Updated on 2021/10/11

写真a

 
TAKEI Kohji
 
Organization
Medicine, Dentistry and Pharmaceutical Sciences Professor
Position
Professor
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Degree

  • 歯学博士 ( 東京歯科大学 )

Research Interests

  • 細胞内小胞輸送

  • 細胞生物

  • cytoskeleton

  • membrane traffic

  • cell biology

  • 細胞骨格

Research Areas

  • Life Science / Cell biology

Education

  • Tokyo Dental College    

    - 1989

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  • Tokyo Dental College   歯学研究科   病理学

    - 1989

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    Country: Japan

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  • Tokyo Dental College    

    - 1985

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  • Tokyo Dental College   College of Dentistry  

    - 1985

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    Country: Japan

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Research History

  • - 岡山大学医歯薬学総合研究科 教授

    2005

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  • - Professor,Graduate School of Medicine, Dentistry and Pharmaceutical Sciences,Okayama University

    2005

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  • 岡山大学医歯学総合研究科 未設定

    2001 - 2005

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  • Research Specialist II,Howard Hughes Medical Institute

    1995 - 1999

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  • ハワードヒューズ 医学財団 リサーチスペシャリスト

    1995 - 1999

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  • Associate Research Scientist,Yale University, School of Medicine

    1993 - 1999

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  • エール大学医学部 アソシエートリサーチサイエンティスト

    1993 - 1999

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  • ハワードヒューズ 医学財団 ポストドクトラルアソシエート

    1992 - 1994

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  • Postdoctoral Associate,Howard Hughes Medical Institute

    1992 - 1994

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  • postdoctoral fellow,Yale Unversity, School of Medicine

    1988 - 1993

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  • エール大学医学部 ポストドクトラルフェロー

    1988 - 1993

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Professional Memberships

 

Books

  • Use of liposomes to study vesicular transport

    Humana Press  2010 

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  • Methods in Molecular Biology “Liposomes”

    Humana Press (USA), Springer Publishing Group,  2009 

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  • 生体の科学55

    医学書院,東京  2004 

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  • 文部科学省科学研究費特定領域研究C「ゲノム」2000年度報告書

    2001 

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  • 平成12?13年度科学研究費補助金(基盤研究(B)(2))研究成果報告書

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MISC

  • Dynamin2 GTPase contributes to invadopodia formation in invasive bladder cancer cells

    Yubai Zhang, Maya Nolan, Hiroshi Yamada, Masami Watanabe, Yasutomo Nasu, Kohji Takei, Tetsuya Takeda

    BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS   480 ( 3 )   409 - 414   2016.11

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    Language:English   Publisher:ACADEMIC PRESS INC ELSEVIER SCIENCE  

    Cancer cell invasion is mediated by actin-based membrane protrusions termed invadopodia. Invadopodia consist of "core" F-actin bundles associated with adhesive and proteolytic machineries promoting cell invasion by degrading extracellular matrix (ECM). Formation of the F-actin core in invadopodia is regulated by various actin-binding proteins including Arp2/3 complex and cortactin. Dynamin GTPase localizes to the invadopodia and is implicated in cancer cell invasion, but its precise role at the invadopodia remained elusive.
    In this study, we examined the roles of dynamin at the invadopodia of bladder cancer cells. Although all three dynamin isoforms (dynamin1, 2 and 3) are expressed in human bladder cancer cell line T24, only dynamin2 localizes to the invadopodia. Inhibition of dynamin2 function, using either RNA interference (RNAi) or the dynamin specific inhibitor Dynasore, caused defects in invadopodia formation and suppressed invasive activity of 124 bladder cancer cells. Structure-function analysis using dynamin2 deletion fragments identified the proline/arginine-rich domain (PRD) of dynamin2 as indispensable for invadopodia formation and invasiveness of 124 cells. Thus, dynamin2 contributes to bladder cancer invasion by controlling invadopodia formation in bladder cancer cells and may prove a valuable therapeutic target. (C) 2016 Elsevier Inc. All rights reserved.

    DOI: 10.1016/j.bbrc.2016.10.063

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  • Actin bundling by dynamin 2 and cortactin is implicated in cell migration by stabilizing filopodia in human non-small cell lung carcinoma cells

    Hiroshi Yamada, Tetsuya Takeda, Hiroyuki Michiue, Tadashi Abe, Kohji Takei

    INTERNATIONAL JOURNAL OF ONCOLOGY   49 ( 3 )   877 - 886   2016.9

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    Language:English   Publisher:SPANDIDOS PUBL LTD  

    The endocytic protein dynamin participates in the formation of actin-based membrane protrusions such as podosomes, pseudopodia, and invadopodia, which facilitate cancer cell migration, invasion, and metastasis. However, the role of dynamin in the formation of actin-based membrane protrusions at the leading edge of cancer cells is unclear. In this study, we demonstrate that the ubiquitously expressed dynamin 2 isoform facilitates cell migration by stabilizing F-actin bundles in filopodia of the lung cancer cell line H1299. Pharmacological inhibition of dynamin 2 decreased cell migration and filopodial formation. Furthermore, dynamin 2 and cortactin mostly colocalized along F-actin bundles in filopodia of serum-stimulated H1299 cells by immunofluorescent and immunoelectron microscopy. Knockdown of dynamin 2 or cortactin inhibited the formation of filopodia in serum-stimulated H1299 cells, concomitant with a loss of F-actin bundles. Expression of wild-type cortactin rescued the punctate-like localization of dynamin 2 and filopodial formation. The incubation of dynamin 2 and cortactin with F-actin induced the formation of long and thick actin bundles, with these proteins colocalizing at F-actin bundles. A depolymerization assay revealed that dynamin 2 and cortactin increased the stability of F-actin bundles. These results indicate that dynamin 2 and cortactin participate in cell migration by stabilizing F-actin bundles in filopodia. Taken together, these findings suggest that dynamin might be a possible molecular target for anticancer therapy.

