis-regulatory elements in the mouse Pax9/Nkx2-9 genomic region : Implication for evolutionary conserved synteny."Genetics. 165. 235-242 (2003)
• [Publications] C.Kokubu: "Undulated short-tail deletion mutation in the mouse ablates Pax1 and leads to ectopic activation of neighboring Nkx2-2 in domains that normally express Pax1."Genetics. 165. 299-307 (2003)
• [Publications] Z.Li: "Defective smooth muscle development in qkI-deficient mice."Develop.Growth Diff.. 45. 449-462 (2003)
• [Publications] M.Sugimoto: "Tetraploid embryos rescue the early defects of t^<w5>/t^<w5> mouse embryos."Genesis. 37. 162-171 (2003)
• [Publications] M.Tsuda: "Conserved role of nanos proteins in germ cell development."Science. 301. 1239-1241 (2003)
• [Publications] K.Abe: "Gene content of the 750 kb critical region for mouse embryonic ectoderm lethal tcl-w5."Mamm.Genome. (in press). (2004)
• [Publications] K.Mitsunaga: "Loss of PGC-specific expression of the orphan nuclear receptor ERR- β results in reduction of germ cell number in mouse embryos."Mech.Dev.. (in press). (2004)
• [Publications] S.Yoshida: "neurogenin3 delineates the earliest stages of spermatogenesis in the mouse testis."Dev.Biol.. (in press). (2004)
• [Publications] 権藤洋一: "ゲノム創薬"(株)化学同人. 199 (2003)
• [Publications] 権藤洋一: "別冊・医学のあゆみ疾患モデル動物-病因解析での役割と限界"医歯薬出版(株). 130 (2004)