KINDAI UNIVERSITY


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MIYAMOTO Kei

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FacultyDepartment of Genetic Engineering / Graduate School of Biology-Oriented Science and Technology
PositionAssociate Professor
Degree
Commentator Guidehttps://www.kindai.ac.jp/meikan/1353-miyamoto-kei.html
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Last Updated :2020/09/30

Education and Career

Education

  •   2000 04  - 2004 03 , Kyoto University, Faculty of Agriculture

Academic & Professional Experience

  •   2020 04 ,  - 現在, Associate Professor, Faculty of Biology-Oriented Science and Technology, Kindai University
  •   2015 04 ,  - 2020 03 , Faculty of Biology-Oriented Science and Technology, Kindai University

Research Activities

Research Areas

  • Life sciences, Zoological sciences

Research Interests

  • Gene expression, Epigenetics, Nuclear transfer, Reprogramming, Molecular and developmental biology

Published Papers

  • Perturbation of maternal PIASy abundance disrupts zygotic genome activation and embryonic development via SUMOylation pathway., Higuchi C, Yamamoto M, Shin SW, Miyamoto K, Matsumoto K, Biology open, Biology open, Oct. 2019 , Refereed
  • Active Fluctuations of the Nuclear Envelope Shape the Transcriptional Dynamics in Oocytes., Almonacid M, Al Jord A, El-Hayek S, Othmani A, Coulpier F, Lemoine S, Miyamoto K, Grosse R, Klein C, Piolot T, Mailly P, Voituriez R, Genovesio A, Verlhac MH, Developmental cell, Developmental cell, Oct. 2019 , Refereed
  • Signs of biological activities of 28,000-year-old mammoth nuclei in mouse oocytes visualized by live-cell imaging., Yamagata K, Nagai K, Miyamoto H, Anzai M, Kato H, Miyamoto K, Kurosaka S, Azuma R, Kolodeznikov II, Protopopov AV, Plotnikov VV, Kobayashi H, Kawahara-Miki R, Kono T, Uchida M, Shibata Y, Handa T, Kimura H, Hosoi Y, Mitani T, Matsumoto K, Iritani A, Scientific reports, Scientific reports, 9(1), 4050, Mar. 2019 , Refereed
  • Various nuclear reprogramming systems using egg and oocyte materials., Miyamoto K, The Journal of reproduction and development, The Journal of reproduction and development, 65(3), 203 - 208, Feb. 2019 , Refereed
  • Chromatin Accessibility Impacts Transcriptional Reprogramming in Oocytes., Miyamoto K, Nguyen KT, Allen GE, Jullien J, Kumar D, Otani T, Bradshaw CR, Livesey FJ, Kellis M, Gurdon JB, Cell reports, Cell reports, 24(2), 304 - 311, Jul. 2018 , Refereed
  • Ubiquitin-proteasome system modulates zygotic genome activation in early mouse embryos and influences full-term development., Higuchi C, Shimizu N, Shin SW, Morita K, Nagai K, Anzai M, Kato H, Mitani T, Yamagata K, Hosoi Y, Miyamoto K, Matsumoto K, The Journal of reproduction and development, The Journal of reproduction and development, Dec. 2017 , Refereed
  • Maternal factors involved in nuclear reprogramming by eggs and oocytes, Kei Miyamoto, Journal of Mammalian Ova Research, Journal of Mammalian Ova Research, 30(3), 68 - 78, Oct. 2013 , Refereed
  • Transcriptional regulation and nuclear reprogramming: roles of nuclear actin and actin-binding proteins., Miyamoto K, Gurdon JB, Cellular and molecular life sciences : CMLS, Cellular and molecular life sciences : CMLS, 70(18), 3289 - 3302, Sep. 2013 , Refereed
  • Nuclear reprogramming of sperm and somatic nuclei in eggs and oocytes., Teperek M, Miyamoto K, Reproductive medicine and biology, Reproductive medicine and biology, 12(4), 133 - 149, Jun. 2013 , Refereed
  • Nuclear actin and transcriptional activation., Miyamoto K, Gurdon JB, Communicative & integrative biology, Communicative & integrative biology, 4(5), 582 - 583, Sep. 2011 , Refereed
  • Zygotic Nuclear F-Actin Safeguards Embryonic Development., Tomomi Okuno, Wayne Yang Li, Yu Hatano, Atsushi Takasu, Yuko Sakamoto, Mari Yamamoto, Zenki Ikeda, Taiki Shindo, Matthias Plessner, Kohtaro Morita, Kazuya Matsumoto, Kazuo Yamagata, Robert Grosse, Kei Miyamoto, Cell reports, Cell reports, 31(13), 107824 - 107824, Jun. 30 2020 , Refereed