    DOI: 10.3892/ijo.2016.3592

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  • Expression of a dynamin 2 mutant associated with Charcot-Marie-Tooth disease leads to aberrant actin dynamics and lamellipodia formation

    Hiroshi Yamada, Kinue Kobayashi, Yubai Zhang, Tetsuya Takeda, Kohji Takei

    NEUROSCIENCE LETTERS   628   179 - 185   2016.8

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    Language:English   Publisher:ELSEVIER IRELAND LTD  

    Specific mutations in dynamin 2 are linked to Charcot-Marie-Tooth disease (CMT), an inherited peripheral neuropathy. However, the effects of these mutations on dynamin function, particularly in relation to the regulation of the actin cytoskeleton remain unclear. Here, selected CMT-associated dynamin mutants were expressed to examine their role in the pathogenesis of CMT in U2OS cells. Ectopic expression of the dynamin CMT mutants 555 Delta 3 and K562E caused an approximately 50% decrease in serum stimulation dependent lamellipodia formation; however, only K562E caused aberrations in the actin cytoskeleton. Immunofluorescence analysis showed that the K562E mutation resulted in the disappearance of radially aligned actin bundles and the simultaneous appearance of F-actin clusters. Live-cell imaging analyses showed F-actin polymers of decreased length assembled into immobile clusters in K562E-expressing cells. The K562E dynamin mutant colocalized with the F-actin clusters, whereas its colocalization with clathrin-coated pit marker proteins was decreased. Essentially the same results were obtained using another cell line, HeLa and NG108-15 cells. The present study is the first to show the association of dynamin CMT mutations with aberrant actin dynamics and lamellipodia, which may contribute to defective endocytosis and myelination in Schwann cells in CMT. (C) 2016 The Authors. Published by Elsevier Ireland Ltd.

    DOI: 10.1016/j.neulet.2016.06.030

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  • Antiobesity Action of ACAM by Modulating the Dynamics of Cell Adhesion and Actin Polymerization in Adipocytes

    Kazutoshi Murakami, Jun Eguchi, Kazuyuki Hida, Atsuko Nakatsuka, Akihiro Katayama, Miwa Sakurai, Haruki Choshi, Masumi Furutani, Daisuke Ogawa, Kohji Takei, Fumio Otsuka, Jun Wada

    DIABETES   65 ( 5 )   1255 - 1267   2016.5

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    Language:English   Publisher:AMER DIABETES ASSOC  

    Coxsackie virus and adenovirus receptor-like membrane protein (CLMP) was identified as the tight junction-associated transmembrane protein of epithelial cells with homophilic binding activities. CLMP is also recognized as adipocyte adhesion molecule (ACAM), and it is upregulated in mature adipocytes in rodents and humans with obesity. Here, we present that aP2 promoter-driven ACAM transgenic mice are protected from obesity and diabetes with the prominent reduction of adipose tissue mass and smaller size of adipocytes. ACAM is abundantly expressed on plasma membrane of mature adipocytes and associated with formation of phalloidin-positive polymerized form of cortical actin (F-actin). By electron microscopy, the structure of zonula adherens with an intercellular space of approximate to 10-20 nm was observed with strict parallelism of the adjoining cell membranes over distances of 1-20 m, where ACAM and -actin are abundantly expressed. The formation of zonula adherens may increase the mechanical strength, inhibit the adipocyte hypertrophy, and improve the insulin sensitivity.

    DOI: 10.2337/db15-1304

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  • Fluvoxamine, an anti-depressant, inhibits human glioblastoma invasion by disrupting actin polymerization

    Keiichiro Hayashi, Hiroyuki Michiue, Hiroshi Yamada, Katsuyoshi Takata, Hiroki Nakayama, Fan-Yan Wei, Atsushi Fujimura, Hiroshi Tazawa, Akira Asai, Naohisa Ogo, Hiroyuki Miyachi, Tei-ichi Nishiki, Kazuhito Tomizawa, Kohji Takei, Hideki Matsui

    SCIENTIFIC REPORTS   6   2016.3

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    Language:English   Publisher:NATURE PUBLISHING GROUP  

    Glioblastoma multiforme (GBM) is the most common malignant brain tumor with a median survival time about one year. Invasion of GBM cells into normal brain is the major cause of poor prognosis and requires dynamic reorganization of the actin cytoskeleton, which includes lamellipodial protrusions, focal adhesions, and stress fibers at the leading edge of GBM. Therefore, we hypothesized that inhibitors of actin polymerization can suppress GBM migration and invasion. First, we adopted a drug repositioning system for screening with a pyrene-actin-based actin polymerization assay and identified fluvoxamine, a clinically used antidepressant. Fluvoxamine, selective serotonin reuptake inhibitor, was a potent inhibitor of actin polymerization and confirmed as drug penetration through the blood-brain barrier (BBB) and accumulation of whole brain including brain tumor with no drug toxicity. Fluvoxamine inhibited serum-induced ruffle formation, cell migration, and invasion of human GBM and glioma stem cells in vitro by suppressing both FAK and Akt/mammalian target of rapamycin signaling. Daily treatment of athymic mice bearing human glioma-initiating cells with fluvoxamine blocked tumor cell invasion and prolonged the survival with almost same dose of anti-depressant effect. In conclusion, fluvoxamine is a promising anti-invasive treatment against GBM with reliable approach.

