Updated on 2024/03/29

写真b

 
OKA Toshihiko
 
*Items subject to periodic update by Rikkyo University (The rest are reprinted from information registered on researchmap.)
Affiliation*
College of Science Department of Life Science
Graduate School of Science Doctoral Program in Life Science
Graduate School of Science Master's Program in Life Science
Title*
Professor
Degree
PhD (Science) ( The University of Tokyo )
Research Theme*
  • 細胞小器官(オルガネラ)はそれぞれ固有の機能と形態をもっている。オルガネラの機能は細胞の生存にとって必須だが、オルガネラの形態は細胞やオルガネラの機能にとってどのような役割があるのか?この命題を、細胞内のエネルギー産生の場であるミトコンドリアに着目し、分子生物学や生化学的手法を用いて、その膜形態の形成と制御機構を理解することで、オルガネラ形態の生理的意義の解明をめざしている。

  • Campus Career*
    • 4 2012 - Present 
      College of Science   Department of Life Science   Professor
    • 4 2012 - Present 
      Graduate School of Science   Master's Program in Life Science   Professor
    • 4 2012 - Present 
      Graduate School of Science   Doctoral Program in Life Science   Professor
     

    Research Areas

    • Life Science / Cell biology

    Research History

    • 4 2012 - Present 
      RIKKYO UNIVERSITY   Graduate School of Science Field of Study: Life Science   Professor

      More details

    • 4 2012 - Present 
      RIKKYO UNIVERSITY   Graduate School of Science Field of Study: Life Science   Professor

      More details

    • 4 2012 - Present 
      RIKKYO UNIVERSITY   College of Science Department of Life Science   Professor

      More details

    Education

    • 4 1991 - 3 1994 
      The University of Tokyo   Graduate School, Division of Science

      More details

      Country: Japan

      researchmap

    • 4 1989 - 3 1991 
      The University of Tokyo   Graduate School, Division of Science

      More details

      Country: Japan

      researchmap

    • 4 1985 - 3 1989 
      Kyushu University   Faculty of Science   Department of Biology

      More details

      Country: Japan

      researchmap

    Papers

    • Neuronal MML-1/MXL-2 regulates systemic aging via glutamate transporter and cell nonautonomous autophagic and peroxidase activity

      Tatsuya Shioda, Ittetsu Takahashi, Kensuke Ikenaka, Naonobu Fujita, Tomotake Kanki, Toshihiko Oka, Hideki Mochizuki, Adam Antebi, Tamotsu Yoshimori, Shuhei Nakamura

      Proceedings of the National Academy of Sciences120 ( 39 )   18 9 2023

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Proceedings of the National Academy of Sciences  

      Accumulating evidence has demonstrated the presence of intertissue-communication regulating systemic aging, but the underlying molecular network has not been fully explored. We and others previously showed that two basic helix–loop–helix transcription factors, MML-1 and HLH-30, are required for lifespan extension in several longevity paradigms, including germlineless Caenorhabditis elegans . However, it is unknown what tissues these factors target to promote longevity. Here, using tissue-specific knockdown experiments, we found that MML-1 and its heterodimer partners MXL-2 and HLH-30 act primarily in neurons to extend longevity in germlineless animals. Interestingly, however, the downstream cascades of MML-1 in neurons were distinct from those of HLH-30. Neuronal RNA interference (RNAi)-based transcriptome analysis revealed that the glutamate transporter GLT-5 is a downstream target of MML-1 but not HLH-30. Furthermore, the MML-1-GTL-5 axis in neurons is critical to prevent an age-dependent collapse of proteostasis and increased oxidative stress through autophagy and peroxidase MLT-7, respectively, in long-lived animals. Collectively, our study revealed that systemic aging is regulated by a molecular network involving neuronal MML-1 function in both neural and peripheral tissues.

      DOI: 10.1073/pnas.2221553120

      researchmap

    • TIM23 facilitates PINK1 activation by safeguarding against OMA1-mediated degradation in damaged mitochondria Peer-reviewed

      Shiori Akabane, Kiyona Watanabe, Hidetaka Kosako, Shun-ichi Yamashita, Kohei Nishino, Masahiro Kato, Shiori Sekine, Tomotake Kanki, Noriyuki Matsuda, Toshiya Endo, Toshihiko Oka

      Cell Reports   112454 - 112454   5 2023

      More details

      Authorship:Corresponding author   Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1016/j.celrep.2023.112454

      researchmap

    • Localization of myoglobin in mitochondria: implication in regulation of mitochondrial respiration in rat skeletal muscle Peer-reviewed International journal

      Rikuhide Koma, Tsubasa Shibaguchi, Claudia Pérez López, Toshihiko Oka, Thomas Jue, Hisashi Takakura, Kazumi Masuda

      Physiological Reports9 ( 5 ) e14769   3 2021

      More details

      Language:English   Publishing type:Research paper (scientific journal)   Publisher:Wiley  

      Mitochondria play a principal role in metabolism, and mitochondrial respiration is an important process for producing adenosine triphosphate. Recently, we showed the possibility that the muscle-specific protein myoglobin (Mb) interacts with mitochondrial complex IV to augment the respiration capacity in skeletal muscles. However, the precise mechanism for the Mb-mediated upregulation remains under debate. The aim of this study was to ascertain whether Mb is truly integrated into the mitochondria of skeletal muscle and to investigate the submitochondrial localization. Isolated mitochondria from rat gastrocnemius muscle were subjected to different proteinase K (PK) concentrations to digest proteins interacting with the outer membrane. Western blotting analysis revealed that the PK digested translocase of outer mitochondrial membrane 20 (Tom20), and the immunoreactivity of Tom20 decreased with the amount of PK used. However, the immunoreactivity of Mb with PK treatment was better preserved, indicating that Mb is integrated into the mitochondria of skeletal muscle. The mitochondrial protease protection assay experiments suggested that Mb localizes within the mitochondria in the inner membrane from the intermembrane space side. These results strongly suggest that Mb inside muscle mitochondria could be implicated in the regulation of mitochondrial respiration via complex IV.

      DOI: 10.14814/phy2.14769

      PubMed

      researchmap

      Other Link: https://onlinelibrary.wiley.com/doi/full-xml/10.14814/phy2.14769

    • The mitochondrial inner membrane protein LETM1 modulates cristae organization through its LETM domain Peer-reviewed

      Seiko Nakamura, Aiko Matsui, Shiori Akabane, Yasushi Tamura, Azumi Hatano, Yuriko Miyano, Hiroshi Omote, Mizuho Kajikawa, Katsumi Maenaka, Yoshinori Moriyama, Toshiya Endo, Toshihiko Oka

      Communications Biology3 ( 1 )   12 2020

      More details

      Authorship:Corresponding author   Publishing type:Research paper (scientific journal)   Publisher:Springer Science and Business Media LLC  

      Abstract

      LETM1 is a mitochondrial inner membrane protein that is required for maintaining the mitochondrial morphology and cristae structures, and regulates mitochondrial ion homeostasis. Here we report a role of LETM1 in the organization of cristae structures. We identified four amino acid residues of human LETM1 that are crucial for complementation of the growth deficiency caused by gene deletion of a yeast LETM1 orthologue. Substituting amino acid residues with alanine disrupts the correct assembly of a protein complex containing LETM1 and prevents changes in the mitochondrial morphology induced by exogenous LETM1 expression. Moreover, the LETM1 protein changes the shapes of the membranes of in vitro-reconstituted proteoliposomes, leading to the formation of invaginated membrane structures on artificial liposomes. LETM1 mutant proteins with alanine substitutions fail to facilitate the formation of invaginated membrane structures, suggesting that LETM1 plays a fundamental role in the organization of mitochondrial membrane morphology.

      DOI: 10.1038/s42003-020-0832-5

      researchmap

      Other Link: http://www.nature.com/articles/s42003-020-0832-5

    • In vitro synthesis of the human calcium transporter Letm1 within cell-sized liposomes and investigation of its lipid dependency Peer-reviewed

      Kosuke Okamura, Shuhei Matsushita, Yasuhiko Kato, Hajime Watanabe, Aiko Matsui, Toshihiko Oka, Tomoaki Matsuura

      Journal of Bioscience and Bioengineering127 ( 5 ) 544 - 548   5 2019

      More details

      Language:English   Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      The human mitochondrion-derived calcium transporter Letm1 was synthesized by reconstituted in vitro transcription-translation (IVTT) in cell-sized liposomes and the dependency of Letm1 on phospholipid composition was investigated. Components for IVTT were encapsulated into cell-sized vesicles together with the DNA encoding Letm1, thereby preparing proteoliposomes. The synthesis of Letm1 and pH-dependent calcium transport activity were confirmed by flow cytometry. Finally, we investigated the effect of phospholipid composition on Letm1 transport activity and found that cardiolipin present in the mitochondrial membrane plays an important role on the transport activity of Letm1.

      DOI: 10.1016/j.jbiosc.2018.11.003

      PubMed

      researchmap

    • Concentration of mitochondrial DNA mutations by cytoplasmic transfer from platelets to cultured mouse cells. Peer-reviewed

      Ishikawa, K, K. Kobayashi, A. Yamada, M. Umehara, T. Oka, K. Nakada

      PLOS ONE   2019

      More details

      Language:English   Publishing type:Research paper (scientific journal)  

      DOI: 10.1371/journal.pone.0213283

      researchmap

    • Inactivation of cardiolipin synthase triggers changes in mitochondrial morphology Peer-reviewed

      Matsumura, A, Higuchi, J, Watanabe, Y, Kato, M, Aoki, K, Akabane, S, Endo, T, Oka, T

      FEBS Letters592 ( 2 ) 209 - 218   1 1 2018

      More details

      Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:Wiley Blackwell  

      Mitochondrial tubular structures are maintained by a balance between membrane fusion and fission that is regulated by various factors, including Drp1 and mitofusin/fzo-1. Here we report the role of cardiolipin (CL) synthase in the regulation of mitochondrial morphology. Knockdown of CL synthase induced mitochondrial elongation in nematode and human cells. Knockdown of both nematode cardiolipin synthase and drp-1 or fzo-1 suggested that knocking down CL synthase decreases mitochondrial division. Mass spectrometric analysis of human CL synthase-knocked down cells revealed a decreased amount of CL and an accumulation of phosphatidylglycerol, a CL precursor. Knockdown of other genes involved in CL synthesis did not influence mitochondrial morphology. Thus, mitochondrial elongation may result from the accumulation of phosphatidylglycerol rather than decreased CL.

      DOI: 10.1002/1873-3468.12948

      Scopus

      researchmap

    • Structural insights into ubiquitin phosphorylation by PINK1 Peer-reviewed International journal

      Okatsu, K, Y. Sato, K. Yamano, N. Matsuda, L. Negishi, A. Takahashi, A. Yamagata, S. Goto-Ito, M. Mishima, Y. Ito, T. Oka, K. Tanaka, S. Fukai

      Sci. Rep.8 ( 1 ) 10382 - 10382   2018

      More details

      Language:English   Publishing type:Research paper (scientific journal)  

      Mutations of PTEN-induced putative kinase 1 (PINK1) and the E3 ubiquitin (Ub) ligase parkin can cause familial parkinsonism. These two proteins are essential for ubiquitylation of damaged mitochondria and subsequent degradation. PINK1 phosphorylates Ser65 of Ub and the Ub-like (UBL) domain of parkin to allosterically relieve the autoinhibition of parkin. To understand the structural mechanism of the Ub/UBL-specific phosphorylation by PINK1, we determined the crystal structure of Tribolium castaneum PINK1 kinase domain (TcPINK1) in complex with a nonhydrolyzable ATP analogue at 2.5 Å resolution. TcPINK1 consists of the N- and C-terminal lobes with the PINK1-specific extension. The ATP analogue is bound in the cleft between the N- and C-terminal lobes. The adenine ring of the ATP analogue is bound to a hydrophobic pocket, whereas the triphosphate group of the ATP analogue and two coordinated Mg ions interact with the catalytic hydrophilic residues. Comparison with protein kinases A and C (PKA and PKC, respectively) unveils a putative Ub/UBL-binding groove, which is wider than the peptide-binding groove of PKA or PKC to accommodate the globular head of Ub or UBL. Further crosslinking analyses suggested a PINK1-interacting surface of Ub. Structure-guided mutational analyses support the findings from the present structural analysis of PINK1.

