ニューロン・グリアの分化決定とニューロンの個性獲得にいたる制御機構の解析

2003 年度 実績報告書

• 研究課題/領域番号: 12210013
• 研究機関: 慶應義塾大学
• 研究代表者: 岡野 栄之 慶應義塾大学, 医学部, 教授 (60160694)
• キーワード: 脳神経系 / 神経幹細胞 / Notchシグナル / Musashi1 / 神経分化 / m-Numb / Neurosphere / 再生医学

研究概要

Musashi1(Msi1)蛋白質は、神経幹細胞を含む神経系の前駆細胞に発現するRNA結合蛋白質であり、Notchシグナルの阻害蛋白質であるm-Numbの翻訳抑制を行っている。本年度において、Msi1を含む蛋白質複合体の構成要素同定を行った。293T細胞またはP19細胞内においてMsi1蛋白質と結合している蛋白質分子の濃縮・精製をTAP (Tandem Affinity Purification)法により行い、二種の蛋白質をMALDI-TOF MS法で解析した結果、PolyA Binding Protein (PABP)、IGF2 mRNA Binding Protein (IMP)が同定された。PABIPはeIF4G, eIF4E, mRNA5'-capを介した翻訳開始複合体に含まれることから、Msi1とPABPの結合が翻訳促進時の翻訳複合体形成を不安定化させる可能性を示唆した(投稿論文準備中)。また現在、ヒト・マウスのmsi1遺伝子間で相同性が非常に高い塩基配列領域を複数発見し、それらの領域をEGFPのレポーター遺伝子に連結したコンストラクトを作成し、ニワトリ胚中枢神経系形成部位にelectroporation法により遺伝子導入する系を用いてmusashi1遺伝子の転写活性化領域の同定を行っている。
Msi1陽性の神経幹細胞の未分化性維持・増殖に必須であるNotchの活性化は、Delta, Jaggedなどのligand依存的に膜直下で切断されることが必要である。本年度、この件について我々は、切断されたNotch1の細胞内領域のN末端を認識する抗体anti-actN1を用いて、発生期のマウス脳の免疫組織化学染色を行い、時空間的なNotch1の活性化状態を可視化し、組織図で示した(Tokunaga et al., J. Neuro. Chem. in press)。