    DOI: 10.1038/srep23372

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  • PtdIns4KII alpha generates endosomal PtdIns(4)P and is required for receptor sorting at early endosomes

    Yuji Henmi, Yoshiaki Morikawa, Natsuko Oe, Narumi Ikeda, Akikazu Fujita, Kohji Takei, Shane Minogue, Kenji Tanabe

    MOLECULAR BIOLOGY OF THE CELL   27 ( 6 )   990 - 1001   2016.3

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    Language:English   Publisher:AMER SOC CELL BIOLOGY  

    Phosphatidylinositol 4-kinase II alpha (PtdIns4KII alpha) localizes to the trans-Golgi network and endosomal compartments and has been implicated in the regulation of endosomal traffic, but the roles of both its enzymatic activity and the site of its action have not been elucidated. This study shows that PtdIns4KII alpha is required for production of endosomal phosphatidylinositol 4-phosphate (PtdIns(4)P) on early endosomes and for the sorting of transferrin and epidermal growth factor receptor into recycling and degradative pathways. Depletion of PtdIns4KII alpha with small interfering RNA significantly reduced the amount of vesicular PtdIns(4) P on early endosomes but not on Golgi membranes. Cells depleted of PtdIns4KII alpha had an impaired ability to sort molecules destined for recycling from early endosomes. We further identify the Eps15 homology domain-containing protein 3 (EHD3) as a possible endosomal effector of PtdIns4KII alpha. Tubular endosomes containing EHD3 were shortened and became more vesicular in PtdIns4KII alpha-depleted cells. Endosomal PtdIns(4,5)P-2 was also significantly reduced in PtdIns4KII alpha-depleted cells. These results show that PtdIns4KII alpha regulates receptor sorting at early endosomes through a PtdIns(4) P-dependent pathway and contributes substrate for the synthesis of endosomal PtdIns(4,5)P-2.

    DOI: 10.1091/mbc.E15-08-0564

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  • Possible role of cortactin phosphorylation by protein kinase C in actin-bundle formation at growth cone

    Hiroshi Yamada, Tatsuya Kikuchi, Toshio Masumoto, Fan-Yan Wei, Tadashi Abe, Tetsuya Takeda, Teiichi Nishiki, Kazuhito Tomizawa, Masami Watanabe, Hideki Matsui, Kohji Takei

    BIOLOGY OF THE CELL   107 ( 9 )   319 - 330   2015.9

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    Language:English   Publisher:WILEY-BLACKWELL  

    Background Information. Cortactin contributes to growth cone morphogenesis by forming with dynamin, ring-shaped complexes that mechanically bundle and stabilise F-actin. However, the regulatory mechanism of cortactin action is poorly understood.
    Results. Immunofluorescence microscopy revealed that protein kinase C (PKC) colocalises with cortactin at growth cone filopodia in SH-SY5Y neuroblastoma cells. PKC activation by phorbol 12-myristate 13-acetate causes cortactin phosphorylation, filopodial retraction and F-actin-bundle loss. Moreover, PKC directly phosphorylates cortactin in vitro at S135/T145/S172, mitigating both cortactin's actin-binding and actin-crosslinking activity, whereas cellular expression of a phosphorylation-mimetic cortactin mutant hinders filopodial formation with a significant decrease of actin bundles.
    Conclusions. Our results indicate that PKC-mediated cortactin phosphorylation might be implicated in the maintenance of growth cone.

    DOI: 10.1111/boc.201500032

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  • Stabilization of actin bundles by a dynamin 1/cortactin ring complex is necessary for growth cone filopodia.

    Yamada, H, Abe, T, Satoh, A, Okazaki, N, Tago, S, Kobayashi, K, Yoshida, Y, Oda, Y, Watanabe, M, Tomizawa, K, Matsui, H, Takei, K

    Journal of Neuroscience   33 ( 10 )   4514 - 4526   2015

  • ダイナミン2による微小管の動態制御とCharcot-Marie-Tooth病

    岡山医学会雑誌   122 ( 2 )   113 - 117   2010

  • Expression pattern of REIC/Dkk-3 in various cell types and the implications of the soluble form in prostatic acinar development

    Int. J. Oncol.   122   113 - 117   2010

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  • Dynamic instability of microtubules requires dynamin 2 and is impaired in a Charcot-Marie-Tooth mutant

    Kenji Tanabe, Kohji Takei

    JOURNAL OF CELL BIOLOGY   185 ( 6 )   939 - 948   2009.6

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    Language:English   Publisher:ROCKEFELLER UNIV PRESS  

    Dynamin is a fission protein that participates in endocytic vesicle formation. Although dynamin was originally identified as a microtubule-binding protein, the physiological relevance of this function was unclear. Recently, mutations in the ubiquitously expressed dynamin 2 (dyn2) protein were found in patients with Charcot-Marie-Tooth (CMT) disease, which is an inherited peripheral neuropathy. In this study, we show that one of these mutations, 551 Delta 3, induces prominent decoration of microtubules with the mutant dyn2. Dyn2 was required for proper dynamic instability of microtubules, and this was impaired in cells expressing the 551 Delta 3 mutant, which showed a remarkable increase in microtubule acetylation, a marker of stable microtubules. Depletion of endogenous dyn2 with a small interfering RNA also resulted in the accumulation of stable microtubules. Furthermore, the formation of mature Golgi complexes, which depends on microtubule-dependent membrane transport, was impaired in both dyn2 knockdown cells and cells expressing the 551 Delta 3 mutant. Collectively, our results suggest that dyn2 regulates dynamic instability of microtubules, which is essential for organelle motility, and that this function may be impaired in CMT disease.