    Summary:After fertilization, sperm and oocyte nuclei are rapidly remodeled to form swollen pronuclei (PN) in mammalian zygotes, and the proper formation and function of PN are key to producing totipotent zygotes. However, how mature PN are formed has been unclear. We find that filamentous actin (F-actin) assembles in the PN of mouse zygotes and is required for fully functional PN. The perturbation of nuclear actin dynamics in zygotes results in the misregulation of genes related to genome integrity and abnormal development of mouse embryos. We show that nuclear F-actin ensures DNA damage repair, thus preventing the activation of a zygotic checkpoint. Furthermore, optogenetic control of cofilin nuclear localization reveals the dynamically regulated F-actin nucleoskeleton in zygotes, and its timely disassembly is needed for developmental progression. Nuclear F-actin is a hallmark of totipotent zygotic PN, and the temporal regulation of its polymerized state is necessary for normal embryonic development.
  • The Actin-Family Protein Arp4 Is a Novel Suppressor for the Formation and Functions of Nuclear F-Actin., Shota Yamazaki, Christian Gerhold, Koji Yamamoto, Yuya Ueno, Robert Grosse, Kei Miyamoto, Masahiko Harata, Cells, Cells, 9(3), Mar. 19 2020 , Refereed
    Summary:The crosstalk between actin and actin-related proteins (Arps), namely Arp2 and Arp3, plays a central role in facilitating actin polymerization in the cytoplasm and also in the nucleus. Nuclear F-actin is required for transcriptional regulation, double-strand break repair, and nuclear organization. The formation of nuclear F-actin is highly dynamic, suggesting the involvement of positive and negative regulators for nuclear actin polymerization. While actin assembly factors for nuclear F-actin have been recently described, information about inhibitory factors is still limited. The actin-related protein Arp4 which is predominantly localized in the nucleus, has been previously identified as an integral subunit of multiple chromatin modulation complexes, where it forms a heterodimer with monomeric actin. Therefore, we tested whether Arp4 functions as a suppressor of nuclear F-actin formation. The knockdown of Arp4 (Arp4 KD) led to an increase in nuclear F-actin formation in NIH3T3 cells, and purified Arp4 potently inhibited F-actin formation in mouse nuclei transplanted into Xenopus laevis oocytes. Consistently, Arp4 KD facilitated F-actin-inducible gene expression (e.g., OCT4) and DNA damage repair. Our results suggest that Arp4 has a critical role in the formation and functions of nuclear F-actin.
  • Combinational Treatment of Trichostatin A and Vitamin C Improves the Efficiency of Cloning Mice by Somatic Cell Nuclear Transfer., Rika Azuma, Kei Miyamoto, Mami Oikawa, Masayasu Yamada, Masayuki Anzai, Journal of visualized experiments : JoVE, Journal of visualized experiments : JoVE, (134), e57036, Apr. 26 2018 , Refereed
    Summary:Somatic cell nuclear transfer (SCNT) provides a unique opportunity to directly produce a cloned animal from a donor cell, and it requires the use of skillful techniques. Additionally, the efficiencies of cloning have remained low since the successful production of cloned animals, especially mice. There have been many attempts to improve the cloning efficiency, and trichostatin A (TSA), a histone deacetylase inhibitor, has been widely used to enhance the efficiency of cloning. Here, we report a dramatically improved cloning method in mice. This somatic cell nuclear transfer method involves usage of Hemagglutinating virus of Japan Envelope (HVJ-E), which enables easy manipulation. Moreover, the treatment using two small molecules, TSA and vitamin C (VC), with deionized bovine serum albumin (dBSA), is highly effective for embryonic development. This approach requires neither additional injection nor genetic manipulation, and thus presents a simple, suitable method for practical use. This method could become a technically feasible approach for researchers to produce genetically modified animals from cultured cells. Furthermore, it might be a useful way for the rescue of endangered animals via cloning.
