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ITO Keisuke
Graduate School of Medicine / Faculty of Medical Sciences
Assistant Professor

Researcher basic information

■ Research Areas
  • Life sciences / Developmental biology

Research activity information

■ Award
  • Jan. 2014 理化学研究所, 理化学研究所 基礎科学・国際特別研究員 研究成果発表会ポスター賞, Analysis of Physiological Meanings of Intracellular Trafficking of the Neurotrophic Receptor, RET – Focusing on Development of the Enteric Nervous System
    Ito Keisuke
    Others

■ Paper
  • Bayu Pratama Putra, Keisuke Ito, Carla Cirillo, Mukhamad Sunardi, Haruhiko Koseki, Toshihiro Uesaka, Hideki Enomoto
    The enteric nervous system (ENS) regulates gut functions independently from the central nervous system (CNS) by its highly autonomic neural circuit that integrates diverse neuronal subtypes. Although several transcription factors are shown to be necessary for the generation of some enteric neuron subtypes, the mechanisms underlying neuronal subtype specification in the ENS remain elusive. In this study, we examined the biological function of Polycomb group RING finger protein 1 (PCGF1), one of the epigenetic modifiers, in the development and differentiation of the ENS by disrupting the Pcgf1 gene selectively in the autonomic-lineage cells. Although ENS precursor migration and enteric neurogenesis were largely unaffected, neuronal differentiation was impaired in the Pcgf1-deficient mice, with the numbers of neurons expressing somatostatin (Sst+ ) decreased in multiple gut regions. Notably, the decrease in Sst+ neurons was associated with the corresponding increase in calbindin+ neurons in the proximal colon. These findings suggest that neuronal subtype conversion may occur in the absence of PCGF1 and that epigenetic mechanism is primarily involved in specification of some enteric neuron subtypes. This article is protected by copyright. All rights reserved.
    Jul. 2023, Development, growth & differentiation, English, Domestic magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Mukhamad Sunardi, Keisuke Ito, Yuya Sato, Toshihiro Uesaka, Mitsuhiro Iwasaki, Hideki Enomoto
    BACKGROUND & AIMS: Hirschsprung disease (HSCR) is a congenital disorder characterized by the absence of the enteric nervous system (ENS). HSCR potentially involves multiple gene aberrations and displays complex patterns of inheritance. Mutations of the RET gene, encoding the RET receptor tyrosine kinase, play a central role in the pathogenesis of HSCR. Although a wide variety of coding RET mutations have been identified, their pathogenetic significance in vivo has remained largely unclear. METHODS: We introduced a HSCR-associated RET missense mutation, RET(S811F), into the corresponding region (S812) of the mouse Ret gene. Pathogenetic impact of Ret(S812F) was assessed by histologic and functional analyses of the ENS and by biochemical analyses. Interactions of the Ret(S812F) allele with HSCR susceptibility genes, the RET9 allele and the Ednrb gene, were examined by genetic crossing in mice. RESULTS: RetS812F/+ mice displayed intestinal aganglionosis (incidence, 50%) or hypoganglionosis (50%), impaired differentiation of enteric neurons, defecation deficits, and increased lethality. Biochemical analyses revealed that Ret(S811F) protein was not only kinase-deficient but also abrogated function of wild-type RET in trans. Moreover, the Ret(S812F) allele interacted with other HSCR susceptibility genes and caused intestinal aganglionosis with full penetrance. CONCLUSIONS: This study demonstrates that a single RET missense mutation alone induces intestinal aganglionosis via a dominant-negative mechanism. The RetS812F/+ mice model HSCR displays dominant inheritance with incomplete penetrance and serves as a valuable platform for better understanding of the pathogenetic mechanism of HSCR caused by coding RET mutations.
    Elsevier BV, Dec. 2022, Cellular and Molecular Gastroenterology and Hepatology, 15(6) (6), 1505 - 1524, English, International magazine
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Mukhamad Sunardi, Keisuke Ito, Hideki Enomoto
    The GDNF Family Ligands (GFLs) regulate neural development and kidney organogenesis by activating the RET receptor tyrosine kinase. Many RET-dependent developmental processes involve long-distance cell-cell communications or cell polarity, which includes cell migration and axon guidance. This suggests that spatiotemporally regulated subcellular localization of RET protein and appropriate propagation of RET signaling in cells are essential for the physiological function of the GFLs. Little is known, however, about the dynamics of RET protein in cells. Addressing this issue requires development of a system that allows visualization of RET in living cells. In this study, we report generation of a novel knock-in mouse line in which the RET-EGFP chimeric receptor is expressed under the Ret promoter. Unlike Ret-deficient mice that die after birth due to the absence of the enteric nervous system (ENS) and kidneys, RetRET-EGFP/RET-EGFP mice were viable and grew to adulthood with no overt abnormality, which indicated that RET-EGFP exerts function comparable to RET. In neurons and ENS progenitors, RET-EGFP signals were detected both on the cell membrane and in the cytoplasm, the latter of which appeared as a punctate pattern. Time-lapse imaging of cultured neural cells and embryos revealed active transport of RET-EGFP puncta in neuronal axons and cell bodies. Immunohistochemical analyses detected RET-EGFP signals in early and recycling endosomes, indicating that RET-EGFP is trafficked via the endocytic pathway. RetRET-EGFP/RET-EGFP mice enable visualization of functional RET protein in vivo for the first time and provide a unique platform to examine the dynamics and physiology of RET trafficking.
    Lead, Wiley, Aug. 2021, Development, Growth & Differentiation, 63(6) (6), 285 - 294, English, Domestic magazine
    [Refereed]
    Scientific journal