    DOI: 10.1083/jcb.200803153

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  • アンフィファイジンとN-WASPのダイナミックな相互作用がアクチン重合を制御する(Dynamic interation of amphiphysin with N-WASP regulates actin assembly)

    山田浩司, セルジ=パディラパーラ, スンジュー=パク, 伊藤俊樹, 名省略, 、ファビオ=ベンフェナティ, ピエトロ=デカメリ, マイテ=コペイモイソン, マルコ=トラミア, テリー=ガリ, 竹居孝二

    ジャーナル・オブ・バイロロジカル・ケミストリー(J. Biol. Chem.)   2009

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  • Dynamin 2 Cooperates with Amphiphysin 1 in Phagocytosis in Sertoli Cells

    Akira Nakanishi, Tadashi Abe, Masami Watanabe, Kohji Takei, Hiroshi Yamada

    ACTA MEDICA OKAYAMA   62 ( 6 )   385 - 391   2008.12

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    Language:English   Publisher:OKAYAMA UNIV MED SCHOOL  

    Testicular Sertoli cells highly express dynamin 2 and amphiphysin 1. Here we demonstrate that dynamin 2 is implicated in phosphatidylserine (PS)-dependent phagocytosis in Sertoli cells. Immunofluorescence and dual-live imaging revealed that dynamin 2 and amphiphysin 1 accumulate simultaneously at ruffles. These proteins are specifically bound in vitro. Over-expression of dominant negative dynamin 2 (K44A) inhibits liposome-uptake and leads to the mis-localization of amphiphysin 1. Thus, the cooperative function of dynamin 2 and amphiphysin 1 in PS-dependent phagocytosis is strongly suggested.

    DOI: 10.18926/AMO/30954

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  • CaM kinase l alpha-induced phosphorylation of Drp1 regulates mitochondrial morphology

    Xiao-Jian Han, Yun-Fei Lu, Shun-Ai Li, Taku Kaitsuka, Yasufumi Sato, Kazuhito Tomizawa, Angus C. Nairn, Kohji Takei, Hideki Matsui, Masayuki Matsushita

    JOURNAL OF CELL BIOLOGY   182 ( 3 )   573 - 585   2008.8

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    Language:English   Publisher:ROCKEFELLER UNIV PRESS  

    Mitochondria are dynamic organelles that frequently move, divide, and fuse with one another to maintain their architecture and functions. However, the signaling mechanisms involved in these processes are still not well characterized. In this study, we analyze mitochondrial dynamics and morphology in neurons. Using time-lapse imaging, we find that Ca(2+) influx through voltage-dependent Ca(2+) channels (VDCCs) causes a rapid halt in mitochondrial movement and induces mitochondrial fission. VDCC-associated Ca(2+) signaling stimulates phosphorylation of dynamin-related protein 1 (Drp1) at serine 600 via activation of Ca(2+)/calmodulin-dependent protein kinase l alpha ( CaMKl alpha). In neurons and HeLa cells, phosphorylation of Drp1 at serine 600 is associated with an increase in Drp1 translocation to mitochondria, whereas in vitro, phosphorylation of Drp1 results in an increase in its affinity for Fis1. CaMKl alpha is a widely expressed protein kinase, suggesting that Ca(2+) is likely to be functionally important in the control of mitochondrial dynamics through regulation of Drp1 phosphorylation in neurons and other cell types.

    DOI: 10.1083/jcb.200802164

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  • Regulation of clathrin-mediated endocytosis by p53

    Yoshie Endo, Atsumi Sugiyama, Shun-Ai Li, Kazuji Ohmori, Hirokazu Ohata, Yusuke Yoshida, Masabumi Shibuya, Kohji Takei, Masato Enari, Yoichi Taya

    GENES TO CELLS   13 ( 4 )   375 - 385   2008.4

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    Language:English   Publisher:BLACKWELL PUBLISHING  

    The p53 gene encodes a multi-functional protein to prevent tumorigenesis. Although there have been many reports of the nuclear functions of p53, little is known about the cytosolic functions of p53. Here, we found that p53 is present in cytosol as well as nuclei under unstressed conditions and binds to clathrin heavy chain (CHC). CHC is known to play a role in receptor-mediated endocytosis. Based on our findings, we examined the effect of p53 on clathrin-mediated endocytosis of epidermal growth factor receptor (EGFR). Surprisingly, p53 co-localized with CHC at the plasma membrane in response to EGF stimulation. In cells with ablated p53 expression by RNAi, EGFR internalization was delayed and intracellular signaling from EGFR was altered. Thus, our findings provide evidence that cytosolic p53 may participate in the regulation of clathrin-mediated endocytosis to control the correct signaling from EGFR.

    DOI: 10.1111/j.1365-2443.2008.01172.x

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  • SNX9 regulates tubular invagination of the plasma membrane through interaction with actin cytoskeleton and dynamin 2

    Narae Shin, Namhui Ahn, Belle Chang-Ileto, Joohyun Park, Kohji Takei, Sang-Gun Ahn, Soo-A Kim, Gilbert Di Paolo, Sunghoe Chang

    JOURNAL OF CELL SCIENCE   121 ( 8 )   1252 - 1263   2008.4

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    Language:English   Publisher:COMPANY OF BIOLOGISTS LTD  