  • Srf destabilizes cellular identity by suppressing cell-type-specific gene expression programs., Takashi Ikeda, Takafusa Hikichi, Hisashi Miura, Hirofumi Shibata, Kanae Mitsunaga, Yosuke Yamada, Knut Woltjen, Kei Miyamoto, Ichiro Hiratani, Yasuhiro Yamada, Akitsu Hotta, Takuya Yamamoto, Keisuke Okita, Shinji Masui, Nature communications, Nature communications, 9(1), 1387 - 1387, Apr. 11 2018 , Refereed
    Summary:Multicellular organisms consist of multiple cell types. The identity of these cells is primarily maintained by cell-type-specific gene expression programs; however, mechanisms that suppress these programs are poorly defined. Here we show that serum response factor (Srf), a transcription factor that is activated by various extracellular stimuli, can repress cell-type-specific genes and promote cellular reprogramming to pluripotency. Manipulations that decrease β-actin monomer quantity result in the nuclear accumulation of Mkl1 and the activation of Srf, which downregulate cell-type-specific genes and alter the epigenetics of regulatory regions and chromatin organization. Mice overexpressing Srf exhibit various pathologies including an ulcerative colitis-like symptom and a metaplasia-like phenotype in the pancreas. Our results demonstrate an unexpected function of Srf via a mechanism by which extracellular stimuli actively destabilize cell identity and suggest Srf involvement in a wide range of diseases.
  • Peroxiredoxin as a functional endogenous antioxidant enzyme in pronuclei of mouse zygotes, Kohtaro Morita, Mikiko Tokoro, Yuki Hatanaka, Chika Higuchi, Haruka Ikegami, Kouhei Nagai, Masayuki Anzai, Hiromi Kato, Tasuku Mitani, Yoshitomo Taguchi, Kazuo Yamagata, Yoshihiko Hosoi, Kei Miyamoto, Kazuya Matsumoto, Journal of Reproduction and Development, Journal of Reproduction and Development, 64(2), 161 - 171, 2018 , Refereed
    Summary:Antioxidant mechanisms to adequately moderate levels of endogenous reactive oxygen species (ROS) are important for oocytes and embryos to obtain and maintain developmental competence, respectively. Immediately after fertilization, ROS levels in zygotes are elevated but the antioxidant mechanisms during the maternal-to-zygotic transition (MZT) are not well understood. First, we identified peroxiredoxin 1 (PRDX1) and PRDX2 by proteomics analysis as two of the most abundant endogenous antioxidant enzymes eliminating hydrogen peroxide (H2O2). We here report the cellular localization of hyperoxidized PRDX and its involvement in the antioxidant mechanisms of freshly fertilized oocytes. Treatment of zygotes at the pronuclear stage with H2O2 enhanced pronuclear localization of hyperoxidized PRDX in zygotes and concurrently impaired the generation of 5-hydroxymethylcytosine (5hmC) on the male genome, which is an epigenetic reprogramming event that occurs at the pronuclear stage. Thus, our results suggest that endogenous PRDX is involved in antioxidant mechanisms and epigenetic reprogramming during MZT.
  • A transient pool of nuclear F-actin at mitotic exit controls chromatin organization, Christian Baarlink, Matthias Plessner, Alice Sherrard, Kohtaro Morita, Shinji Misu, David Virant, Eva-Maria Kleinschnitz, Robert Harniman, Dominic Alibhai, Stefan Baumeister, Kei Miyamoto, Ulrike Endesfelder, Abderrahmane Kaidi, Robert Grosse, NATURE CELL BIOLOGY, NATURE CELL BIOLOGY, 19(12), 1389 - +, Dec. 2017 , Refereed
    Summary:Re-establishment of nuclear structure and chromatin organization after cell division is integral for genome regulation or development and is frequently altered during cancer progression. The mechanisms underlying chromatin expansion in daughter cells remain largely unclear. Here, we describe the transient formation of nuclear actin filaments (F-actin) during mitotic exit. These nuclear F-actin structures assemble in daughter cell nuclei and undergo dynamic reorganization to promote nuclear protrusions and volume expansion throughout early G1 of the cell cycle. Specific inhibition of this nuclear F-actin assembly impaired nuclear expansion and chromatin decondensation after mitosis and during early mouse embryonic development. Biochemical screening for mitotic nuclear F-actin interactors identified the actin-disassembling factor cofilin-1. Optogenetic regulation of cofilin-1 revealed its critical role for controlling timing, turnover and dynamics of F-actin assembly inside daughter cell nuclei. Our findings identify a cell-cycle-specific and spatiotemporally controlled form of nuclear F-actin that reorganizes the mammalian nucleus after mitosis.