      DOI: 10.1038/s41598-018-28656-8

      PubMed

      researchmap

    • Molecular basis of selective mitochondrial fusion by heterotypic action between OPA1 and cardiolipin. Peer-reviewed International journal

      Tadato Ban, Takaya Ishihara, Hiroto Kohno, Shotaro Saita, Ayaka Ichimura, Katsumi Maenaka, Toshihiko Oka, Katsuyoshi Mihara, Naotada Ishihara

      Nature cell biology19 ( 7 ) 856 - 863   7 2017

      More details

      Language:English   Publishing type:Research paper (scientific journal)  

      Mitochondria are highly dynamic organelles that undergo frequent fusion and fission. Optic atrophy 1 (OPA1) is an essential GTPase protein for both mitochondrial inner membrane (IM) fusion and cristae morphology. Under mitochondria-stress conditions, membrane-anchored L-OPA1 is proteolytically cleaved to form peripheral S-OPA1, leading to the selection of damaged mitochondria for mitophagy. However, molecular details of the selective mitochondrial fusion are less well understood. Here, we showed that L-OPA1 and cardiolipin (CL) cooperate in heterotypic mitochondrial IM fusion. We reconstituted an in vitro membrane fusion reaction using purified human L-OPA1 protein expressed in silkworm, and found that L-OPA1 on one side of the membrane and CL on the other side are sufficient for fusion. GTP-independent membrane tethering through L-OPA1 and CL primes the subsequent GTP-hydrolysis-dependent fusion, which can be modulated by the presence of S-OPA1. These results unveil the most minimal intracellular membrane fusion machinery. In contrast, independent of CL, a homotypic trans-OPA1 interaction mediates membrane tethering, thereby supporting the cristae structure. Thus, multiple OPA1 functions are modulated by local CL conditions for regulation of mitochondrial morphology and quality control.

      DOI: 10.1038/ncb3560

      PubMed

      researchmap

    • Constitutive Activation of PINK1 Protein Leads to Proteasome-mediated and Non-apoptotic Cell Death Independently of Mitochondrial Autophagy Peer-reviewed

      Shiori Akabane, Kohei Matsuzaki, Shun-ichi Yamashita, Kana Arai, Kei Okatsu, Tomotake Kanki, Noriyuki Matsuda, Toshihiko Oka

      JOURNAL OF BIOLOGICAL CHEMISTRY291 ( 31 ) 16162 - 16174   7 2016

      More details

      Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC  

      Phosphatase and tensin homolog-induced putative kinase 1 (PINK1), a Ser/Thr kinase, and PARKIN, a ubiquitin ligase, are causal genes for autosomal recessive early-onset parkinsonism. Multiple lines of evidence indicate that PINK1 and PARKIN cooperatively control the quality of the mitochondrial population via selective degradation of damaged mitochondria by autophagy. Here, we report that PINK1 and PARKIN induce cell death with a 12-h delay after mitochondrial depolarization, which differs from the time profile of selective autophagy of mitochondria. This type of cell death exhibited definite morphologic features such as plasma membrane rupture, was insensitive to a pan-caspase inhibitor, and did not involve mitochondrial permeability transition. Expression of a constitutively active form of PINK1 caused cell death in the presence of a pan-caspase inhibitor, irrespective of the mitochondrial membrane potential. PINK1-mediated cell death depended on the activities of PARKIN and proteasomes, but it was not affected by disruption of the genes required for autophagy. Furthermore, fluorescence and electron microscopic analyses revealed that mitochondria were still retained in the dead cells, indicating that PINK1-mediated cell death is not caused by mitochondrial loss. Our findings suggest that PINK1 and PARKIN play critical roles in selective cell death in which damaged mitochondria are retained, independent of mitochondrial autophagy.

      DOI: 10.1074/jbc.M116.714923

      researchmap

    • PKA Regulates PINK1 Stability and Parkin Recruitment to Damaged Mitochondria through Phosphorylation of MIC60 Peer-reviewed

      Shiori Akabane, Midori Uno, Naoki Tani, Shunta Shimazaki, Natsumi Ebara, Hiroki Kato, Hidetaka Kosako, Toshihiko Oka

      MOLECULAR CELL62 ( 3 ) 371 - 384   5 2016

      More details

      Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:CELL PRESS  

      A mitochondrial kinase, PTEN-induced putative kinase 1 (PINK1), selectively recruits the ubiquitin ligase Parkin to damaged mitochondria, which modifies mitochondria by polyubiquitination, leading to mitochondrial autophagy. Here, we report that treatment with an adenylate cyclase agonist or expression of protein kinase A (PKA) impairs Parkin recruitment to damaged mitochondria and decreases PINK1 protein levels. We identified a mitochondrial membrane protein, MIC60 (also known as mitofilin), as a PKA substrate. Mutational and mass spectrometric analyses revealed that the Ser528 residue of MIC60 undergoes PKA-dependent phosphorylation. MIC60 transiently interacts with PINK1, and MIC60 downregulation leads to a reduction in PINK1 and mislocalization of Parkin. Phosphorylation-mimic mutants of MIC60 fail to restore the defect in Parkin recruitment in MIC60-knocked down cells, whereas a phosphorylation-deficient MIC60 mutant facilitates the mitochondrial localization of Parkin. Our findings indicate that PKA-mediated phosphorylation of MIC60 negatively regulates mitochondrial clearance that is initiated by PINK1 and Parkin.

      DOI: 10.1016/j.molcel.2016.03.037

      researchmap

    • Unconventional PINK1 localization to the outer membrane of depolarized mitochondria drives Parkin recruitment Peer-reviewed

      Kei Okatsu, Mayumi Kimura, Toshihiko Oka, Keiji Tanaka, Noriyuki Matsuda

      JOURNAL OF CELL SCIENCE128 ( 5 ) 964 - 978   3 2015

      More details

      Language:English   Publishing type:Research paper (scientific journal)   Publisher:COMPANY OF BIOLOGISTS LTD  

      Dysfunction of PTEN-induced putative kinase 1 (PINK1), a Ser/Thr kinase with an N-terminal mitochondrial-targeting sequence (MTS), causes familial recessive parkinsonism. Reduction of the mitochondrial membrane potential limits MTS-mediated matrix import and promotes PINK1 accumulation on the outer mitochondrial membrane (OMM) of depolarized mitochondria. PINK1 then undergoes autophosphorylation and phosphorylates ubiquitin and Parkin, a cytosolic ubiquitin ligase, for clearance of damaged mitochondria. The molecular basis for PINK1 localization on the OMM of depolarized mitochondria rather than release to the cytosol is poorly understood. Here, we disentangle the PINK1 localization mechanism using deletion mutants and a newly established constitutively active PINK1 mutant. Disruption of the MTS through N-terminal insertion of aspartic acid residues results in OMM localization of PINK1 in energized mitochondria. Unexpectedly, the MTS and putative transmembrane domain (TMD) are dispensable for OMM localization, whereas mitochondrial translocase Tom40 (also known as TOMM40) and an alternative mitochondrial localization signal that resides between the MTS and TMD are required. PINK1 utilizes a mitochondrial localization mechanism that is distinct from that of conventional MTS proteins and that presumably functions in conjunction with the Tom complex in OMM localization when the conventional N-terminal MTS is inhibited.

      DOI: 10.1242/jcs.161000

      researchmap

    • A dimeric pink1-containing complex on depolarized mitochondria stimulates parkin recruitment Peer-reviewed

      Kei Okatsu, Midori Uno, Fumika Koyano, Etsu Go, Mayumi Kimura, Toshihiko Oka, Keiji Tanaka, Noriyuki Matsuda

      Journal of Biological Chemistry288 ( 51 ) 36372 - 36384   20 12 2013

      More details

      Language:English   Publishing type:Research paper (scientific journal)  

      Parkinsonism typified by sporadic Parkinson disease is a prevalent neurodegenerative disease. Mutations in PINK1 (PTENinduced putative kinase 1), a mitochondrial Ser/Thr protein kinase, or PARKIN, a ubiquitin-protein ligase, cause familial parkinsonism. The accumulation and autophosphorylation of PINK1 on damaged mitochondria results in the recruitment of Parkin, which ultimately triggers quarantine and/or degradation of the damaged mitochondria by the proteasome and autophagy. However, the molecular mechanism of PINK1 in dissipation of the mitochondrial membrane potential (ΔΨm) has not been fully elucidated. Here we show by fluorescence-based techniques that the PINK1 complex formed following a decrease in ΔΨm is composed of two PINK1 molecules and is correlated with intermolecular phosphorylation of PINK1. Disruption of complex formation by the PINK1 S402A mutation weakened Parkin recruitment onto depolarized mitochondria. The most disease-relevant mutations of PINK1 inhibit the complex formation. Taken together, these results suggest that formation of the complex containing dyadic PINK1 is an important step for Parkin recruitment onto damaged mitochondria. © 2013 by The American Society for Biochemistry and Molecular Biology.

      DOI: 10.1074/jbc.M113.509653

      Scopus

      PubMed

      researchmap

    • Fis1 acts as a mitochondrial recruitment factor for TBC1D15 that is involved in regulation of mitochondrial morphology. Peer-reviewed International journal

      Kenta Onoue, Akihiro Jofuku, Reiko Ban-Ishihara, Takaya Ishihara, Maki Maeda, Takumi Koshiba, Takashi Itoh, Mitsunori Fukuda, Hidenori Otera, Toshihiko Oka, Hiroyoshi Takano, Noboru Mizushima, Katsuyoshi Mihara, Naotada Ishihara

      Journal of cell science126 ( Pt 1 ) 176 - 85   1 1 2013

      More details

      Language:English   Publishing type:Research paper (scientific journal)  

      In yeast, C-tail-anchored mitochondrial outer membrane protein Fis1 recruits the mitochondrial-fission-regulating GTPase Dnm1 to mitochondrial fission sites. However, the function of its mammalian homologue remains enigmatic because it has been reported to be dispensable for the mitochondrial recruitment of Drp1, a mammalian homologue of Dnm1. We identified TBC1D15 as a Fis1-binding protein in HeLa cell extracts. Immunoprecipitation revealed that Fis1 efficiently interacts with TBC1D15 but not with Drp1. Bacterially expressed Fis1 and TBC1D15 formed a direct and stable complex. Exogenously expressed TBC1D15 localized mainly in cytoplasm in HeLa cells, but when coexpressed with Fis1 it localized to mitochondria. Knockdown of TBC1D15 induced highly developed mitochondrial network structures similar to the effect of Fis1 knockdown, suggesting that the TBC1D15 and Fis1 are associated with the regulation of mitochondrial morphology independently of Drp1. These data suggest that Fis1 acts as a mitochondrial receptor in the recruitment of mitochondrial morphology protein in mammalian cells.