    Dynamic membrane remodeling during intracellular trafficking is controlled by the intricate interplay between lipids and proteins. BAR domains are modules that participate in endocytic processes by binding and deforming the lipid bilayer. Sorting nexin 9 (SNX9), which functions in clathrin-mediated endocytosis, contains a BAR domain, however, the properties of this domain are not well understood. Here we show that SNX9 shares many properties with other BAR domain-containing proteins, such as amphiphysin and endophilin. SNX9 is able to deform the plasma membrane, as well as liposomes, into narrow tubules and recruit N-WASP and dynamin 2 to these tubules via its SH3 domain. SNX9-induced tubulation is antagonized by N-WASP and dynamin 2 while it is enhanced by perturbation of actin dynamics. However, SNX9 also has several unique properties. The tubulating activity requires the BAR and PX domains, as well as the low-complexity (LC) domain, which binds the Arp2/3 complex. SNX9 also binds to PtdIns(4)P-5-kinases via its PX domain and its tubulating activity is regulated by phosphoinositides. In addition, the kinase activity of PtdIns(4)P-5-kinases is stimulated by interaction with SNX9, suggesting a positive feedback interaction between SNX9 and PtdIns(4)P-5-kinases. These results suggest that SNX9 functions in the coordination of membrane remodeling and fission via interactions with actin-regulating proteins, endocytic proteins and PtdIns(4,5)P-2-metabolizing enzymes.

    DOI: 10.1242/jcs.016709

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  • Involvement of calcineurin in glutamate-induced mitochondrial dynamics in neurons

    Xiao-Jian Han, Yun-Fei Lu, Shun-Ai Li, Kazuhito Tomizawa, Kohji Takei, Masayuki Matsushita, Hideki Matsui

    NEUROSCIENCE RESEARCH   60 ( 1 )   114 - 119   2008.1

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    Language:English   Publisher:ELSEVIER IRELAND LTD  

    Alterations in the morphology and movement of mitochondria influence neuronal viability. However, the precise mechanisms of such alterations are unclear. In this study, we showed calcineurin was involved in the regulation of mitochondrial dynamics. Glutamate stimulation inhibited mitochondrial movement and decreased mitochondrial length in neurons. FK506 and cyclosporine A, calcineurin inhibitors, attenuated the effects of glutamate on mitochondrial dynamics. It was also found that glutamate treatment dephosphorylated, a proapoptotic protein, Bad and promoted its translocation to mitochondria in neurons via calcineurin. These results provide important new insights into intracellular signaling pathways that regulate mitochondrial dynamics and neuronal cell death. (c) 2007 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.

    DOI: 10.1016/j.neures.2007.09.012

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  • Amphiphysin 1 is important for actin polymerization During phagocytosis

    Hiroshi Yamada, Emiko Ohashi, Tadashi Abe, Norihiro Kusumi, Shun-AI Li, Yumi Yoshida, Masami Watanabe, Kazuhito Tomizawa, Yuji Kashiwakura, Hiromi Kumon, Hideki Matsui, Kohji Takei

    MOLECULAR BIOLOGY OF THE CELL   18 ( 11 )   4669 - 4680   2007.11

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    Language:English   Publisher:AMER SOC CELL BIOLOGY  

    Amphiphysin 1 is involved in clathrin-mediated endocytosis. In this study, we demonstrate that amphiphysin 1 is essential for cellular phagocytosis and that it is critical for actin polymerization. Phagocytosis in Sertoli cells was induced by stimulating phosphatidylserine receptors. This stimulation led to the formation of actin-rich structures, including ruffles, phagocytic cups, and phagosomes, all of which showed an accumulation of amphiphysin 1. Knocking out amphiphysin 1 by RNA interference in the cells resulted in the reduction of ruffle formation, actin polymerization, and phagocytosis. Phagocytosis was also drastically decreased in amph 1(-/-) Sertoli cells. In addition, phosphatidylinositol-4,5-bisphosphate-induced actin polymerization was decreased in the knockout testis cytosol. The addition of recombinant amphiphysin 1 to the cytosol restored the polymerization process. Ruffle formation in small interfering RNA-treated cells was recovered by the expression of constitutively active Rac1, suggesting that amphiphysin 1 functions upstream of the protein. These findings support that amphiphysin 1 is important in the regulation of actin dynamics and that it is required for phagocytosis.

    DOI: 10.1091/mbc.E07-04-0296

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  • cAMP-response element binding protein (CREB) regulates cyclosporine-A-mediated down-regulation of cathepsin B and L synthesis

    Kazuhiro Omori, Koji Naruishi, Tomoko Yamaguchi, Shun-Ai Li, Mayumi Yamaguchi-Morimoto, Kaori Matsuura, Hideo Arai, Kohji Takei, Shogo Takashiba

    CELL AND TISSUE RESEARCH   330 ( 1 )   75 - 82   2007.10

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    Language:English   Publisher:SPRINGER  

    Cyclosporin A (CsA) is an immunosuppressant with severe side effects including gingival overgrowth. We have previously reported that CsA impairs the activity of the lysosomal enzymes cathepsin B and L in human gingival fibroblasts (HGFs). Here, we have examined the effects of CsA on the DNA-binding activity of the cyclic AMP response element-binding protein (CREB) and cell viability, and the effects of CREB on cathepsin B and L synthesis and activity in HGFs. We have confirmed that CsA down-regulates cathepsin B and L synthesis. Further, CsA has no effect on cell viability and dramatically impairs CREB-DNA binding activity. Importantly, the synthesis of cathepsin B and L is down-regulated, and their activity is also significantly impaired in HGFs transfected with plasmid expressing dominant-negative CREB. These results suggest that CREB is essential for the CsA-mediated down-regulation of cathepsin B and L synthesis in HGFs.