  • Mitsumasa Okamoto, Toshihiro Uesaka, Keisuke Ito, Hideki Enomoto
    Society for Neuroscience, May 2021, eneuro, 8(3) (3), ENEURO.0534 - 20.2021, English
    [Refereed]
    Scientific journal
    Link to Kobe Univ. Repository (Kernel)

  • Keisuke Ito, Hideki Enomoto
    Lead, Oxford University Press (OUP), Aug. 2016, Journal of Biochemistry, 160(2) (2), 77 - 85, English
    [Refereed]
    Scientific journal

  • Goto A, Sumiyama K, Kamioka Y, Nakasho E, Ito K, Iwasaki M, Enomoto H, Matsuda M
    2013, Journal of Neuroscience, 33(11) (11), 4901 - 4912, English
    [Refereed]
    Scientific journal

  • Keisuke Ito, Takahiko Kawasaki, Seiji Takashima, Ikuo Matsuda, Atsu Aiba, Tatsumi Hirata
    Ventral tangential migration of neurons is the most prominent mode of neuronal translocation during earliest neurogenesis in the mouse telencephalon. A typical example of the neurons that adopt this migration mode is guidepost neurons in the lateral olfactory tract designated as lot cells. These neurons are generated from the neocortical neuroepithelium and migrate tangentially down to the ventral edge of the neocortex abutting the ganglionic eminence, on which the future lateral olfactory tract develops. We show here that this migration stream is repelled by a secreted axon guidance molecule, semaphorin 3F through interaction with its specific receptor, neuropilin-2. Accordingly, in mutant mice for semaphorin 3F or neuropilin-2, lot cells ectopically penetrated into the deep brain domain, which normally expresses semaphorin 3F. These results reveal that semaphorin 3F is an important regulator of the ventral tangential migration stream, confining the migrating neurons on the telencephalon surface by repelling from the deeper domain.
    Lead, Apr. 2008, The Journal of neuroscience : the official journal of the Society for Neuroscience, 28(17) (17), 4414 - 22, English, International magazine
    [Refereed]
    Scientific journal

  • Netrin 1 regulates ventral tangential migration of guidepost neurons in the lateral olfactory tract.
    Takahiko Kawasaki, Keisuke Ito, Tatsumi Hirata
    In the developing nervous system, functional neural networks are constructed with intricate coordination of neuronal migrations and axonal projections. We have previously reported a ventral tangential migration of a special type of cortical neurons, lot cells, in the mouse embryo. These neurons originate from the ventricular zone of the entire neocortex, tangentially migrate in the surface layer of the neocortex into the ventral direction, align in the future pathway of the lateral olfactory tract (LOT) and eventually guide the projection of LOT axons. In this study, we developed an organotypic culture system to investigate the regulation of this cell migration in the developing telencephalon. Our data show that the neocortex contains the signals that direct lot cells ventrally, that the ganglionic eminence excludes lot cells by repelling the migration and that lot cells are attracted to netrin 1, an axon guidance factor. Furthermore, we demonstrate that mutations in the genes encoding netrin 1 and its functional receptor Dcc lead to inappropriate distribution of lot cells and subsequent partial disruption of LOT projection. These results suggest that netrin 1 regulates the migration of lot cells and LOT projections, possibly by ensuring the correct distribution of these guidepost neurons.
    Mar. 2006, Development (Cambridge, England), 133(5) (5), 845 - 53, English, International magazine
    [Refereed]