      DOI: 10.1242/jcs.111211

      PubMed

      researchmap

    • PINK1 autophosphorylation upon membrane potential dissipation is essential for Parkin recruitment to damaged mitochondria Peer-reviewed

      Kei Okatsu, Toshihiko Oka, Masahiro Iguchi, Kenji Imamura, Hidetaka Kosako, Naoki Tani, Mayumi Kimura, Etsu Go, Fumika Koyano, Manabu Funayama, Kahori Shiba-Fukushima, Shigeto Sato, Hideaki Shimizu, Yuko Fukunaga, Hisaaki Taniguchi, Masaaki Komatsu, Nobutaka Hattori, Katsuyoshi Mihara, Keiji Tanaka, Noriyuki Matsuda

      NATURE COMMUNICATIONS3   8 2012

      More details

      Language:English   Publishing type:Research paper (scientific journal)   Publisher:NATURE PUBLISHING GROUP  

      Dysfunction of PINK1, a mitochondrial Ser/Thr kinase, causes familial Parkinson's disease (PD). Recent studies have revealed that PINK1 is rapidly degraded in healthy mitochondria but accumulates on the membrane potential (Delta Psi m)-deficient mitochondria, where it recruits another familial PD gene product, Parkin, to ubiquitylate the damaged mitochondria. Despite extensive study, the mechanism underlying the homeostatic control of PINK1 remains unknown. Here we report that PINK1 is autophosphorylated following a decrease in Delta Psi mm and that most disease-relevant mutations hinder this event. Mass spectrometric and mutational analyses demonstrate that PINK1 autophosphorylation occurs at Ser228 and Ser402, residues that are structurally clustered together. Importantly, Ala mutation of these sites abolishes autophosphorylation of PINK1 and inhibits Parkin recruitment onto depolarized mitochondria, whereas Asp (phosphorylation-mimic) mutation promotes mitochondrial localization of Parkin even though autophosphorylation was still compromised. We propose that autophosphorylation of Ser228 and Ser402 in PINK1 is essential for efficient mitochondrial localization of Parkin.

      DOI: 10.1038/ncomms2016

      researchmap

    • KLP6: a newly identified kinesin that regulates the morphology and transport of mitochondria in neuronal cells Peer-reviewed

      Kousuke Tanaka, Yoshimi Sugiura, Ryohei Ichishita, Katsuyoshi Mihara, Toshihiko Oka

      JOURNAL OF CELL SCIENCE124 ( 14 ) 2457 - 2465   7 2011

      More details

      Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:COMPANY OF BIOLOGISTS LTD  

      Mitochondria utilize diverse cytoskeleton-based mechanisms to control their functions and morphology. Here, we report a role for kinesin-like protein KLP6, a newly identified member of the kinesin family, in mitochondrial morphology and dynamics. An RNA interference screen using Caenorhabditis elegans led us to identify a C. elegans KLP-6 involved in maintaining mitochondrial morphology. We cloned a cDNA coding for a rat homolog of C. elegans KLP-6, which is an uncharacterized kinesin in vertebrates. A rat KLP6 mutant protein lacking the motor domain induced changes in mitochondrial morphology and significantly decreased mitochondrial motility in HeLa cells, but did not affect the morphology of other organelles. In addition, the KLP6 mutant inhibited transport of mitochondria during anterograde movement in differentiated neuro 2a cells. To date, two kinesins, KIF1B alpha and kinesin heavy chain (KHC; also known as KIF5) have been shown to be involved in the distribution of mitochondria in neurons. Expression of the kinesin heavy chain/KIF5 mutant prevented mitochondria from entering into neurites, whereas both the KLP6 and KIF1B alpha mutants decreased mitochondrial transport in axonal neurites. Furthermore, both KLP6 and KIF1B alpha bind to KBP, a KIF1-binding protein required for axonal outgrowth and mitochondrial distribution. Thus, KLP6 is a newly identified kinesin family member that regulates mitochondrial morphology and transport.

      DOI: 10.1242/jcs.086470

      researchmap

    • Characterization of the mitochondrial protein LETM1, which maintains the mitochondrial tubular shapes and interacts with the AAA-ATPase BCS1L Peer-reviewed

      Shoko Tamai, Hiroshi Iida, Sadaki Yokota, Tomoko Sayano, Shoko Kiguchiya, Naotada Ishihara, Jun-Ichi Hayashi, Katsuyoshi Mihara, Toshihiko Oka

      JOURNAL OF CELL SCIENCE121 ( 15 ) 2588 - 2600   8 2008

      More details

      Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:COMPANY OF BIOLOGISTS LTD  

      LETM1 is located in the chromosomal region that is deleted in patients suffering Wolf-Hirschhorn syndrome; it encodes a homolog of the yeast protein Mdm38 that is involved in mitochondrial morphology. Here, we describe the LETM1-mediated regulation of the mitochondrial volume and its interaction with the mitochondrial AAA-ATPase BCS1L that is responsible for three different human disorders. LETM1 is a mitochondrial inner-membrane protein with a large domain extruding to the matrix. The LETM1 homolog LETM2 is a mitochondrial protein that is expressed preferentially in testis and sperm. LETM1 downregulation caused mitochondrial swelling and cristae disorganization, but seemed to have little effect on membrane fusion and fission. Formation of the respiratory-chain complex was impaired by LETM1 knockdown. Cells lacking mitochondrial DNA lost active respiratory chains but maintained mitochondrial tubular networks, indicating that mitochondrial swelling caused by LETM1 knockdown is not caused by the disassembly of the respiratory chains. LETM1 was co-precipitated with BCS1L and formation of the LETM1 complex depended on BCS1L levels, suggesting that BCS1L stimulates the assembly of the LETM1 complex. BCS1L knockdown caused disassembly of the respiratory chains as well as LETM1 downregulation and induced distinct changes in mitochondrial morphology.

      DOI: 10.1242/jcs.026625

      researchmap

    • Identification of a novel protein MICS1 that is involved in maintenance of mitochondrial morphology and apoptotic release of cytochrome c Peer-reviewed

      Toshihiko Oka, Tomoko Sayano, Shoko Tamai, Sadaki Yokota, Hiroki Kato, Gen Fujii, Katsuyoshi Mihara

      MOLECULAR BIOLOGY OF THE CELL19 ( 6 ) 2597 - 2608   6 2008

      More details

      Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:AMER SOC CELL BIOLOGY  

      Mitochondrial morphology dynamically changes in a balance of membrane fusion and fission in response to the environment, cell cycle, and apoptotic stimuli. Here, we report that a novel mitochondrial protein, MICS1, is involved in mitochondrial morphology in specific cristae structures and the apoptotic release of cytochrome c from the mitochondria. MICS1 is an inner membrane protein with a cleavable presequence and multiple transmembrane segments and belongs to the Bi-1 super family. MICS1 down-regulation causes mitochondrial fragmentation and cristae disorganization and stimulates the release of proapoptotic proteins. Expression of the anti-apoptotic protein Bcl-XL does not prevent morphological changes of mitochondria caused by MICS1 down-regulation, indicating that MICS1 plays a role in maintaining mitochondrial morphology separately from the function in apoptotic pathways. MICS1 overproduction induces mitochondrial aggregation and partially inhibits cytochrome c release during apoptosis, regardless of the occurrence of Bax targeting. MICS1 is cross-linked to cytochrome c without disrupting membrane integrity. Thus, MICS1 facilitates the tight association of cytochrome c with the inner membrane. Furthermore, under low-serum condition, the delay in apoptotic release of cytochrome c correlates with MICS1 up-regulation without significant changes in mitochondrial morphology, suggesting that MICS1 individually functions in mitochondrial morphology and cytochrome c release.

      DOI: 10.1091/mbc.E07-12-1205

      researchmap

    • An RNAi screen for mitochondrial proteins required to maintain the morphology of the organelle in Caenorhabditis elegans Peer-reviewed

      Ryohei Ichishita, Kousuke Tanaka, Yoshimi Sugiura, Tomoko Sayano, Katsuyoshi Mihara, Toshihiko Oka

      JOURNAL OF BIOCHEMISTRY143 ( 4 ) 449 - 454   4 2008

      More details

      Authorship:Corresponding author   Language:English   Publishing type:Research paper (scientific journal)   Publisher:OXFORD UNIV PRESS  

      Mitochondria are dynamic organelles that frequently divide and fuse together, resulting in the formation of intracellular tubular networks. In yeast and mammals, several factors including Drp1/Dnm1 and Mfn/Fzo1 are known to regulate mitochondrial morphology by controlling membrane fission or fusion. Here, we report the systematic screening of Caenorhabditis elegans mitochondrial proteins required to maintain the morphology of the organelle using an RNA interference feeding library. In C. elegans body wall muscle cells, mitochondria usually formed tubular structures and were severely fragmented by the mutation in fzo-1 gene, indicating that the body wall muscle cells are suitable for monitoring changes in mitochondrial morphology due to gene silencing. Of 719 genes predicted to code for most of mitochondrial proteins, knockdown of >80% of them caused abnormal mitochondrial morphology, including fragmentation and elongation. These findings indicate that most fundamental mitochondrial functions, including metabolism and oxidative phosphorylation, are necessary for maintenance of the tubular networks as well as membrane fission and fusion. This is the first evidence that known mitochondrial activities are prerequisite for regulating the morphology of the organelle. Furthermore, 88 uncharacterized or poorly characterized genes were found in the screening to be implicated in mitochondrial morphology.

      DOI: 10.1093/jb/mvm245

      researchmap

    • Mitotic Phosphorylation of Dynamin-related GTPase Drp1 Participates in Mitochondrial Fission Peer-reviewed

      Naoko Taguchi, Naotada Ishihara, Akihiro Jofuku, Toshihiko Oka, Katsuyoshi Mihara

      Journal of Biological Chemistry282 ( 15 ) 11521 - 11529   4 2007

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.m607279200

      researchmap

    • Identification and Characterization of a New Tom40 Isoform, a Central Component of Mitochondrial Outer Membrane Translocase Peer-reviewed

      J.-y. Kinoshita, K. Mihara, T. Oka

      Journal of Biochemistry141 ( 6 ) 897 - 906   27 3 2007

      More details

      Authorship:Corresponding author   Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

      DOI: 10.1093/jb/mvm097

      researchmap

    • A novel insertion pathway of mitochondrial outer membrane proteins with multiple transmembrane segments Peer-reviewed

      Otera H, Taira Y, Horie C, Suzuki Y, Suzuki H, Setoguchi K, Kato H, Oka T, Mihara K

      Journal of Cell Biology179 ( 7 ) 1355 - 1363   2007

    • Identification of a novel protein that regulates mitochondrial fusion by modulating mitofusin (Mfn) protein function Peer-reviewed

      Yuka Eura, Naotada Ishihara, Toshihiko Oka, Katsuyoshi Mihara

      Journal of Cell Science119 ( 23 ) 4913 - 4925   1 12 2006

      More details

      Publishing type:Research paper (scientific journal)   Publisher:The Company of Biologists  

      Mitofusin proteins 1 and 2 (Mfn1 and Mfn2, respectively) of the mammalian mitochondrial outer membrane are homologues of Drosophila FZO and yeast Fzo1, and both are essential for GTP-dependent mitochondrial fusion. We identified a 55-kDa Mfn-binding protein named MIB. It is a member of the medium-chain dehydrogenase/reductase protein superfamily, and has a conserved coenzyme-binding domain (CBD). The majority of MIB is localized in the cytoplasm but a small amount is associated with mitochondria. Exogenous expression of MIB in HeLa cells induced mitochondrial fragmentation, which was prevented by coexpression of Mfn1, suggesting a functional interaction of MIB with Mfn proteins; the GGVG sequence in the CBD of MIB is essential for its function. By contrast, MIB knockdown resulted in growth arrest of the cells, although apoptotic sensitivity was not affected by either its knockdown or its overexpression. Furthermore, MIB knockdown induced a large extension of mitochondrial network structures. By contrast, a double knockdown of MIB and Mfn1 resulted in mitochondrial fragmentation and reversal of the growth arrest, the morphology and growth phenotype induced by knockdown of Mfn1 alone, again suggesting that MIB modulates Mfn1 function. Together, these findings suggest that MIB is essential for cellular function by regulating mitochondrial membrane dynamics in cooperation with Mfn proteins.