    DOI: 10.1007/s00441-007-0457-8

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  • Truncations of amphiphysin I by calpain inhibit vesicle endocytosis during neural hyperexcitation

    Yumei Wu, Shuang Liang, Yoshiya Oda, Iori Ohmori, Tei-Ichi Nishiki, Kohji Takei, Hideki Matsui, Kazuhito Tomizawa

    EMBO JOURNAL   26 ( 12 )   2981 - 2990   2007.6

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    Language:English   Publisher:NATURE PUBLISHING GROUP  

    Under normal physiological conditions, synaptic vesicle endocytosis is regulated by phosphorylation and Ca2+-dependent dephosphorylation of endocytic proteins such as amphiphysin and dynamin. To investigate the regulatory mechanisms that may occur under the conditions of excessive presynaptic Ca2+ influx observed preceding neural hyperexcitation, we examined hippocampal slices following high-potassium or high-frequency electrical stimulation (HFS). In both cases, three truncated forms of amphiphysin I resulted from cleavage by the protease calpain. In vitro, the binding of truncated amphiphysin I to dynamin I and copolymerization into rings with dynamin I were inhibited, but its interaction with liposomes was not affected. Moreover, overexpression of the truncated form of amphiphysin I inhibited endocytosis of transferrin and synaptic vesicles. Inhibiting calpain prevented HFS-induced depression of presynaptic transmission. Finally, calpain-dependent amphiphysin I cleavage attenuated kainate-induced seizures. These results suggest that calpain-dependent cleavage of amphiphysin I inhibits synaptic vesicle endocytosis during neural hyperexcitation and demonstrate a novel post-translational regulation of endocytosis.

    DOI: 10.1038/sj.emboj.7601741

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  • Implication of amphiphysin 1 and dynamin 2 in tubulobulbar complex formation and spermatid release.

    Kusumi N, Watanabe M, Yamada H, Li SA, Kashiwakura Y, Matsukawa T, Nagai A, Nasu Y, Kumon H, Takei K

    Cell Struc Func   2 ( 2 )   101 - 113   2007

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  • Regulatory mechanisms of dynamin-dependent endocytosis

    K Takei, Y Yoshida, H Yamada

    JOURNAL OF BIOCHEMISTRY   137 ( 3 )   243 - 247   2005.3

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    Language:English   Publishing type:Book review, literature introduction, etc.   Publisher:JAPANESE BIOCHEMICAL SOC  

    Extensive studies on endocytosis in the last decade have resulted in identification of several key molecules that function in clathrin- and dynamin-dependent endocytosis. Most endocytic molecules contain multiple binding motifs that mediate protein-protein or protein-lipid interactions, which must be modulated spatially and temporally during endocytosis. Regulation of these interactions is the molecular basis of regulatory mechanisms involved in endocytosis. This review first describes current models of the mechanism of dynamin-dependent fission, then introduces several mechanisms that modulate dynamin GTPase activity and dynamin-dependent vesicle formation. Such mechanisms include regulation by inositol phospholipids, especially phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P-2], and their metabolism. It concludes by describing the regulation of dynamin 1 by its binding partner, amphiphysin 1, and regulation by cyclin-dependent kinase 5 (Cdk5)-dependent phosphorylation of dynamin 1 and amphiphysin 1. These mechanisms help endocytic molecules to function properly, and cooperatively regulate dynamin-dependent endocytosis.

    DOI: 10.1093/jb/mvi052

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  • A human beta-cell line for transplantation therapy to control type 1 diabetes.

    Narushima M, Kobayashi N, Okitsu T, Tanaka Y, Li SA, Chen Y, Miki A, Tanaka K, Nakaji S, Takei K, Gutierrez AS, Rivas-Carrillo JD, Navarro-Alvarez N, Jun HS, Westerman KA, Noguchi H

    Nature Biotech   2005

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  • Dynamin-2 regulates oxidized low-density lipoprotein-induced apoptosis of vascular smooth muscle cell

    Y Kashiwakura, M Watanabe, N Kusumi, K Sumiyoshi, Y Nasu, H Yamada, T Sawamura, H Kumon, K Takei, H Daida

    CIRCULATION   110 ( 21 )   3329 - 3334   2004.11

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    Language:English   Publisher:LIPPINCOTT WILLIAMS & WILKINS  

    Background-On exposure to oxidized low-density lipoprotein (oxLDL), vascular cells generally undergo apoptosis, which is one of the major pathogenic factors of atherosclerosis. In this study, we examined the role of dynamin ( a crucial GTPase protein in endocytosis) in oxLDL-induced apoptosis of vascular smooth muscle cells (VSMC).
    Methods and Results-After oxLDL stimulation, dynamin-2 colocalized with LOX-1 around the cell surface, as well as oxLDL in the cytoplasm, suggesting that dynamin-2 was involved in scavenger receptor-mediated oxLDL endocytosis. Downregulation of dynamin-2 induced by dynamin-2 dominant negative plasmid (K44A) resulted in a decrease of oxLDL uptake and thereby in a reduction of apoptosis. These data demonstrated that dynamin-2 was involved in oxLDL-induced apoptosis via the oxLDL endocytotic pathway. On the other hand, dynamin-2 wild-type plasmid transfection promoted oxLDL-induced apoptosis without increasing oxLDL uptake. Interestingly, the p53 inhibitor pifithrin-alpha (PFT) significantly reduced apoptosis promoted by wild-type dynamin-2 (78% reduction compared with the PFT[-] condition). These results indicated that dynamin-2 enhanced oxLDL-induced apoptosis of VSMC by participating in the p53 pathway, probably as a signal transducer. Moreover, we demonstrated that, in advanced plaques of apolipoprotein E-/- mice, dynamin-2 expression was often enhanced in apoptotic VSMC, suggesting that dynamin-2 might participate in apoptosis of VSMC even in vivo.
    Conclusions - Our data demonstrated that dynamin-2 at least partially regulated oxLDL-induced apoptosis of VSMC by participating in 2 independent pathways: the oxLDL endocytotic pathway and the p53 pathway. These findings suggest that dynamin-2 may serve as a new research or therapeutic target in vascular disease.