  • Reprogramming towards totipotency is greatly facilitated by synergistic effects of small molecules, Kei Miyamoto, Yosuke Tajima, Koki Yoshida, Mami Oikawa, Rika Azuma, George E. Allen, Tomomi Tsujikawa, Tomomasa Tsukaguchi, Charles R. Bradshaw, Jerome Jullien, Kazuo Yamagata, Kazuya Matsumoto, Masayuki Anzai, Hiroshi Imai, John B. Gurdon, Masayasu Yamada, BIOLOGY OPEN, BIOLOGY OPEN, 6(4), 415 - 424, Apr. 2017 , Refereed
    Summary:Animal cloning has been achieved in many species by transplanting differentiated cell nuclei to unfertilized oocytes. However, the low efficiencies of cloning have remained an unresolved issue. Here we find that the combination of two small molecules, trichostatin A (TSA) and vitamin C (VC), under culture condition with bovine serum albumin deionized by ion-exchange resins, dramatically improves the cloning efficiency in mice and 15% of cloned embryos develop to term by means of somatic cell nuclear transfer (SCNT). The improvement was not observed by adding the non-treated, rather than deionized, bovine serum. RNA-seq analyses of SCNT embryos at the two-cell stage revealed that the treatment with TSA and VC resulted in the upregulated expression of previously identified reprogramming-resistant genes. Moreover, the expression of early-embryo-specific retroelements was upregulated by the TSA and VC treatment. The enhanced gene expression was relevant to the VC-mediated reduction of histone H3 lysine 9 methylation in SCNT embryos. Our study thus shows a simply applicable method to greatly improve mouse cloning efficiency, and furthers our understanding of how somatic nuclei acquire totipotency.
  • Nuclear Actin in Development and Transcriptional Reprogramming, Shinji Misu, Marina Takebayashi, Kei Miyamoto, FRONTIERS IN GENETICS, FRONTIERS IN GENETICS, 8, 27, Mar. 2017 , Refereed
    Summary:Actin is a highly abundant protein in eukaryotic cells and dynamically changes its polymerized states with the help of actin-binding proteins. Its critical function as a constituent of cytoskeleton has been well-documented. Growing evidence demonstrates that actin is also present in nuclei, referred to as nuclear actin, and is involved in a number of nuclear processes, including transcriptional regulation and chromatin remodeling. The contribution of nuclear actin to transcriptional regulation can be explained by its direct interaction with transcription machineries and chromatin remodeling factors and by controlling the activities of transcription factors. In both cases, polymerized states of nuclear actin affect the transcriptional outcome. Nuclear actin also plays an important role in activating strongly silenced genes in somatic cells for transcriptional reprogramming. When these nuclear functions of actin are considered, it is plausible to speculate that nuclear actin is also implicated in embryonic development, in which numerous genes need to be activated in a well-coordinated manner. In this review, we especially focus on nuclear actin's roles in transcriptional activation, reprogramming and development, including stem cell differentiation and we discuss how nuclear actin can be an important player in development and cell differentiation.