      DOI: 10.1242/jcs.03253

      researchmap

    • IntraGolgi distribution of the Conserved Oligomeric Golgi (COG) complex Peer-reviewed

      Eliza Vasile, Toshihiko Oka, Maria Ericsson, Nobuhiro Nakamura, Monty Krieger

      Experimental Cell Research312 ( 16 ) 3132 - 3141   10 2006

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1016/j.yexcr.2006.06.005

      researchmap

    • Regulation of mitochondrial morphology through proteolytic cleavage of OPA1 Peer-reviewed

      Naotada Ishihara, Yuu Fujita, Toshihiko Oka, Katsuyoshi Mihara

      The EMBO Journal25 ( 13 ) 2966 - 2977   12 7 2006

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Wiley  

      DOI: 10.1038/sj.emboj.7601184

      researchmap

    • Subunit Architecture of the Conserved Oligomeric Golgi Complex Peer-reviewed

      Daniel Ungar, Toshihiko Oka, Eliza Vasile, Monty Krieger, Frederick M. Hughson

      Journal of Biological Chemistry280 ( 38 ) 32729 - 32735   9 2005

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.m504590200

      researchmap

    • Genetic Analysis of the Subunit Organization and Function of the Conserved Oligomeric Golgi (COG) Complex Peer-reviewed

      Toshihiko Oka, Eliza Vasile, Marsha Penman, Carl D. Novina, Derek M. Dykxhoorn, Daniel Ungar, Frederick M. Hughson, Monty Krieger

      Journal of Biological Chemistry280 ( 38 ) 32736 - 32745   9 2005

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.m505558200

      researchmap

    • The COG and COPI Complexes Interact to Control the Abundance of GEARs, a Subset of Golgi Integral Membrane Proteins Peer-reviewed

      Toshihiko Oka, Daniel Ungar, Frederick M. Hughson, Monty Krieger

      Molecular Biology of the Cell15 ( 5 ) 2423 - 2435   5 2004

      More details

      Publishing type:Research paper (scientific journal)   Publisher:American Society for Cell Biology (ASCB)  

      The conserved oligomeric Golgi (COG) complex is a soluble hetero-octamer associated with the cytoplasmic surface of the Golgi. Mammalian somatic cell mutants lacking the Cog1 (ldlB) or Cog2 (ldlC) subunits exhibit pleiotropic defects in Golgi-associated glycoprotein and glycolipid processing that suggest COG is involved in the localization, transport, and/or function of multiple Golgi processing proteins. We have identified a set of COG-sensitive, integral membrane Golgi proteins called GEARs (mannosidase II, GOS-28, GS15, GPP130, CASP, giantin, and golgin-84) whose abundances were reduced in the mutant cells and, in some cases, increased in COG-overexpressing cells. In the mutants, some GEARs were abnormally localized in the endoplasmic reticulum and were degraded by proteasomes. The distributions of the GEARs were altered by small interfering RNA depletion of ϵ-COP in wild-type cells under conditions in which COG-insensitive proteins were unaffected. Furthermore, synthetic phenotypes arose in mutants deficient in both ϵ-COP and either Cog1 or Cog2. COG and COPI may work in concert to ensure the proper retention or retrieval of a subset of proteins in the Golgi, and COG helps prevent the endoplasmic reticulum accumulation and degradation of some GEARs.

      DOI: 10.1091/mbc.e03-09-0699

      researchmap

    • From Lysosomes to the Plasma Membrane Peer-reviewed

      Takao Toyomura, Yoshiko Murata, Akitsugu Yamamoto, Toshihiko Oka, Ge-Hong Sun-Wada, Yoh Wada, Masamitsu Futai

      Journal of Biological Chemistry278 ( 24 ) 22023 - 22030   6 2003

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.m302436200

      researchmap

    • Characterization of a mammalian Golgi-localized protein complex, COG, that is required for normal Golgi morphology and function Peer-reviewed

      Daniel Ungar, Toshihiko Oka, Elizabeth E. Brittle, Eliza Vasile, Vladimir V. Lupashin, Jon E. Chatterton, John E. Heuser, Monty Krieger, M. Gerard Waters

      Journal of Cell Biology157 ( 3 ) 405 - 415   29 4 2002

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Rockefeller University Press  

      Multiprotein complexes are key determinants of Golgi apparatus structure and its capacity for intracellular transport and glycoprotein modification. Three complexes that have previously been partially characterized include (a) the Golgi transport complex (GTC), identified in an in vitro membrane transport assay, (b) the ldlCp complex, identified in analyses of CHO cell mutants with defects in Golgi-associated glycosylation reactions, and (c) the mammalian Sec34 complex, identified by homology to yeast Sec34p, implicated in vesicular transport. We show that these three complexes are identical and rename them the conserved oligomeric Golgi (COG) complex. The COG complex comprises four previously characterized proteins (Cog1/ldlBp, Cog2/ldlCp, Cog3/Sec34, and Cog5/GTC-90), three homologues of yeast Sec34/35 complex subunits (Cog4, -6, and -8), and a previously unidentified Golgi-associated protein (Cog7). EM of ldlB and ldlC mutants established that COG is required for normal Golgi morphology. “Deep etch” EM of purified COG revealed an ∼37-nm-long structure comprised of two similarly sized globular domains connected by smaller extensions. Consideration of biochemical and genetic data for mammalian COG and its yeast homologue suggests a model for the subunit distribution within this complex, which plays critical roles in Golgi structure and function.

      DOI: 10.1083/jcb.200202016

      researchmap

    • V-o and V-1 subunit isoforms of mouse vacuolar proton ATPase V1Vo Peer-reviewed

      GH Sun-Wada, T Yoshimizu, Y Murata, T Oka, Y Wada, M Futai

      FASEB JOURNAL16 ( 4 ) A154 - A154   3 2002

      More details

      Language:English   Publisher:FEDERATION AMER SOC EXP BIOL  

      researchmap

    • Structure and properties of isoforms of mouse vacuolar proton ATPase subunits

      GH Sun-Wada, T Yoshimizu, Y Murata, T Oka, Y Wada, M Futai

      MOLECULAR BIOLOGY OF THE CELL12   251A - 251A   11 2001

      More details

      Language:English   Publisher:AMER SOC CELL BIOLOGY  

      researchmap

    • a4, a Unique Kidney-specific Isoform of Mouse Vacuolar H+-ATPase Subunit a Peer-reviewed

      Toshihiko Oka, Yoshiko Murata, Miwako Namba, Takao Yoshimizu, Takao Toyomura, Akitsugu Yamamoto, Ge-Hong Sun-Wada, Naotaka Hamasaki, Yoh Wada, Masamitsu Futai

      Journal of Biological Chemistry276 ( 43 ) 40050 - 40054   10 2001

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.m106488200

      researchmap

    • Four Subunit a Isoforms ofCaenorhabditis elegans Vacuolar H+-ATPase Peer-reviewed

      Toshihiko Oka, Takao Toyomura, Kenta Honjo, Yoh Wada, Masamitsu Futai

      Journal of Biological Chemistry276 ( 35 ) 33079 - 33085   8 2001

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.m101652200

      researchmap

    • Requirement of V-ATPase for Ovulation and Embryogenesis inCaenorhabditis elegans Peer-reviewed

      Toshihiko Oka, Masamitsu Futai

      Journal of Biological Chemistry275 ( 38 ) 29556 - 29561   9 2000

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.m002756200

      researchmap

    • Upstream Regions Directing Heart-Specific Expression of the GATA6 Gene During Mouse Early Development Peer-reviewed

      G.-H. Sun-Wada, S. Manabe, T. Yoshimizu, C. Yamaguchi, T. Oka, Y. Wada, M. Futai

      Journal of Biochemistry127 ( 4 ) 703 - 709   1 4 2000

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

      DOI: 10.1093/oxfordjournals.jbchem.a022660

      researchmap

    • Three Subunit a Isoforms of Mouse Vacuolar H+-ATPase Peer-reviewed

      Takao Toyomura, Toshihiko Oka, Chie Yamaguchi, Yoh Wada, Masamitsu Futai

      Journal of Biological Chemistry275 ( 12 ) 8760 - 8765   3 2000

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.275.12.8760

      researchmap

    • Disruption of clh-1, a chloride channel gene, results in a wider body of Caenorhabditis elegans Peer-reviewed

      Mark I.R Petalcorin, Toshihiko Oka, Makoto Koga, Ken-ichi Ogura, Yoh Wada, Yasumi Ohshima, Masamitsu Futai

      Journal of Molecular Biology294 ( 2 ) 347 - 355   11 1999

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1006/jmbi.1999.3241

      researchmap

    • A pair of invertedly repeated genes in Chlamydomonas reinhardtii encodes a zygote-specific protein whose expression is UV-sensitive Peer-reviewed

      Hidenobu Uchida, Lena Suzuki, Toyoaki Anai, Koji Doi, Hiroyoshi Takano, Hirofumi Yamashita, Toshihiko Oka, Shigeyuki Kawano, Ken-Ichi Tomizawa, Tamotsu Kawazu, Haruko Kuroiwa, Tsuneyoshi Kuroiwa

      Current Genetics36 ( 4 ) 232 - 240   13 10 1999

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Springer Science and Business Media LLC  

      DOI: 10.1007/s002940050495

      researchmap

      Other Link: http://link.springer.com/article/10.1007/s002940050495/fulltext.html

    • Identification of SEC12, SED4, Truncated SEC16, and EKS1/HRD3 as Multicopy Suppressors of ts Mutants of Sari GTPase Peer-reviewed International journal

      Y. Saito, T. Yamanushi, T. Oka, A. Nakano

      Journal of Biochemistry125 ( 1 ) 130 - 137   1 1 1999

      More details

      Language:English   Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

      The yeast SAR1 gene encodes a low-molecular-weight GTPase which is essential for the formation of transport vesicles from the endoplasmic reticulum (ER). To understand how the Sar1p function is regulated in its GTPase cycle, we searched for multicopy suppressors of sar1 temperature-sensitive mutants and identified SEC12, SED4, truncated SEC16, and EKS1. EKS1 turns out to be identical to HRD3, which was independently isolated as a gene implicated in the degradation of an HMG-CoA reductase isozyme, Hmg2p. In this paper, we show that the product of EKS1/HRD3 is a type-I transmembrane glycoprotein and resides in the ER. The eks1/hrd3 disrupted cells are normal in growth and transport of cargo proteins, but missecrete BiP (Kar2p). The overexpression of EKS1/HRD3, which stabilizes Hmg2p, did not affect the stability of wild-type or mutant Sar1p or any early Sec proteins we examined. These results suggest that the role of Eks1p/Hrd3p is not involved in the ER protein degradation in general but rather required for the maintenance of the ER membrane functions. The novel genetic interactions unveiled between SAR1, SEC12, SEC16, and SED4 will provide useful information as to how the complex machinery of vesicle budding operates.