    DOI: 10.1161/01.CIR.0000147828.86593.85

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  • The stimulatory action of amphiphysin on dynamin function is dependent on lipid bilayer curvature

    Y Yoshida, M Kinuta, T Abe, S Liang, K Araki, O Cremona, G Di Paolo, Y Moriyama, T Yasuda, P De Camilli, K Takei

    EMBO JOURNAL   23 ( 17 )   3483 - 3491   2004.9

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    Language:English   Publisher:NATURE PUBLISHING GROUP  

    Amphiphysin is a major dynamin-binding partner at the synapse; however, its function in fission is unclear. Incubation of large unilamellar liposomes with mice brain cytosol led to massive formation of small vesicles, whereas cytosol of amphiphysin 1 knockout mice was much less efficient in this reaction. Vesicle formation from large liposomes by purified dynamin was also strongly enhanced by amphiphysin. In the presence of liposomes, amphiphysin strongly affected dynamin GTPase activity and the recruitment of dynamin to the liposomes, but this activity was highly dependent on liposome size. Deletion from amphiphysin of its central proline-rich stretch dramatically potentiated its effect on dynamin, possibly by relieving an inhibitory intramolecular interaction. These results suggest a model in which maturation of endocytic pits correlates with the oligomerization of dynamin with either amphiphysin or other proteins with similar domain structure. Formation of these complexes is coupled to the activation of dynamin GTPase activity, thus explaining how deep invagination of the pit leads to fission.

    DOI: 10.1038/sj.emboj.7600355

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  • Immunohistochemical localization of Klotho protein in brain, kidney, and reproductive organs of mice.

    Li S-A, Watanabe M, Yamada H, Nagai A, Kinuta M, Takei K

    Cell Struct Funct   2004

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  • ダイナミンとエンドサイトーシス。

    竹居孝二

    蛋白質核酸酵素 増刊『細胞における蛋白質の一生』   49,938-941   2004

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  • Formation of S-[2-carboxy-1-(1H-imidazol-4-yl) ethyl]glutathione, a new metabolite of L-histidine, from cis-urocanic acid and glutathione by the action of glutathione S-transferase

    M Kinuta, K Kinuta, H Yamada, T Abe, Y Yoshida, K Araki, SA Li, A Otsuka, A Nakanishi, Y Moriyama, K Takei

    ELECTROPHORESIS   24 ( 18 )   3212 - 3218   2003.9

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    Language:English   Publisher:WILEY-V C H VERLAG GMBH  

    Exposure of the skin to sunlight results in an increase of the content of epidermal trans-urocanic acid, a key metabolite Of L-histidine, and also in occurrence of the isomerization of trans-urocanic acid to the cis isomer. S-[2-Carboxy-1-(1H-imidazol-4-yl)ethyl]glutathione (GS(CIE)), An adduct of urocanic acid and glutathione, is a presumed origin of a urinary compound S-[2-carboxy-1-(1H-imidazol-4-yl)ethyl]-L-cysteine (Cys(CIE)). The formation of GS(CIE) is stimulated by exposing the skin to sunlight irradiation. In this study we investigated an enzymatic formation of GS(CIE) from glutathione and cis-urocanic acid by incubation with rat liver extract that contained glutathione S-transferase (GST) at high activity. The formation of GS(CIE) was suppressed significantly when a liver extract depleted of GST activity was used. Enzymatic degradation of GS(CIE) with gamma-glutamyl transpeptidase resulted in the formation of N-{S-[2-carboxy-1-(1H-imidazol-4-yl)ethyl]-L-cysteinyl}glycine, a metabolic intermediate between the glutathione adduct and Cys(CIE). A hydrolyzed product of GS(CIE) by HCl was identical with the urinary Cys(CIE). Compounds were analyzed by high-voltage paper electrophoresis, capillary electrophoresis, and fast atom bombardment mass spectrometry. From these results, we suggest that GS(CIE) formed from cis-urocanic acid and glutathione is an origin of the urinary compound Cys(CIE) and that the formation reaction is catalyzed mostly by the action of GST.

    DOI: 10.1002/elps.200305582

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  • ARF6 stimulates clathrin/AP-2 recruitment to synaptic membranes by activating phosphatidylinositol phosphate kinase type I gamma

    M Krauss, M Kinuta, MR Wenk, P De Camilli, K Takei, Haucke, V

    JOURNAL OF CELL BIOLOGY   162 ( 1 )   113 - 124   2003.7

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    Language:English   Publisher:ROCKEFELLER UNIV PRESS  

    Clathrin-mediated endocytosis of synaptic vesicle membranes involves the recruitment of clathrin and AP-2 adaptor complexes to the presynaptic plasma membrane. Phosphoinositides have been implicated in nucleating coat assembly by directly binding to several endocytotic proteins including AP-2 and AP180. Here, we show that the stimulatory effect of ATP and GTPgammaS on clathrin coat recruitment is mediated at least in part by increased levels Of PIP2. We also provide evidence for a role of ADP-ribosylation factor 6 (ARF6) via direct stimulation of a synaptically enriched phosphatidylinositol 4-phosphate 5-kinase type Igamma (PIPKIgamma), in this effect. These data suggest a model according to which activation of PIPKIgamma by ARF6-GTP facilitates clathrin-coated pit assembly at the synapse.

    DOI: 10.1083/jcb.200301006

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  • Formation of meso, N-diphenylprotoporphyrin IX by an aerobic reaction of phenylhydrazine with oxyhemoglobins.