  • Gene Resistance to Transcriptional Reprogramming following Nuclear Transfer Is Directly Mediated by Multiple Chromatin-Repressive Pathways, Jerome Jullien, Munender Vodnala, Vincent Pasque, Mami Oikawa, Kei Miyamoto, George Allen, Sarah Anne David, Vincent Brochard, Stan Wang, Charles Bradshaw, Haruhiko Koseki, Vittorio Sartorelli, Nathalie Beaujean, John Gurdon, MOLECULAR CELL, MOLECULAR CELL, 65(5), 873 - +, Mar. 2017 , Refereed
    Summary:Understanding the mechanism of resistance of genes to reactivation will help improve the success of nuclear reprogramming. Using mouse embryonic fibroblast nuclei with normal or reduced DNA methylation in combination with chromatin modifiers able to erase H3K9me3, H3K27me3, and H2AK119ub1 from transplanted nuclei, we reveal the basis for resistance of genes to transcriptional reprogramming by oocyte factors. A majority of genes is affected by more than one type of treatment, suggesting that resistance can require repression through multiple epigenetic mechanisms. We classify resistant genes according to their sensitivity to 11 chromatin modifier combinations, revealing the existence of synergistic as well as adverse effects of chromatin modifiers on removal of resistance. We further demonstrate that the chromatin modifier USP21 reduces resistance through its H2AK119 deubiquitylation activity. Finally, we provide evidence that H2A ubiquitylation also contributes to resistance to transcriptional reprogramming in mouse nuclear transfer embryos.
  • Sperm is epigenetically programmed to regulate gene transcription in embryos, Marta Teperek, Angela Simeone, Vincent Gaggioli, Kei Miyamoto, George E. Allen, Serap Erkek, Taejoon Kwon, Edward M. Marcotte, Philip Zegerman, Charles R. Bradshaw, Antoine H. F. M. Peters, John B. Gurdon, Jerome Jullien, GENOME RESEARCH, GENOME RESEARCH, 26(8), 1034 - 1046, Aug. 2016 , Refereed
    Summary:For a long time, it has been assumed that the only role of sperm at fertilization is to introduce the male genome into the egg. Recently, ideas have emerged that the epigenetic state of the sperm nucleus could influence transcription in the embryo. However, conflicting reports have challenged the existence of epigenetic marks on sperm genes, and there are no functional tests supporting the role of sperm epigenetic marking on embryonic gene expression. Here, we show that sperm is epigenetically programmed to regulate embryonic gene expression. By comparing the development of sperm-and spermatid-derived frog embryos, we show that the programming of sperm for successful development relates to its ability to regulate transcription of a set of developmentally important genes. During spermatid maturation into sperm, these genes lose H3K4me2/3 and retain H3K27me3 marks. Experimental removal of these epigenetic marks at fertilization de-regulates gene expression in the resulting embryos in a paternal chromatin-dependent manner. This demonstrates that epigenetic instructions delivered by the sperm at fertilization are required for correct regulation of gene expression in the future embryos. The epigenetic mechanisms of developmental programming revealed here are likely to relate to the mechanisms involved in transgenerational transmission of acquired traits. Understanding how parental experience can influence development of the progeny has broad potential for improving human health.
  • The Expression of TALEN before Fertilization Provides a Rapid Knock-Out Phenotype in Xenopus laevis Founder Embryos, Kei Miyamoto, Ken-ichi T. Suzuki, Miyuki Suzuki, Yuto Sakane, Tetsushi Sakuma, Sarah Herberg, Angela Simeone, David Simpson, Jerome Jullien, Takashi Yamamoto, J. B. Gurdon, PLOS ONE, PLOS ONE, 10(11), e0142946, Nov. 2015 , Refereed
    Summary:Recent advances in genome editing using programmable nucleases have revolutionized gene targeting in various organisms. Successful gene knock-out has been shown in Xenopus, a widely used model organism, although a system enabling less mosaic knock-out in founder embryos (F0) needs to be explored in order to judge phenotypes in the F0 generation. Here, we injected modified highly active transcription activator-like effector nuclease (TALEN) mRNA to oocytes at the germinal vesicle (GV) stage, followed by in vitro maturation and intracytoplasmic sperm injection, to achieve a full knock-out in F0 embryos. Unlike conventional injection methods to fertilized embryos, the injection of TALEN mRNA into GV oocytes allows expression of nucleases before fertilization, enabling them to work from an earlier stage. Using this procedure, most of developed embryos showed full knock-out phenotypes of the pigmentation gene tyrosinase and/or embryonic lethal gene pax6 in the founder generation. In addition, our method permitted a large 1 kb deletion. Thus, we describe nearly complete gene knock-out phenotypes in Xenopus laevis F0 embryos. The presented method will help to accelerate the production of knock-out frogs since we can bypass an extra generation of about 1 year in Xenopus laevis. Meantime, our method provides a unique opportunity to rapidly test the developmental effects of disrupting those genes that do not permit growth to an adult able to reproduce. In addition, the protocol shown here is considerably less invasive than the previously used host transfer since our protocol does not require surgery. The experimental scheme presented is potentially applicable to other organisms such as mammals and fish to resolve common issues of mosaicism in founders.