      DOI: 10.1093/oxfordjournals.jbchem.a022249

      PubMed

      researchmap

    • Identification and Cloning of Rat Galectin-2: Expression Is Predominantly in Epithelial Cells of the Stomach Peer-reviewed

      Toshihiko Oka, Seiko Murakami, Yoichiro Arata, Jun Hirabayashi, Ken-Ichi Kasai, Yoh Wada, Masamitsu Futai

      Archives of Biochemistry and Biophysics361 ( 2 ) 195 - 201   1 1999

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1006/abbi.1998.0968

      researchmap

    • Identification of the copper chaperone, CUC-1, in<i>Caenorhabditis elegans</i>: tissue specific co-expression with the copper transporting ATPase, CUA-1 Peer-reviewed

      Tokumitsu Wakabayashi, Norihiro Nakamura, Yoshihiro Sambongi, Yoh Wada, Toshihiko Oka, Masamitsu Futai

      FEBS Letters440 ( 1-2 ) 141 - 146   27 11 1998

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Wiley  

      DOI: 10.1016/s0014-5793(98)01431-8

      researchmap

    • Activities of Mutant Sar1 Proteins in Guanine Nucleotide Binding, GTP Hydrolysis, and Cell-Free Transport from the Endoplasmic Reticulum to the Golgi Apparatus Peer-reviewed

      Y. Saito, K. Kimura, T. Oka, A. Nakano

      Journal of Biochemistry124 ( 4 ) 816 - 823   1 10 1998

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

      DOI: 10.1093/oxfordjournals.jbchem.a022185

      researchmap

    • Multiple Genes for Vacuolar-type ATPase Proteolipids inCaenorhabditis elegans Peer-reviewed

      Toshihiko Oka, Ryuji Yamamoto, Masamitsu Futai

      Journal of Biological Chemistry273 ( 35 ) 22570 - 22576   8 1998

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.273.35.22570

      researchmap

    • Three vha Genes Encode Proteolipids ofCaenorhabditis elegans Vacuolar-type ATPase Peer-reviewed

      Toshihiko Oka, Ryuji Yamamoto, Masamitsu Futai

      Journal of Biological Chemistry272 ( 39 ) 24387 - 24392   9 1997

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1074/jbc.272.39.24387

      researchmap

    • Caenorhabditis elegans cDNA for a Menkes/Wilson Disease Gene Homologue and Its Function in a Yeast CCC2 Gene Deletion Mutant Peer-reviewed

      Y. Sambongi, T. Wakabayashi, T. Yoshimizu, H. Omote, T. Oka, M. Futai

      Journal of Biochemistry121 ( 6 ) 1169 - 1175   1 6 1997

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

      DOI: 10.1093/oxfordjournals.jbchem.a021711

      researchmap

    • Characterization of Yeast sar1 Temperature-Sensitive Mutants, Which Are Defective in Protein Transport from the Endoplasmic Reticulum Peer-reviewed

      T. Yamanush, A. Hirata, T. Oka, A. Nakano

      Journal of Biochemistry120 ( 2 ) 452 - 458   1 8 1996

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

      DOI: 10.1093/oxfordjournals.jbchem.a021432

      researchmap

    • The Rat Intrinsic Factor Gene: Its 5′-Upstream Region and Chief Cell-Specific Transcription1 Peer-reviewed

      Masatomo Maeda, Seikyo Asahara, Tsuyoshi Nishi, Sotaro Mushiake, Toshihiko Oka, Shoichi Shimada, Tsutomu Chiba, Masaya Tohyama, Masamitsu Futai

      The Journal of Biochemistry117 ( 6 ) 1305 - 1311   6 1995

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

      DOI: 10.1093/oxfordjournals.jbchem.a124859

      researchmap

    • Identification of the Promoter Region of the Human Histamine H2-Receptor Gene Peer-reviewed

      T. Nishi, T. Koike, T. Oka, M. Maeda, M. Futai

      Biochemical and Biophysical Research Communications210 ( 2 ) 616 - 623   5 1995

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

      DOI: 10.1006/bbrc.1995.1703

      researchmap

    • Mutational Analysis of the Sar1 Protein, a Small GTPase Which Is Essential for Vesicular Transport from the Endoplasmic Reticulum1 Peer-reviewed

      Akihiko Nakano, Hideshi Otsuka, Mami Yamagishi, Eiichi Yamamoto, Keitaro Kimura, Shuh-ichi Nishikawa, Toshihiko Oka

      The Journal of Biochemistry116 ( 2 ) 243 - 247   8 1994

      More details

      Publishing type:Research paper (scientific journal)   Publisher:Oxford University Press (OUP)  

      DOI: 10.1093/oxfordjournals.jbchem.a124513

      researchmap

    • Inhibition of GTP hydrolysis by Sar1p causes accumulation of vesicles that are a functional intermediate of the ER-to-Golgi transport in yeast Peer-reviewed

      Toshihiko Oka, Akihiko Nakano

      Journal of Cell Biology124 ( 4 ) 425 - 434   1994

      More details

      Language:English   Publishing type:Research paper (scientific journal)   Publisher:Rockefeller University Press  

      The SAR1 gene product (Sar1p), a 21-kD GTPase, is a key component of the ER-to-Golgi transport in the budding yeast. We previously reported that the in vitro reconstitution of protein transport from the ER to the Golgi was dependent on Sar1p and Sec12p (Oka, T., S. Nishikawa, and A. Nakano. 1991. J. Cell Biol. 114:671-679). Sec12p is an integral membrane protein in the ER and is essential for the Sar1 function. In this paper, we show that Sar1p can remedy the temperature-sensitive defect of the sec12 mutant membranes, which is in the formation of ER-to-Golgi transport vesicles. The addition of Sar1p promotes vesicle formation from the ER irrespective of the GTP- or GTPγS- bound form, indicating that the active form of Sar1p but not the hydrolysis of GTP is required for this process. The inhibition of GTP hydrolysis blocks transport of vesicles to the Golgi and thus causes their accumulation. The accumulating vesicles, which carry Sar1p on them, can be separated from other membranes, and, after an appropriate wash that removes Sar1p, are capable of delivering the content to the Golgi when added back to fresh membranes. Thus we have established a new method for isolation of functional intermediate vesicles in the ER-to-Golgi transport. The sec23 mutant is defective in activation of Sar1 GTPase (Yoshihisa, T., C. Barlowe, and R. Schekman. 1993. Science (Wash. DC). 259:1466-1468). The membranes and cytosol from the sec23 mutant show only a partial defect in vesicle formation and this defect is also suppressed by the increase of Sar1p. Again GTP hydrolysis is not needed for the suppression of the defect in vesicle formation. Based on these results, we propose a model in which Sar1p in the GTP-bound form is required for the formation of transport vesicles from the ER and the GTP hydrolysis by Sar1p is essential for entering the next step of vesicular transport to the Golgi apparatus.

      DOI: 10.1083/jcb.124.4.425

      Scopus

      PubMed

      researchmap

    • Reconstitution of GTP-binding Sar1 protein function in ER to Golgi transport. Peer-reviewed

      Toshihiko Oka, Shuh-ichi Nishikawa, Akihiko Nakano

      Journal of Cell Biology114 ( 4 ) 671 - 679   15 8 1991

      More details

      Authorship:Corresponding author   Publishing type:Research paper (scientific journal)   Publisher:Rockefeller University Press  

      In the yeast secretory pathway, two genes SEC12 and SAR1, which encode a 70-kD integral membrane protein and a 21-kD GTP-binding protein, respectively, cooperate in protein transport from the ER to the Golgi apparatus. In vivo, the elevation of the SAR1 dosage suppresses temperature sensitivity of the sec12 mutant. In this paper, we show cell-free reconstitution of the ER-to-Golgi transport that depends on both of these gene products. First, the membranes from the sec12 mutant cells reproduce temperature sensitivity in the in vitro ER-to-Golgi transport reaction. Furthermore, the addition of the Sar1 protein completely suppresses this temperature-sensitive defect of the sec12 membranes. The analysis of Sar1p partially purified by E. coli expression suggests that GTP hydrolysis is essential for Sar1p to execute its function.

      DOI: 10.1083/jcb.114.4.671

      researchmap

    ▼display all

    Misc.

    • Insights into the regulation of mitochondrial functions by PKA-mediated phosphorylation. Invited International journal

      Shiori Akabane, Toshihiko Oka

      Journal of biochemistry   29 9 2023

      More details

      Language:English  

      Cyclic AMP (cAMP) - protein kinase A (PKA) signaling is a highly conserved pathway in eukaryotes and plays a central role in cell signaling cascades in response to environmental changes. Elevated cAMP levels promote the activation of PKA, which phosphorylates various downstream proteins. Many cytosolic and nuclear proteins, such as metabolic enzymes and transcriptional factors, have been identified as substrates for PKA, suggesting that PKA-mediated regulation occurs predominantly in the cytosol. Mitochondrial proteins are also phosphorylated by PKA, and PKA-mediated phosphorylation of mitochondrial proteins is considered to control a variety of mitochondrial functions, including oxidative phosphorylation, protein import, morphology, and quality control. In this review, we outline PKA mitochondrial substrates and summarize the regulation of mitochondrial functions through PKA-mediated phosphorylation.

      DOI: 10.1093/jb/mvad075

      PubMed

      researchmap

    • PKAはMIC60のリン酸化を介してPINK1とParkinによるミトコンドリア品質管理を制御する Invited

      赤羽しおり, 岡 敏彦

      実験医学   2016

      More details

      Language:Japanese   Publishing type:Article, review, commentary, editorial, etc. (trade magazine, newspaper, online media)  

      researchmap

    • ミトコンドリア形態異常と疾患 Invited

      岡 敏彦

      医学のあゆみ   2015

      More details

      Language:Japanese   Publishing type:Article, review, commentary, editorial, etc. (trade magazine, newspaper, online media)  

      researchmap

    • Regulation and Physiologic Functions of GTPases in Mitochondrial Fusion and Fission in Mammals Invited

      Naotada Ishihara, Hidenori Otera, Toshihiko Oka, Katsuyoshi Mihara

      ANTIOXIDANTS & REDOX SIGNALING19 ( 4 ) 389 - 399   8 2013

      More details

      Language:English   Publishing type:Book review, literature introduction, etc.   Publisher:MARY ANN LIEBERT, INC  

      Significance: Mitochondria are double membrane-bound organelles with tubular network structures that are essential for oxidative ATP production and play pivotal roles in regulating calcium homeostasis and apoptosis. Furthermore, mitochondria produce large amounts of reactive oxygen species that are fatal to cellular functions through uncoupled respiration. These organelles dynamically change their morphology by frequent fusion and fission, and three types of high molecular weight GTPase proteins have been identified as core components of the fusion and fission machineries. Recent Advances: Here, we review recent advances in the study of mitochondrial fission and fusion GTPases and their physiologic roles in mammalian cells. The regulation of mitochondrial dynamics coupled with a quality control system is essential for cellular homeostasis, development, and tissue differentiation. Defects of these mechanisms cause various disorders, including neurodegenerative diseases, such as Parkinson's disease, Huntington's disease, and Alzheimer's disease. Critical Issues: Although a significant amount of relevant data has accumulated on the regulation of mammalian mitochondrial fusion and fission, mechanistic molecular details and cellular functions still remain insufficiently defined. Future Directions: Elucidating the physiologic roles of mitochondrial fusion and fission in highly differentiated cells using tissue-specific knockout mice remains a challenge for the future.

      DOI: 10.1089/ars.2012.4830

      researchmap

    • ミトコンドリア膜構造変化とアポトーシス制御 Invited

      大寺秀典, 岡 敏彦

      細胞工学   2010

      More details

      Language:Japanese   Publishing type:Article, review, commentary, editorial, etc. (trade magazine, newspaper, online media)  

      researchmap

    • Retrograde transport on the COG railway Invited

      D Ungar, T Oka, M Krieger, FM Hughson

      TRENDS IN CELL BIOLOGY16 ( 2 ) 113 - 120   2 2006

      More details

      Language:English   Publishing type:Book review, literature introduction, etc.   Publisher:ELSEVIER SCIENCE LONDON  

      The conserved oligomeric Golgi (COG) complex is essential for establishing and/or maintaining the structure and function of the Golgi apparatus. The Golgi apparatus, in turn, has a central role in protein sorting and glycosylation within the eukaryotic secretory pathway. As a consequence, COG mutations can give rise to human genetic diseases known as congenital disorders of glycosylation. We review recent results from studies of yeast, worm, fly and mammalian COG that provide evidence that COG might function in retrograde vesicular trafficking within the Golgi apparatus. This hypothesis explains the impact of COG mutations by postulating that they impair the retrograde flow of resident Golgi proteins needed to maintain normal Golgi structure and function.