研究成果

• [文献書誌] Tonchev AB, Okano, H, et al.: "Proliferation of neural and neuronal progenitors after global brain ischemia in young adult macaque monkeys."Mol Cell Neurosci. 23. 292-301 (2003)
• [文献書誌] Kayahara T, Okano H, et al.: "Candidate markers for stem and early progenitor cells, Musashi-1 and Hes1, are expressed in crypt base columnar cells of mouse small intestine."FEBS Letter. 535. 131-135 (2003)
• [文献書誌] Sasaki T, Okano H, et al.: "Implication of Cyclooxygenase-2 on Enhanced Proliferation of Neural Progenitor Cells in the Adult Mouse Hippocampus After Ischemia."J Neurosci Res. 72. 461-471 (2003)
• [文献書誌] Nakamura Y, Okano H, et al.: "Expression of tubulin beta II in neural stem/progenitor cells and radial fibers during human fetal brain development"Lab Invest. 83. 479-489 (2003)
• [文献書誌] Yuasa Y, Okano H, et al.: "Drosophila homeodomain protein REPO controls glial differentiation by cooperating with ETS and BTB transcription factors."Development. 130. 2419-2428 (2003)
• [文献書誌] Uchida K, Okano H, et al.: "Grafted swine neuroepithelial stem cells can form myelinated axons and both efferent and afferent synapses with xenogeneic rat neurons."J Neurosci Res. 72. 661-669 (2003)
• [文献書誌] Kokuzawa J, Okano H, et al.: "Hepatocyte growth factor promotes proliferation and neural differentiation of neural stem cells from mouse embryos."Mol Cell Neurosci. 24. 190-197 (2003)
• [文献書誌] Ishizuya-Oka A, Okano H, et al.: "Thyroid hormone-upregulated expression of Musashi-1 is specific for progenitor cells of the adult epithelium during amphibian gastrointestinal remodeling."J Cell Sci. 116. 3157-3164 (2003)
• [文献書誌] Kanuka H, Okano H, et al.: "Cytosol-endoplasmic reticulum interplay by Sec61alpha translocon in polyglutamine-mediated neurotoxicity in Drosophila."Proc Natl Acad Sci USA. 100. 11723-11728 (2003)
• [文献書誌] Baker H, Okano H, et al.: "Cortical and striatal expression of tyrosine hydroxylase mRNA in neonatal and adult mice."Cell Mol Neurobiol. 23. 503-518 (2003)
• [文献書誌] Tamaki T, Okano H, et al.: "Growth and differentiation potential of main-and side-population cells derived from murine skeletal muscle."Exp Cell Res. 291. 83-90 (2003)
• [文献書誌] Miyanomori Y, Okano H, et al.: "Origin of figher affinity to RNA of the N-terminal RNA-binding domain than that of the C-terminal one of a mouse neural protein, musashi1, as revealed by comparison of their structures, modes of interaction, surface electrostatic potentials, and backbone dynamics."J Biol Chem. 278. 41309-41315 (2003)
• [文献書誌] Yoshida T, Okano, H: "Distinct expression patterns of splicing isoforms of mNumb in the endocrine lineage of developing pancreas."Differentiation. 71. 486-495 (2003)
• [文献書誌] Hu QD, Okano H, et al.: "F3/Contactin acts as a functional ligand for Notch during oligodendrocyte maturation."Cell. 535. 131-135 (2003)
• [文献書誌] Matsuzaki Y, Kinjo K, Mulligan RC, Okano H et al.: "Drosophila homeodomain protein REPO controls glial differentiation by cooperating with ETS and BTB transcription factors."Development. 130. 2419-2428 (2003)
• [文献書誌] Uchida K, Okano H, et al.: "Grafted swine neuroepithelial stem cells can form myelinated axons and both efferent and afferent synapses with xenogeneic rat neurons."J Neurosci Res. 72. 661-669 (2003)
• [文献書誌] Okada S, Okano H, et al.: "Implication of Cyclooxygenase-2 on Enhanced Proliferation of Neural Progenitor Cells in the Adult Mouse Hippocampus After Ischemia."J Neurosci Res. (In press). (2004)
• [文献書誌] Yamashima T, Okano H et al.: "Vascular adventitia generates neuronal progenitors in monkey hippocampus after ischemia."Hippocampus. (In press). (2004)
• [文献書誌] Ieda M, Okano H, et al.: "Distinct expression patterns of splicing isoforms of mNumb in the endocrine lineage of developing pancreas."Differentiation. (In press). (2004)
• [文献書誌] Watanabe K, Okano H, et al.: "Comparison between fetal spinal cord-and forebrain-derived neural stem/progenitor cells as a source of transplantation for spinal cord injury."J Neurosci Res. (In press). (2004)
• [文献書誌] Ohba H, Okano H, et al.: "Sox21 is a repressor of neuronal differentiation and is antagonized by YB-1."Neurosci Lett. (In press). (2004)
• [文献書誌] Mikami Y, Okano H, et al.: "Implantation of dendritic cells in the injured adult spinal coed results in activation of the endogenous neural/progenitor cells for den novo neurogenesis and axonal regeneration, leading to functional recovery."J Neurosci Res. (In press). (2004)
• [文献書誌] Ozawa Y, Okano H, et al.: "Down-regulation of STAT3 activation is required as an intrinsic factor to complete rod photoreceptor differentiation."Mol Cell Neurosci. (In press). (2004)
• [文献書誌] Tokunaga A, Okano H et al.: "Mapping spatio-temporal activation of Notch signaling during neurogenesis and gliogenesis in the developing mouse brain."J Neurochem. (In press). (2004)