  • Histone H3 lysine 9 trimethylation is required for suppressing the expression of an embryonically activated retrotransposon in Xenopus laevis, Sarah Herberg, Angela Simeone, Mami Oikawa, Jerome Jullien, Charles R. Bradshaw, Marta Teperek, John Gurdon, Kei Miyamoto, SCIENTIFIC REPORTS, SCIENTIFIC REPORTS, 5, 14236, Sep. 2015 , Refereed
    Summary:Transposable elements in the genome are generally silenced in differentiated somatic cells. However, increasing evidence indicates that some of them are actively transcribed in early embryos and the proper regulation of retrotransposon expression is essential for normal development. Although their developmentally regulated expression has been shown, the mechanisms controlling retrotransposon expression in early embryos are still not well understood. Here, we observe a dynamic expression pattern of retrotransposons with three out of ten examined retrotransposons (1a11, lambda-olt 2-1 and xretpos( L)) being transcribed solely during early embryonic development. We also identified a transcript that contains the long terminal repeat (LTR) of lambda-olt 2-1 and shows a similar expression pattern to lambda-olt 2-1 in early Xenopus embryos. All three retrotransposons are transcribed by RNA polymerase II. Although their expression levels decline during development, the LTRs are marked by histone H3 lysine 4 trimethylation. Furthermore, retrotransposons, especially lambda-olt 2-1, are enriched with histone H3 lysine 9 trimethylation (H3K9me3) when their expression is repressed. Overexpression of lysine-specific demethylase 4d removes H3K9me3 marks from Xenopus embryos and inhibits the repression of lambda-olt 2-1 after gastrulation. Thus, our study shows that H3K9me3 is important for silencing the developmentally regulated retrotransposon in Xenopus laevis.
  • Manipulation and In Vitro Maturation of Xenopus laevis Oocytes, Followed by Intracytoplasmic Sperm Injection, to Study Embryonic Development, Kei Miyamoto, David Simpson, John B. Gurdon, JOVE-JOURNAL OF VISUALIZED EXPERIMENTS, JOVE-JOURNAL OF VISUALIZED EXPERIMENTS, (96), e52496, Feb. 2015 , Refereed
    Summary:Amphibian eggs have been widely used to study embryonic development. Early embryonic development is driven by maternally stored factors accumulated during oogenesis. In order to study roles of such maternal factors in early embryonic development, it is desirable to manipulate their functions from the very beginning of embryonic development. Conventional ways of gene interference are achieved by injection of antisense oligonucleotides (oligos) or mRNA into fertilized eggs, enabling under-or over-expression of specific proteins, respectively. However, these methods normally require more than several hours until protein expression is affected, and, hence, the interference of gene functions is not effective during early embryonic stages. Here, we introduce an experimental system in which expression levels of maternal proteins can be altered before fertilization. Xenopus laevis oocytes obtained from ovaries are defolliculated by incubating with enzymes. Antisense oligos or mRNAs are injected into defolliculated oocytes at the germinal vesicle (GV) stage. These oocytes are in vitro matured to eggs at the metaphase II (MII) stage, followed by intracytoplasmic sperm injection (ICSI). By this way, up to 10% of ICSI embryos can reach the swimming tadpole stage, thus allowing functional tests of specific gene knockdown or overexpression. This approach can be a useful way to study roles of maternally stored factors in early embryonic development.