      DOI: 10.1016/j.tcb.2005.12.004

      researchmap

    • A railroad switch in mitochondrial protein import Invited

      T Oka, K Mihara

      MOLECULAR CELL18 ( 2 ) 145 - 146   4 2005

      More details

      Language:English   Publishing type:Other   Publisher:CELL PRESS  

      Chacinska et al. (2005) recently clarified how translocation machineries of the mitochondrial outer and inner membranes cooperate to correctly sort preproteins destined for the mitochondrial matrix and inner membrane.

      DOI: 10.1016/j.molcel.2005.03.022

      researchmap

    • Multi-component protein complexes and Golgi membrane trafficking Invited

      T Oka, M Krieger

      JOURNAL OF BIOCHEMISTRY137 ( 2 ) 109 - 114   2 2005

      More details

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

      Several large cytosolic protein complexes with multiple components have been proposed to play key roles in mediating or controlling membrane trafficking. Among these complexes, TRAPP, COG and GARP/VFT have been implicated in multiple steps of Golgi membrane trafficking. The importance of these complexes for Golgi function has been established using in vitro biochemical assays and yeast and mammalian somatic cell genetics. Furthermore, mutations in the genes encoding subunits of either TRAPP or the COG complex have been shown to be responsible for human genetic disorders. We here review recent studies exploring the structures and functions of these three oligomeric complexes.

      DOI: 10.1093/jb/mvi024

      researchmap

    • Oligosaccharide specificity of galectins: a search by frontal affinity chromatography

      Jun Hirabayashi, Tomomi Hashidate, Yoichiro Arata, Nozomu Nishi, Takanori Nakamura, Mitsuomi Hirashima, Tadasu Urashima, Toshihiko Oka, Masamitsu Futai, Werner E.G Muller, Fumio Yagi, Ken-ichi Kasai

      Biochimica et Biophysica Acta (BBA) - General Subjects1572 ( 2-3 ) 232 - 254   9 2002

      More details

      Publisher:Elsevier BV  

      DOI: 10.1016/s0304-4165(02)00311-2

      researchmap

    • Luminal acidification of diverse organelles by V-ATPase in animal cells

      M. Futai, T. Oka, G. Sun-Wada, Y. Moriyama, H. Kanazawa, Y. Wada

      Journal of Experimental Biology203 ( 1 ) 107 - 116   1 1 2000

      More details

      Publisher:The Company of Biologists  

      Eukaryotic cells contain organelles bounded by a single membrane in the cytoplasm. These organelles have differentiated to carry out various functions in the pathways of endocytosis and exocytosis. Their lumina are acidic, with pH ranging from 4.5 to 6.5. This article describes recent studies on these animal cell organelles focusing on (1) the primary proton pump (vacuolar-type H(+)-ATPase) and (2) the functions of the organelle luminal acidity. We also discuss similarities and differences between vacuolar-type H(+)-ATPase and F-type ATPase. Our own studies and interests are emphasized.

      DOI: 10.1242/jeb.203.1.107

      researchmap

    • Diverse Roles of Single Membrane Organelles: Factors Establishing the Acid Lumenal pH

      M. Futai, T. Oka, Y. Moriyama, Y. Wada

      Journal of Biochemistry124 ( 2 ) 259 - 267   1 8 1998

      More details

      Publisher:Oxford University Press (OUP)  

      DOI: 10.1093/oxfordjournals.jbchem.a022105

      researchmap

    • Purification and assay of yeast Sarlp

      Keitarou Kimura, Toshihiko Oka, Akihiko Nakano

      Small GTPases and Their Regulators Part C: Proteins Involved in Transport   41 - 49   1995

      More details

      Publisher:Elsevier  

      DOI: 10.1016/s0076-6879(95)57008-x

      researchmap

    • ER-to-Golgi trabsport in yeast

      Toshihiko Oka, Akihiko Nakano

        下   615 - 623   1992

      More details

    ▼display all

    Research Projects

    • ミトコンドリア外シグナルによるミトコンドリア品質管理の制御機構

      日本学術振興会  科学研究費助成事業 

      岡 敏彦

      More details

      4 2020 - 3 2023

      Grant number:20H03257

      Grant amount:\17810000 ( Direct Cost: \13700000 、 Indirect Cost:\4110000 )

      ミトコンドリアは多彩な細胞機能を担っており,その機能は細胞により様々な調節を受けている。特に,オートファジーを介した障害を受けたミトコンドリアの排斥は,ミトコンドリア品質管理として,近年,その詳細な分子メカニズムが明らかになってきた。私達は,PINK1/Parkinを介したミトコンドリアの品質管理が,cAMP/PKA経路によるリン酸化修飾で調節されることを証明した。これにより,様々なミトコンドリア外のシグナル経路が品質管理を調節する可能性が示された。本研究では,ミトコンドリア外シグナルによるミトコンドリア品質管理調節機構の分子メカニズムの全体像の理解を目指す。令和3年度は,次の2点について研究を進めた。
      1.ミトコンドリア外膜上におけるヘキソキナーゼの機能を解析するために,3種類のアイソフォーム(HK1, HK2, HK4)を用いて,その酵素活性と細胞内局在の関連性を蛍光抗体法により解析した結果,HK1はミトコンドリアに,HK4は細胞質に,HK2はミトコンドリアと細胞質に局在することが明らかとなった。さらに,酵素活性を失ったHK2変異体は,ミトコンドリア局在を示さなかったことから,酵素活性に依存したミトコンドリアへの標的化機構が示唆された。
      2.ヘキソキナーゼアイソフォームHK2のタンパク質複合体の形成に関わる因子を検索するために,ミトコンドリアの脱分極に依存してヘキソキナーゼに結合するタンパク質の同定を試みたが,まだ特異的なタンパク室の同定には至っていない。

      researchmap

    • Phosphorylation-dependent regulation of mitochondrial functions and quality control

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      OKA Toshihiko

      More details

      4 2017 - 3 2020

      Grant number:17H03676

      Grant amount:\17420000 ( Direct Cost: \13400000 、 Indirect Cost:\4020000 )

      Mitochondria with low membrane potential are eliminated by autophagy, which is called mitochondrial quality control. It is still unclear how the mitochondrial quality control is regulated by cellular signals.
      We have focused on PKA-dependent phosphorylation and determined that intracellular localization of the PKA catalytic subunit isoforms. Furthermore, we found a role of the enzyme in the glycolytic pathway in mitochondrial quality control.

      researchmap

    • PINK1-mediated regulation of mitochondrial morphology and functions

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      Oka Toshihiko

      More details

      4 2014 - 3 2017

      Grant number:26440106

      Grant amount:\5200000 ( Direct Cost: \4000000 、 Indirect Cost:\1200000 )

      Mitochondria mainly contributes to cellular ATP production, and their morphology dynamically changes, i.e. they continuously divide and fuse each other. How mitochondrial morphology relates to their functions is watched with interest. Using nematode and mammalian cell lines, we indicated that PINK1, a key factor for mitochondrial quality control, controls mitochondrial morphology in nematode and demonstrated that PINK1activation induces necrosis-like cell death in mammalian cell lines. In particular, it is of interest in the relationship between PINK1-mediated cell death and loss of dopaminergic neurons in patients with Parkinson's disease.

      researchmap

    • The mechanism of Drp1-dependent mitochondrial fission

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      MIHARA Katsuyoshi

      More details

      4 2012 - 3 2014

      Grant number:24657138

      Grant amount:\4160000 ( Direct Cost: \3200000 、 Indirect Cost:\960000 )

      Mitochondria participate in a variety of physiologic processes such as the ATP poroduction and apoptosis. Mitochondria dynamically change morphology through fusion and fission and high MW GTPases Mfn1 and Mfn2 of outer membrane and OPA1 of inner membrane are involved in fusion, whereas cytoplasimic Drp1 regulates fission. Here, (1) Since outer membrane proteins Mff, Mid49 and Mid51 function as the Drp1 receptor, we analyzed relative importance of these receptors and found that Mff as the major Drp1 receptor. (2) We have generated Mff-KO mice; they were embryonic lethal (E6.5). Consideing that Drp1-KO mice are lethal at E12.5, Mff seems to have important developmental function in addition to the Drp1-receptor function. Furhtermore, we have demonstrated that (3) Drp1 is involved in segregation of mitochondrial nucleoids, Drp1 is esssential for making correctly arranged heart mucle fibers, and konckout of liver Drp1 induced NASH phenotype in high fat diet-fed rats by the ER stress.

      researchmap

    • Analysis of mitochondrial morphology and movement

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      OKA Toshihiko

      More details

      2011 - 2013

      Grant number:23570232

      Grant amount:\5460000 ( Direct Cost: \4200000 、 Indirect Cost:\1260000 )

      An RNA interference screen using nematode led us to identification of a novel kinasin KLP6 involved in mitochondrial morphology and movement. The dominant-negative KLP6 induced abnormal mitochondrial morphology in HeLa cells. Furthermore, anterograde transport of mitochondria in the neurites of differentiated neuro2a cells was significantly reduced when the dominant negative form was overexpressed, indicating that KLP6 regulates intracellular transport of mitochondria.
      Downregulation of an inner membrane protein LETM1 induces mitochondrial swelling and fragmentation. Moreover, cristae structure clearly disappeared in the LETM1-knockdown cells. To investigate the LETM1 functions, we attempted to in vitro reconstitute membrane invagination using LETM1 recombinant protein and artificial liposome. This is a good tool to analyze the roles of LETM1 on the formation of mitochondrial inner membrane structures.

      researchmap

    • Regulation of mitochondrial dynamics in mammals

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      MIHARA Katsuyoshi, MIHARA Hidenori, ISIHARA Naotada

      More details

      2010 - 2012

      Grant number:22370071

      Grant amount:\18850000 ( Direct Cost: \14500000 、 Indirect Cost:\4350000 )

      (1) Generation and phenotype analysis of whole body and neuron-specific Drp1-KO mice.Tissue-specific Drp1-KO mice including liver, oocyte. heart muscle, and pancreatic beta-cell have been generated and their phenotypes are now under investigation.
      (2) Identification of Mff of mitochondrial outer membrane (MOM) as the major Drp1 receptor.
      (3) Identification of a new KIF family protein KLP6 functioning as an axonal mitochondria transport motor.
      (4) Identified a Rab-GAP protein TBC1D15 as the ligand of MOM hFis1 protein involved in mitochondrial morphology regulation.
      (5) Demonstration that Drp1 depletion or inhibition of its function increases mitochondrial nucleoid size concomitant with the mitochondrial network extension and formation of tightly packed cristae structure as a response to prevent cells from apoptotic death.

      researchmap

    • ミトコンドリア分裂・融合因子の作動機序

      日本学術振興会  科学研究費助成事業 

      三原 勝芳, 岡 敏彦, 大寺 秀典

      More details

      2010 - 2011

      Grant number:22020026

      Grant amount:\6400000 ( Direct Cost: \6400000 )