    Nakanishi A, Kinuta K, Abe T, Araki K, Yoshida Y, Liang S, Li SA, Takei K, Kinuta M

    Acta Med Okayama   2003

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  • Cophosphorylation of amphiphysin I and dynamin I by Cdk5 regulates clathrin-mediated endocytosis of synaptic vesicles.

    Tomizawa K, Sunada S, Lu Y-F, Oda Y, Kinuta M, Ohshima T, Saito T, Wei F-Y, Matsushita M, Li S-Ti, Tsutsui K, Hisanaga S, Mikoshiba K, Takei K, Matsui H

    J. Cell Biol.   2003

  • イノシトールリン脂質によるエンドサイトーシスの制御機構

    絹田正裕, 竹居孝二

    生化学   75,1449-1452   2003

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  • Phosphatidylinositol 4,5-bisphosphate stimulates vesicle formation from liposomes by brain cytosol

    M Kinuta, H Yamada, T Abe, M Watanabe, SA Li, A Kamitani, T Yasuda, T Matsukawa, H Kumon, K Takei

    PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA   99 ( 5 )   2842 - 2847   2002.3

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    Language:English   Publisher:NATL ACAD SCIENCES  

    As a step toward the elucidation of mechanisms in vesicle budding, a cell-free assay that measures cytosol-induced vesicle generation from liposomes was established. This assay then was used to explore the role of phosphoinositides in vesicle formation. Liposomes incubated with brain cytosol in the presence of ATP and GTP massively generated small vesicles, as assessed both quantitatively and qualitatively by a dynamic light-scattering assay. Both ATP and GTP were required. Vesicle formation was inhibited greatly by the immunodepletion of dynamin 1 from the cytosol, indicating a major contribution of this GTPase in this reaction and suggesting that it mimics endocytic vesicle fission. Increasing the concentration Of L-alpha-phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P-2] but not of L-alpha-phosphatidylinositol 4-monophosphate or L-alpha-phosphatidylinositol in the lipid membranes enhanced vesicle formation. Lipid analysis revealed rapid degradation of PtdIns(4,5)P-2 to L-alpha-phosphatidylinositol during the incubation with the reaction reaching a maximum within 5 sec, whereas vesicle formation proceeded with a longer time course. PtdIns(4,5)P-2 degradation was independent of vesicle formation and occurred also in the presence of guanosine 5'-O-(thiotriphosphate), where few vesicle formations occurred. These results suggest that PtdIns(4,5)P-2 plays a critical role in the early step of vesicle formation, possibly in the recruitment of coats and fission factors to membranes.

    DOI: 10.1073/pnas.261715599

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  • Norrpinephrine-evoked 5-hydroxytryptamine secretion is Ca2+ dependent and mediated through exocytosis in rat pinealocytes.

    Yamada H, Hayashi M, Kinoshita M, Uehara S, Watanabe M, Takei K, Moriyama Y

    J Neurochem   2002

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  • Cdk5/P35 regulates neurotransmitter release through phosphorylation and downregulation of P/Q-type voltage-dependent calcium channel activity.

    Tomizawa K, Ohta J, Matsushita M, Moriwaki A, Li S-T, Takei K, Matsui H

    J Neurosci   2002

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  • Distribution of dynamins in testis, and their possible relation to spermatogenesis.

    Kamitani A, Yamada H, Kinuta M, Watanabe M, Li S-I, Matsukawa T, McNiven M, Kumon H, Takei K

    Biochem Biophys Res Commun   2002

  • Utilization of liposomes in vesicular transport studies.

    Kinuta M, Takei K

    Cell Struct Funct   27,2,63-69   2002

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  • Clathrin-mediated endocytosis: membrane factors pull the trigger.

    Takei K, Haucke V

    Trends in Cell Biol   11,385-391   2001

  • Determination of S-[2-carboxy-1-(1H-imidazol-4-yl)ethyl] glutathione, a novel metabolite of L-histidine, in tissue extracts from sunlight-irradiated rat by capillary electrophoresis.

    Kinuta M, Ohta J, Yamada H, Kinuta K, Abe T, Li SA, Otsuka A, Nakanishi A, Takei K

    Electrophoresis   2001

  • Interactions of dynamin and amphiphysin with liposomes.

    Takei K, Slepnev VI, De Camilli P

    Methods in Enzymol   2001

  • Generation of high curvature membranes mediated by direct endophilin-bilayer interactions.

    Farsad K, Ringstad N, Takei K, Floyd S R, Rose K, De Camilli P

    J Cell Biol   2001

  • Characterization of synaptojanin 2B isoforms expressed in brain and tests.

    Nemoto Y, Wenk M R, Watanabe M, Daniel L, Murakami T, Nemoto T, Ringstad N, Yamada H, Takei K, De Camilli P

    J Biol Chem   2001

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Presentations

  • Dynamin-Cortactin Ring Complex Physically Bundles Actin Filaments and Protects the Bundle from Depolarization

    IGER国際シンポジウム「Now in actin study: Motor protein research reaching a new stage」  2016 

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  • Dynamin-Cortactin Ring Complex Physically Bundles Actin Filaments and Protects the Bundle from Depolarization

    IGER International Symposium on Now in actin study: Motor protein research reaching a new stage]  2016 

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  • 神経細胞成長におけるアクチンダイナミクスの制御(Regulation of actin dynamics in growing neurons)

    10th Cell Transplant Society Congress  2009 

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  • Mechanism of actin-regulation by dynamin/cortactin complex

    米国細胞生物学会/日本細胞生物学会/理化学研究所 発生・再生科学総合研究センター合同ミーティング  2009 

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