  • Sperm and Spermatids Contain Different Proteins and Bind Distinct Egg Factors, Marta Teperek, Kei Miyamoto, Angela Simeone, Renata Feret, Michael J. Deery, John B. Gurdon, Jerome Jullien, INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES, 15(9), 16719 - 16740, Sep. 2014 , Refereed
    Summary:Spermatozoa are more efficient at supporting normal embryonic development than spermatids, their immature, immediate precursors. This suggests that the sperm acquires the ability to support embryonic development during spermiogenesis (spermatid to sperm maturation). Here, using Xenopus laevis as a model organism, we performed 2-D Fluorescence Difference Gel Electrophoresis (2D-DIGE) and mass spectrometry analysis of differentially expressed proteins between sperm and spermatids in order to identify factors that could be responsible for the efficiency of the sperm to support embryonic development. Furthermore, benefiting from the availability of egg extracts in Xenopus, we also tested whether the chromatin of sperm could attract different egg factors compared to the chromatin of spermatids. Our analysis identified: (1) several proteins which were present exclusively in sperm; but not in spermatid nuclei and (2) numerous egg proteins binding to the sperm (but not to the spermatid chromatin) after incubation in egg extracts. Amongst these factors we identified many chromatin-associated proteins and transcriptional repressors. Presence of transcriptional repressors binding specifically to sperm chromatin could suggest its preparation for the early embryonic cell cycles, during which no transcription is observed and suggests that sperm chromatin has a unique protein composition, which facilitates the recruitment of egg chromatin remodelling factors. It is therefore likely that the acquisition of these sperm-specific factors during spermiogenesis makes the sperm chromatin suitable to interact with the maternal factors and, as a consequence, to support efficient embryonic development.
  • Hierarchical Molecular Events Driven by Oocyte-Specific Factors Lead to Rapid and Extensive Reprogramming, Jerome Jullien, Kei Miyamoto, Vincent Pasque, George E. Allen, Charles R. Bradshaw, Nigel J. Garrett, Richard P. Halley-Stott, Hiroshi Kimura, Keita Ohsumi, John B. Gurdon, MOLECULAR CELL, MOLECULAR CELL, 55(4), 524 - 536, Aug. 2014 , Refereed
    Summary:Nuclear transfer to oocytes is an efficient way to transcriptionally reprogram somatic nuclei, but its mechanisms remain unclear. Here, we identify a sequence of molecular events that leads to rapid transcriptional reprogramming of somatic nuclei after transplantation to Xenopus oocytes. RNA-seq analyses reveal that reprogramming by oocytes results in a selective switch in transcription toward an oocyte rather than pluripotent type, without requiring new protein synthesis. Time-course analyses at the single-nucleus level show that transcriptional reprogramming is induced in most transplanted nuclei in a highly hierarchical manner. We demonstrate that an extensive exchange of somatic-for oocyte-specific factors mediates reprogramming and leads to robust oocyte RNA polymerase II binding and phosphorylation on transplanted chromatin. Moreover, genome-wide binding of oocyte-specific linker histone B4 supports its role in transcriptional reprogramming. Thus, our study reveals the rapid, abundant, and stepwise loading of oocyte-specific factors onto somatic chromatin as important determinants for successful reprogramming.
  • Nuclear Wave1 Is Required for Reprogramming Transcription in Oocytes and for Normal Development, Kei Miyamoto, Marta Teperek, Kosuke Yusa, George E. Allen, Charles R. Bradshaw, J. B. Gurdon, SCIENCE, SCIENCE, 341(6149), 1002 - 1005, Aug. 2013 , Refereed
    Summary:Eggs and oocytes have a remarkable ability to induce transcription of sperm after normal fertilization and in somatic nuclei after somatic cell nuclear transfer. This ability of eggs and oocytes is essential for normal development. Nuclear actin and actin-binding proteins have been shown to contribute to transcription, although their mode of action is elusive. Here, we find that Xenopus Wave1, previously characterized as a protein involved in actin cytoskeleton organization, is present in the oocyte nucleus and is required for efficient transcriptional reprogramming. Moreover, Wave1 knockdown in embryos results in abnormal development and defective hox gene activation. Nuclear Wave1 binds by its WHD domain to active transcription components, and this binding contributes to the action of RNA polymerase II. We identify Wave1 as a maternal reprogramming factor that also has a necessary role in gene activation in development.