      ミトコンドリア(Mt)は外膜、内膜2枚の膜に囲まれ細胞における好気的ATP産生、アポトーシス、カルシウムシグナリングなどに関わる真核細胞に必須のオルガネラであり、細胞の環境や病態に応じて融合・分裂を介してダイナミックに形態を変化させるが、この動態の調節は細胞の生と死に重要な役割を果たす。この調節には4つの高分子量GTPaseが主要な因子として関与する;細胞質に局在しMtにリクルートされてMt分裂を行うDrp1、外膜の融合に関わるMfn1とMfn2、ならびに内膜の融合とクリステ構造維持に関わる膜間部のOPA1(内膜結合)である。本申請は哺乳類Mtの融合・分裂の分子機構を明らかにすることを目的とし、平成23年度は以下の事柄を明らかにした。
      (1)Mtの分裂に関わる細胞質の因子Drp1の全身Drp1-KOならびに神経Drp1-KOマウスの作成に続いて肝臓、膵臓β細胞、心筋、卵巣でのDrp1-KOマウスを作製し解析を行っている。膵臓β細胞では2型糖尿病の、心筋では心筋梗塞の症状を示しまた卵巣では卵の分裂停止を惹起することが明らかになった(投稿準備中)。(2)Mt外膜上のDrp1受容体の実態は長らく不明であったが、我々は従来考えられていたFis1ではなく新規な外膜蛋白質MffがDrp1受容体として働くことを明らかにし現在反応機構解析を行っている。(3)Mffの高次機能を解明する目的でMff-KOマウスの作成に着手しES細胞を胚盤胞に注入しキメラマウスを作成する段階に到っている。

      researchmap

    • Analysis of mitochondrial proteins required for regulation of mitochondrial morphology

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      OKA Toshihiko

      More details

      2008 - 2010

      Grant number:20570185

      Grant amount:\4940000 ( Direct Cost: \3800000 、 Indirect Cost:\1140000 )

      To identify genes for regulation of mitochondrial morphology, we applied a feeding RNA interference in nematode C. elegans, and searched genes for mitochondrial proteins and molecular motors. We showed that MICS1 and KLP6 are required for mitochondrial morphology and transport, respectively.

      researchmap

    • Membrane morphology -An attempt to reconstitute the cristae structure in liposome-

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      OKA Toshihiko

      More details

      2008 - 2010

      Grant number:20200058

      Grant amount:\29900000 ( Direct Cost: \23000000 、 Indirect Cost:\6900000 )

      Biomembranes are various in form, and the diversity of the shapes of biomembranes are critical for cell viability. To understand the function of the shapes in intra-and inter-cellular activities, we attempted to reconstitute in vitro the cristae structure, an invaginated inner membrane of mitochondria, onto liposomes. Using a recombinant protein of LETM1, a mitochondrial protein required for maintenance of the cristae structure, multiple membrane invagination was successfully formed on liposomes.

      researchmap

    • ミトコンドリア外膜の生合成と分裂・融合の分子機構

      日本学術振興会  科学研究費助成事業 

      三原 勝芳, 岡 敏彦, 大寺 秀典

      More details

      2008 - 2010

      Grant number:20059024

      Grant amount:\7000000 ( Direct Cost: \7000000 )

      ミトコンドリア(Mt)は外膜、内膜2枚の膜に囲まれ細胞における好気的なATP産生、アポトーシス、カルシウムシグナル伝達などに関わる必須のオルガネラである。本申請は特にインターフェースとして様々な機能が集積しているMtの外膜に焦点を絞り、外膜を構成する蛋白質のMtへの配送、蛋白質輸送装置による外膜蛋白質の外膜への組み込み、ならびに外膜の分裂融合装置によるMt膜ダイナミクスの制御機構を明らかにすることを目的とし、以下の事柄を明らかにした。
      (1)ミトコンドリアDNAにコードされる蛋白質のマトリクス側から内膜への組み込みに関わる因子OXA1を同定し、この内膜組み込みの機構を明らかにした。さらに最初の膜貫通領域の直上に膜間部へのエクスポートに必須な領域があることを見いだした。(2)高分子量GTPase Drp1は通常細胞質に分布し、Mt表面にリクルートされてのMtの分裂に関わる因子であるがその生理的な重要性は不明であった。我々はDrp1遺伝子のノックアウトマウス作成に初めて成功し、個体ならびに細胞レベルでの表現形の解析を行なった。その結果Drp1は神経のシナプス形成に必須であることを明らかにした。(3)Mtの軸索輸送に関わる新規な因子KLP6の同定に成功した。
      (4) Mt外膜上のDrp1受容体は不明であったが我々は従来推定されていたFis1ではなくMffと呼ばれる外膜因子が受容体として働くことを初めて明らかにした。

      researchmap

    • ミトコンドリア外膜タンパク質のプロテアソーム分解とその生理的役割

      日本学術振興会  科学研究費助成事業 

      岡 敏彦, 大寺 秀典

      More details

      2007 - 2008

      Grant number:19044033

      Grant amount:\6100000 ( Direct Cost: \6100000 )

      1) ミトコンドリア上でのユビキチン化Tom20とその制を因子の解析
      Tom20はTom22非存在下でミトコンドリア膜上でのユビキチン化されていた。プロテアソーム阻害剤存在下で、ユビキチン化Tom20はミトコンドリア膜上に蓄積しており、膜から引き抜かれる際のどのような因子が必要かを検討したとこと、小胞体での膜タンパク質分解に関与するp97は必要でなかった。
      2) 低酸素条件下でのTom20の分解
      筋繊維ではTom20が虚血(低酸素条件下)で特異的に分解される報告がある。培養細胞においても、ガスパックにより酸素を減らすことでTom20の分解が促進されることを確認している。この分解がプロテアソーム依存であるのかを検討したが、この分解はプロテアソーム阻害剤により阻害されなかった。
      3) Tom20をユビキチン化する酵素
      これまでミトコンドリア機能に影響を与えることが知られているMarchV/MITOLや新規タンパク質MAPL/MULANなどのミトコンドリアユビキチンリガーゼは、Tom20の分解に関与しないことがこれまでの私たちの研究で明らかとなった。さらに今回、酵母Mdm30p(F-boxタンパク質)の相同遺伝子であるFbx3やCullinもまた、RNA干渉法ではTom20の分解への影響は見られなかった。

      researchmap

    • 様々な酸性環境を作り出す液胞型ATPaseの役割と機能

      日本学術振興会  科学研究費助成事業 

      岡 敏彦

      More details

      2001 - 2001

      Grant number:13680784

      Grant amount:\2200000 ( Direct Cost: \2200000 )

      researchmap

    • 線虫における液胞型ATPaseの役割とクロライドイオンによる調節

      日本学術振興会  科学研究費助成事業 

      岡 敏彦

      More details

      1999 - 2000

      Grant number:11780445

      Grant amount:\2200000 ( Direct Cost: \2200000 )

      本年度の研究行なった。具体的には以下の示す。
      1)新たな線虫V-ATPase遺伝子の同定
      本研究では、V-ATPaseの作り出す多様な酸性環境の役割を明らかにするために、複数存在するV-ATPaseサブユニットのアイソフォームに着目し、アイソフォームが酸性環境の多様性にどう影響しているかを理解しようとしている。そのために、線虫でアイソフォーム遺伝子の欠損変異体の作成、解析を行い、その構築している酸性環境を同定することを目指している。そのために、線虫のデータベースより、V-ATPaseサブユニットaに似たESTクローンを検索し、4種類のクローンを同定した。ESTクローンと5'-RACE法で得られたPCR産物の塩基配列を決定し、全長のcDNAの配列を明らかにした。これかV-ATPaseサブユニットaイソフォーム遺伝子(vha-5,vha-6,vha-7,unc-32)は、それぞれ873,865,966,894アミノ酸残基より成るタンパク質をコードしていた。
      2)線虫V-ATPase変異体の構築
      平成12年度は、新らたに同定したV-ATPaseサブユニット遺伝子)の欠損変異体の構築を目指した。まず2本鎖RNA(dsRNA)を用いたRNA interference法により一時的にvha遺伝子の発現を遮断し、目的とする欠損変異体の表現型を解析した。vha-5遺伝子のdsRNAを成虫に導入することにより、その子孫が2令幼虫期で死滅したことから、vha-5遺伝子が線虫の3令幼虫への発生に重要な役割を担っていることが示唆された。また、vha-6やunc-32遺伝子のdsRNAを導入すると、1令幼虫や胚の段階で発生が押さえられ死滅することが分かった。vha-7遺伝子のdsRNA導入では子孫に何の変化も見られなかった。これら原因としては、それぞれの発生段階で特異的なサブユニット構成のV-ATPaseの活性が必要とされていることが推定される。現在は、これら遺伝子の機能欠損による致死の原因を分子レベルで検討中である。

      researchmap

    • 胃におけるGATA転写因子の制御機構とクロライドイオン輸送の解析

      日本学術振興会  科学研究費助成事業 

      岡 敏彦

      More details

      1998 - 1999

      Grant number:09780575

      Grant amount:\1800000 ( Direct Cost: \1800000 )

      本年度の研究行なった。具体的には以下の示す。
      1、 転写囚子GATA-GTlまたはGATA-GT2と相互作用するタンパク質の検索
      融合タンパク質を用いた因子の検索:GATA-GTl、GATA-GT2と直接的に相互作用するタンパク質を調べるために、HA(ヘマグルチニン)タグを付けたGATA-GTlまたはGATA-GT2との融合タンパク質を大腸菌で大量に調製した。そのれら融合タンパク質をプローブとして、胃粘膜組織より調製したcDNAライブラリーの発現系を用いて、相互作用するタンパク質の分離を行なった。スクリーニングにより複数の陽性クローンを分離し、その塩基配列の決定した。幾つかの新規のクローンの中に、ノーザンブロットなどにより組織特異的発現が観察されるものがあった。現在は酵母Two-hybrid法滋を用いて相互作用に必要な領域を検討中である。
      2、 胃におけるクロライドイオンの輸送とそのチャンネルの同定
      胃のクロライドチャンネルを同定するために、アフリカツメガエルの卵を用いた発現クローニングを行なった。胃粘膜より調製したcDNAをアフリカツメガエルの卵に打ち込み、クロライドイオンの透過を電気生理学的に測定したが、これまでに特異的なクローンは得られていない。胃粘膜のcDNAの再調製と測定感度について再度検討しなおしたが、チャンネル活性を示すクローンは同定できなかった。

      researchmap

    • 高等植物における液胞形成の制御と組織構築

      日本学術振興会  科学研究費助成事業 

      和田 洋, 岡 敏彦, 表 弘志

      More details

      1998 - 1998

      Grant number:10182208

      Grant amount:\3000000 ( Direct Cost: \3000000 )

      液胞はその内部を酸性に保ち,液胞膜を介した化学浸透圧的エネルギー差を利用して代謝産.物・無機イオンを輸送する.高等植物の液胞膜を介した化学浸透圧エネルギー差は,H^+-ATPase,H^+-PPaseの二種のイオンポンプの機能による,本研究では、高等植物の形態・組織構築に果たす液胞の生理的機能を明らかに.する研究の一環としてオルガネラ酸性化の制御機構に焦点を結び,細胞基質と液胞のイオン環境の調節を分子レベルで解析する系の構築を進めた.シロイヌナズナの液胞膜H^+-PPaseのcDNA(AVP3)をライブラリーから取得し,これを出芽酵母の構成的かつ強発現性プロモータの下流において出芽酵母に導入した.細胞分画法・蛍光抗体法の両手法で,66-kDaのシロイヌナズナH^+-PPaseが出芽酵母の液胞に発現されることを確認した.この形質転換酵母より液胞膜小胞を単離し,ATP(出芽酵母内在性のH^+-ATPaseを駆動する),およびPPi(シロイヌナズナH^+-PPKSeを駆動する)依存に形成されるpH差.膜電位差を色素の蛍光クエンチングを指標に測定した.ATPase、PPaseの両者をもつ液胞膜小胞は,ATPもしくはPPi単独でH^+輸送を駆動したときよりも,ATP+PPiで駆動した場合の方が,大きなpH差を形成するが,一方,膜電位差は大きくは変わらない.このことは,液胞膜のイオン環境の制御には,膜電位差が重要な,おそらく制限的な因子として作用していることを示している.