  • Plexin-a4 mediates axon-repulsive activities of both secreted and transmembrane semaphorins and plays roles in nerve fiber guidance.
    Fumikazu Suto, Keisuke Ito, Masato Uemura, Masayuki Shimizu, Yutaka Shinkawa, Makoto Sanbo, Tomoyasu Shinoda, Miu Tsuboi, Seiji Takashima, Takeshi Yagi, Hajime Fujisawa
    It has been proposed that four members of the plexin A subfamily (plexin-As; plexin-A1, -A2, -A3, and -A4) and two neuropilins (neuropilin-1 and neuropilin-2) form complexes and serve as receptors for class 3 secreted semaphorins (Semas), potent neural chemorepellents. The roles of given plexin-As in semaphorin signaling and axon guidance, however, are mostly unknown. Here, to elucidate functions of plexin-A4 in semaphorin signaling and axon guidance events in vivo, we generated plexin-A4 null mutant mice by targeted disruption of the plexin-A4 gene. Plexin-A4 mutant mice were defective in the trajectory and projection of peripheral sensory axons and sympathetic ganglion (SG) axons and the formation of the anterior commissure and the barrels. The defects in peripheral sensory and SG axons were fundamentally related to those of neuropilin-1 or Sema3A mutant embryos reported but were more moderate than the phenotype in these mutants. The growth cone collapse assay showed that dorsal root ganglion axons and SG axons of plexin-A4 mutant embryos partially lost their responsiveness to Sema3A. These results suggest that plexin-A4 plays roles in the propagation of Sema3A activities and regulation of axon guidance and that other members of the plexin-A subfamily are also involved in the propagation of Sema3A activities. Plexin-A4-deficient SG axons did not lose their responsiveness to Sema3F, suggesting that plexin-A4 serves as a Sema3A-specific receptor, at least in SG axons. In addition, the present study showed that plexin-A4 bound class 6 transmembrane semaphorins, Sema6A and Sema6B, and mediated their axon-repulsive activities, independently of neuropilin-1. Our results imply that plexin-A4 mediates multiple semaphorin signals and regulates axon guidance in vivo.
    Lead, Apr. 2005, The Journal of neuroscience : the official journal of the Society for Neuroscience, 25(14) (14), 3628 - 37, English, International magazine
    [Refereed]

■ MISC
  • Repulsive effect of Sema3F on lot cells, the guidepost neurons in the lateral olfactory tract
    Keisuke Ito, Takahiko Kawasaki, Tatsurni Hirata
    Lead, 2007, NEUROSCIENCE RESEARCH, 58, S38 - S38, English
    Summary international conference

  • Effect of a protein kinase inhibitor, K252a on migratory neurons
    Keisuke Ito, Takahiko Kawasaki, Tatsumi Hirata
    Lead, 2006, NEUROSCIENCE RESEARCH, 55, S216 - S216, English
    Summary international conference

■ Lectures, oral presentations, etc.
  • RET 活性化型変異 C618F は遺伝子量減少によりヒルシュスプルング病を誘導する
    ITO KEISUKE, 岡本 光正, UESAKA TOSHIHIRO, 前田 貢作, ENOMOTO HIDEKI
    CSMIリトリート「若手道場」, Jan. 2019, Japanese, 淡路, Domestic conference
    Oral presentation

  • Elevated levels of RET signaling coupled with decreased RET dose causes Hirschsprung's disease in mice
    ITO KEISUKE
    第2回神戸理研・神戸大学合同シンポジウム, Jan. 2019, English, 神戸, Domestic conference
    Poster presentation

  • RET 活性化型変異 C618F は遺伝子量減少によりヒルシュスプルング病を誘導する
    ITO KEISUKE, 岡本 光正, UESAKA TOSHIHIRO, 前田 貢作, ENOMOTO HIDEKI
    第94回日本解剖学会近畿支部学術集会, Nov. 2018, Japanese, 神戸, Domestic conference
    Oral presentation

  • CRISPR/Cas9システムによる遺伝子ノックアウト研究の進展と今後の課題
    ITO KEISUKE
    Neuroscience network in Kobe 定例研究会, Sep. 2018, Japanese, 神戸, Domestic conference
    Oral presentation

  • Polarized intracellular trafficking of RET tyrosine kinase in living neurons
    Ito Keisuke, Enomoto Hideki
    The 38th Annual Meeting of the Japan Neuroscience Society, Jul. 2015, English, 日本神経科学, 神戸, Domestic conference
    Poster presentation

  • Polarized trafficking of RET tyrosine kinase in migrating enteric-neural crestderived cells
    Ito Keisuke, Enomoto Hideki
    4th International Symposium on "Development of the enteric nervous system:Cells,Singnals,Genes and Therapy", Apr. 2015, English, ENS meeting scientific organizing committee:Robert Hofstra,Alan Burns(local organizer), Rotterdam, The Netherlands, International conference
    Poster presentation