      researchmap

    • マラリア原虫における細胞内小胞輸送の分子機構

      日本学術振興会  科学研究費助成事業 

      和田 洋, 岡 敏彦, 表 弘志

      More details

      1998 - 1998

      Grant number:10166204

      Grant amount:\1700000 ( Direct Cost: \1700000 )

      マラリア原虫の血球内生育サイクルにおいて,エンドサイトーシス・エクソサイトーシスといった細胞膜と細胞内小器官膜との間の小胞輸送過程が,原虫のエネルギー獲得と物質の取り込みに必須な役割を果たす.したがって,小胞輸送過程の分子機構の解明は,マラリア原虫の生育生理を理解する上で極めて重要であるのみならず,マラリア制圧に向けた新奇の戦略を構想する点においても重要である.しかしながら,本生物の細胞内小胞輸送の分子機構はほとんど明らかにされていない.細胞内小胞輸送に直接的に関与する分子として,N-ethylmaleimide sensitive fusion protcin(NSF)がPlasmodium falciparumのゲノム上に存在することを,Genomedatabaseの検索の結果,明らかにした.この分子の機能を,まず,出芽酵母発現系を用いて解析することを考え,酵母発現ベクターに遺伝子断片を組み込むことを進めている.しかしながら,Plasmodiumの遺伝子のAT-richである特性のため,通常の遺伝子操作法では特異的遺伝子断片の取得と同定が困難であり,現在のところ酵母での発現にまで至っていない.

      researchmap

    • ATP駆動型プロトントランスポーターとCI-チェンネルの構造・機能・協関

      日本学術振興会  科学研究費助成事業 

      二井 將光, 岡 敏彦

      More details

      1997 - 1997

      Grant number:09257228

      Grant amount:\1800000 ( Direct Cost: \1800000 )

      ATPの分解によって駆動されるプロトントランスポーターとクロライドチャネルについて詳細な研究を展開し、以下にまとめた主な成果を得た。
      1)F_0F_1ATPaseのプロトン・チャネル部分であるF_0部分を精製し、リポソームに埋め込み原子力間顕微鏡によって観察した。その結果、直径約130Åで中央部に深さが約18Åのくぼみがある構造を観察した。原子力間顕微鏡の限界を考慮し、この構造がF_0であることを確認した。さらにF_0を形成しているサブユニットの一つであるaサブユニットについて膜外にあると推定される四個所に対応するSite-directed抗体を作成し、F_0部分に関する反応性を検討した。これらの結果から3種のサブユニットからなるF_0部分のモデルを提出した。
      2)リソソーム、エンドソームに局在する液胞型プロトンポンプが膀胱の最表層の細胞に存在することを示した。このポンプが実際に尿を酸性化していることを示した。
      3)線虫においては3つの遺伝子(vha1,vha2,vha3)が液胞型プロトン・ポンプ(液胞型ATPase)のプロトン・チャネルをコードしていること、これらが線虫の発生に伴なってStage-Specificに、しかも細胞特異的に発現しているを示した。さらに線虫に存在するClC型のクロライドチャネルに対応するcDNAをすべてクローン化し、これらの機能を解析した。

      researchmap

    • 胃壁細胞におけるプロトンポンプの転写機構とクロライドイオン輸送の解析

      日本学術振興会  科学研究費助成事業 

      岡 敏彦

      More details

      1996 - 1996

      Grant number:08780587

      Grant amount:\1000000 ( Direct Cost: \1000000 )

      本年度の研究行なった。具体的には以下の示す。
      1、転写因子GATA-GT1またはGATA-GT2と相互作用するタンパク質の検索
      融合タンパク質を用いた因子の検索:GATA-GT1、GATA-GT2と直接的に相互作用するタンパク質を調べるために、HA(ヘマグルチニン)タグを付けたGATA-GT1またはGATA-GT2との融合タンパク質を大腸菌で大量に調製した。そのれら融合タンパク質をプローブとして、胃粘膜組織より調製したcDNAライブラリーの発現系を用いて、相互作用するタンパク質の分離を行なっている。現在までに、複数の陽性クローンを分離できた。さらに、この方法によって分離されたクローンの塩基配列の決定を進めるとともに、ノーザンブロットなどを用いて得たクローンを選別していく予定である。
      2、胃におけるクロライドイオンの輸送とそのチャンネルの同定
      胃のクロライドチャンネルを同定するために、アフリカツメガエルの卵を用いた発現クローニングを行なった。胃粘膜より調製したcDNAをアフリカツメガエルの卵に打ち込み、クロライドイオンの透過を電気生理学的に測定したが、現在までにこの方法により、特異的なクローンは得られていない。今後は、胃粘膜のcDNAの再調製と測定感度について再度検討しなおし、チャンネルを同定を進めていく予定である。

      researchmap

    • ATP駆動型プロトントランスポーターとCl^-チャンネルの構造・機能・協関

      日本学術振興会  科学研究費助成事業 

      二井 將光, 岡 敏彦

      More details

      1996 - 1996

      Grant number:08268232

      Grant amount:\1500000 ( Direct Cost: \1500000 )

      ATPによって駆動される、H^+トランスポーターについて詳細な研究を展開した。本年度は特に、液胞型ATPaseとF型ATPaseを中心に以下の成果を得た。
      (1)F型ATPaseのプロトン輸送路サブユニットの1つである、アブユニットaの膜を横切るトポロジーを決定した。また、原子間力顕微鏡(AFM)によってFoの全体的な構造を明らかにした。
      (2)液胞型ATPaseのプロトン輸送路を形成しているサブユニットの遺伝子が、線虫には3種存在していること、発現に細胞特異的性があることを示した。液胞型ATPaseと協同して、液胞系オルガネラ内部pHを調節しているCl^-チャネルに関して線虫を中心に研究を進め、遺伝子およびcDNAをクローン化し、欠失動物を作製した。
      (3)液胞型ATPaseが膀胱の最外層の細胞の形質膜に存在し、膀胱内部を酸性化していることを明らかにした。

      researchmap

    • Mechanism of H^+ transport by H^+ATPase

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      FUTAI Masamitsu, OKA Toshihiko

      More details

      1995 - 1996

      Grant number:07458159

      Grant amount:\7300000 ( Direct Cost: \7300000 )

      H^+-ATPase transport protons coupling with the energy of ATP hydrosis, and ATP synthase synthesizes ATP coupling with proton movement. They are the two of the most the important basic enzymes in organisms. In this project, we have focused on FoF_1 type ATPase and vacuolar type ATPases.
      From the extensive mutagenesis studies, we could show Lys-155, Thr-156, Glu-181 and Arg-182 of the beta subunit are forming catalytic site. We also could show that Glu-185 of the subunit is essential for the catalytic cooperativity of the enzyme. The carboxyl and amino terminal helices of the gamma subunit were shown to be essential for energy coupling between catalysis and proton transport.
      The H^+ pathway of Fo is formed from the a, b, and c subunits we could show that the a subunit has six transmembrane domains and the entire Fo has a ring like structure. We have studied effects of a series of lipophilic cations on FoF_1-ATPase and vacuolar type ATPase and chloropromazine, quinacrine could in habit both enzymes. We also showed inhibitory effects of concanamycin on vacuolar type ATPase.

      researchmap

    • プロトンポンプの胃壁細胞における特異的転写と細胞内輸送機構の解析

      日本学術振興会  科学研究費助成事業 

      岡 敏彦

      More details

      1995 - 1995

      Grant number:07780536

      Grant amount:\1000000 ( Direct Cost: \1000000 )

      転写因子GATA-GT1またはGATA-GT2と相互作用するタンパク質の検索を目的として、本年度の研究行なった。具体的には以下の示す。
      1、融合タンパク質を用いた因子の検索:GATA-GT1、GATA-GT2と直接的に相互作用するタンパク質を調べるために、HA(ヘマグルチニン)タグを付けたGATA-GT1またはGATA-GT2との融合タンパク質を大腸菌で大量に調製した。そのれら融合タンパク質をプローブとして、胃粘膜組織より調製したcDNAライブラリーの発現系を用いて、相互作用するタンパク質の分離を行なっている。現在までに、2つの陽性クローンを分離できた。さらに、この方法によるクローンの分離を進めるとともに現在得ているクローンについても詳細に解析していく予定である。
      2、酵母two-hybrid法を用いた因子の検索:GATA-GT1、GATA-GT2と直接的に相互作用するタンパク質を調べるために、酵母GAL4のDNA結合ドメインとGATA-GT1又はGATA-GT2との融合タンパク質を酵母内で発現させ、それに結合しうる因子をtwo-hybrid法により検索した。現在、胃粘膜組織より調製したcDNAライブラリーの調製が終わり、スクリーニングを開始した。今後、このスクリーニングにより得たクローンをさらに解析してゆき、GATA-GT1、GATA-GT2の転写活性化の機構に関与している因子の役割を明らかにしていく。

      researchmap

    • Mechanism of Energy Coupling between Catalysis (ATP Synthesis and Hydrolysis) and H^+ Transport in ATP Synthase

      Japan Society for the Promotion of Science  Grants-in-Aid for Scientific Research 

      FUTAI Masamitsu, OKA Toshihiko, IWAMOTO Atsuko, MORIYAMA Yoshinori

      More details

      1993 - 1994

      Grant number:05454630

      Grant amount:\6400000 ( Direct Cost: \6400000 )

      The ATP synthase (FoFl, H^+ATPase) catalyzes ATP synthesis of hydrolysis coupling with proton translocation. Escherichia coli enzyme is similar to those found in inner mitochondrial or chloroplast thylakoid membranes, and has contributed greatly to the understanding of this complicated enzyme. The ctalytic site of the enzyme is in the beta subunit or at the interface between the alpha and beta subunits of the membrane extrinsic Fl sector. The proton pathway is fomed from the a b, and c subunit of the membrane intrinsic Fo sector. The gamma, delta, and epsilon subunits of the Fl are required funcionally and structrually to connect the catalyic subunits to the Fo sector. The mechanism of ATP hydrolysis can be studied using purified (Fl-ATPase).
      In this project we defined, by mutational analysis of the E.Coli enzyme, the catalytic site in the beta subunit and the active role (s) of the gamma subunit in the energy coupling between the chemical (ATP synthesis/hydrolysis) and osmotic reaction (proton translocation).
      Results obtained for the beta subunit catalytic site were : (1) betaLys-155 and betaThr-156 in the glycine-rich sequence (Cly-Cly-Ala-Cly-Val-Cly-Lys-Thr, residues 149-156) are essential for catalysis ;
      (2) betaClu 181 and betaArg-182 in the conserved GER sequence are essential catalytic residues ;
      (3) betaCly-149 is close to betaCly-172, betaSer-174, betaClu-192 and betaVal-198. From these results, amodel of the catalytic site is proposed that is consistent with the inhibitor sensitvities of the mutant enzmes. This model is consistent with that obtained from recent X ray structure of Fl. Genetic studies suggested that the gamma subunit plays a role in regulation energy coupling (coupling between catalysis and proton transport) : gammaMet-23, gammaArg-242, and the region between gammaCln-269 and gammaVal-280 are close to each other and interract for efficient energy coupling.

      researchmap

    ▼display all