  • Polarized trafficking of RET tyrosine kinase in living cells
    Ito Keisuke, Enomoto Hideki
    Neurovascular Wiring 2nd International Symposium 2015, Jan. 2015, English, 新学術領域研究「血管-神経ワイヤリングにおける相互依存性の正立機構」, 京都, Domestic conference
    Poster presentation

  • Polarized trafficking of RET tyrosine kinase in living neurons
    Ito Keisuke
    第2回神緑会ヤングイベスティゲーターアワード発表会, Oct. 2014, Japanese, Kobe University School of Medicine, 神戸, Domestic conference
    Poster presentation

  • Analysis of Physiological Meanings of Intracellular Trafficking of the Neurotrophic Receptor, RET – Focusing on Development of the Enteric Nervous System
    Ito Keisuke, Enomoto Hideki
    RIKEN Special Postdoctoral Researchers Program & Foreign Postdoctoral Researchers Program, Poster Session of Research Results, Jan. 2014, English, RIKEN, 和光, Wako, Domestic conference
    Poster presentation

  • Intracellular Trafficking of a GDNF Receptor, RET- Roles in Neural Development? -
    Ito Keisuke, Enomoto Hideki
    Neuro2013, Jun. 2013, English, The Japan Neuroscience Society, The Japanese Society for Neurochemistry, Japanese Neural Network Society, 京都, Domestic conference
    Oral presentation

■ Research Themes
  • 前肢に存在する「神経様細胞」の異常産生と、神経芽腫発症の共通分子基盤を探る
    伊藤 圭祐
    日本学術振興会, 科学研究費助成事業 基盤研究(C), 基盤研究(C), 神戸大学, 01 Apr. 2022 - 31 Mar. 2025

  • Analysis of multipotency and its underlying molecular mechanisms of sympathetic nerve-associating Schwann cell precursors
    伊藤 圭祐
    Japan Society for the Promotion of Science, Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C), Grant-in-Aid for Scientific Research (C), Kobe University, 01 Apr. 2019 - 31 Mar. 2022

  • 伊藤 圭祐
    学術研究助成基金助成金／若手研究(B), Apr. 2015 - Mar. 2017, Principal investigator
    Competitive research funding

  • 神経芽腫に対する新規治療剤の探索
    榎本 秀樹
    国立研究開発法人日本医療研究開発機構, 創薬支援推進事業・創薬総合支援事業, 2017
    Competitive research funding

  • 嗅索道標細胞(lot細胞)の移動機構の解析
    伊藤 圭祐
    日本学術振興会, 科学研究費助成事業 特別研究員奨励費, 特別研究員奨励費, 2006 - 2007
    本研究の目的はlot細胞の移動を制御する分子を探索する事にある。昨年度の候補分子のスクリーニングによりFGF8とSDF1に誘引の傾向が、EphB1に反発の傾向がある事が分かった。そこで今年度はこれらの分子のlot細胞に対する効果の検証を行った。FGF8とSDFに関しては発現パターンや薬剤投与の結果から、lot細胞の移動に機能する可能性が低いと判断した。一方EphB1に関しては興味深い結果が多く得られた。培養実験、発現解析などの結果から、EphB1とephrinBとの間の双方向シグナルがlot細胞の移動に関与している事が示唆された。またlot細胞は外套・副外套境界と呼ばれる終脳の背腹を分ける境界に配列するため、今後この解析によりそれまで不明であった脳内における細胞移動と境界形成との間の関連も解明できる可能性がある。 昨年度から継続して行った研究に、Sema3Fのlot細胞に対する反発効果の研究があり、遺伝子破壊マウスの表現型解析からSema3Fが移動後のlot細胞を表層に留めるという新しい機能を明らかにした。この研究は論文にまとめ、受理された。昨年度からの継続研究のもう一つにK252aという薬剤の効果の解析がある。この薬剤を与えると移動神経細胞の細胞体の動きは止まる一方で先導突起は止まらないという結果が観察された。このためこれまで不明であった、神経細胞が細胞移動から軸索伸長へ切り替わるメカニズムの解明が期待された。実験を進めた結果、まずK252aはlot細胞以外の神経細胞にも同様の効果を持つ事が分かり、またCDKの阻害剤を与えるとK252aとよく似た効果が表れた。そこでCDK5に着目して機能欠損の実験を行った所、細胞体の移動は減少する一方で突起はそのまま伸長することが分かった。このため、CDK5の活性が神経細胞の移動と軸索身長を切り替えるスイッチとして働く可能性が示唆された

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