  • Reprogramming and development in nuclear transfer embryos and in interspecific systems, Patrick Narbonne, Kei Miyamoto, J. B. Gurdon, CURRENT OPINION IN GENETICS & DEVELOPMENT, CURRENT OPINION IN GENETICS & DEVELOPMENT, 22(5), 450 - 458, Oct. 2012 , Refereed
    Summary:Nuclear transfer (NT) remains the most effective method to reprogram somatic cells to totipotency. Somatic cell nuclear transfer (SCNT) efficiency however remains low, but recurrent problems occurring in partially reprogrammed cloned embryos have recently been identified and some remedied. In particular, the trophectoderm has been identified as a lineage whose reprogramming success has a large influence on SCNT embryo development. Several interspecific hybrid and cybrid reprogramming systems have been developed as they offer various technical advantages and potential applications, and together with SCNT, they have led to the identification of a series of reprogramming events and responsible reprogramming factors. Interspecific incompatibilities hinder full exploitation of cross-species reprogramming systems, yet recent findings suggest that these may not constitute insurmountable obstacles.
  • Epigenetic factors influencing resistance to nuclear reprogramming, Vincent Pasque, Jerome Jullien, Kei Miyamoto, Richard P. Halley-Stott, J. B. Gurdon, TRENDS IN GENETICS, TRENDS IN GENETICS, 27(12), 516 - 525, Dec. 2011 , Refereed
    Summary:Patient-specific somatic cell reprogramming is likely to have a large impact on medicine by providing a source of cells for disease modelling and regenerative medicine. Several strategies can be used to reprogram cells, yet they are generally characterised by a low reprogramming efficiency, reflecting the remarkable stability of the differentiated state. Transcription factors, chromatin modifications, and noncoding RNAs can increase the efficiency of reprogramming. However, the success of nuclear reprogramming is limited by epigenetic mechanisms that stabilise the state of gene expression in somatic cells and thereby resist efficient reprogramming. We review here the factors that influence reprogramming efficiency, especially those that restrict the natural reprogramming mechanisms of eggs and oocytes. We see this as a step towards understanding the mechanisms by which nuclear reprogramming takes place.
  • Nuclear actin in transcriptional reprogramming by oocytes Are actin nucleators key players?, Kei Miyamoto, Vincent Pasque, John B. Gurdon, CELL CYCLE, CELL CYCLE, 10(18), 3040 - 3041, Sep. 2011 , Refereed
  • Mechanisms of nuclear reprogramming by eggs and oocytes: a deterministic process?, Jerome Jullien, Vincent Pasque, Richard P. Halley-Stott, Kei Miyamoto, J. B. Gurdon, NATURE REVIEWS MOLECULAR CELL BIOLOGY, NATURE REVIEWS MOLECULAR CELL BIOLOGY, 12(7), 453 - 459, Jul. 2011 , Refereed
    Summary:Differentiated cells can be experimentally reprogrammed back to pluripotency by nuclear transfer, cell fusion or induced pluripotent stem cell technology. Nuclear transfer and cell fusion can lead to efficient reprogramming of gene expression. The egg and oocyte reprogramming process includes the exchange of somatic proteins for oocyte proteins, the post-translational modification of histones and the demethylation of DNA. These events occur in an ordered manner and on a defined timescale, indicating that reprogramming by nuclear transfer and by cell fusion rely on deterministic processes.
  • Nuclear actin polymerization is required for transcriptional reprogramming of Oct4 by oocytes, Kei Miyamoto, Vincent Pasque, Jerome Jullien, John B. Gurdon, GENES & DEVELOPMENT, GENES & DEVELOPMENT, 25(9), 946 - 958, May 2011 , Refereed
    Summary:Amphibian oocytes can rapidly and efficiently reprogram the transcription of transplanted somatic nuclei. To explore the factors and mechanisms involved, we focused on nuclear actin, an especially abundant component of the oocyte's nucleus (the germinal vesicle). The existence and significance of nuclear actin has long been debated. Here, we found that nuclear actin polymerization plays an essential part in the transcriptional reactivation of the pluripotency gene Oct4 (also known as Pou5f1). We also found that an actin signaling protein, Toca-1, enhances Oct4 reactivation by regulating nuclear actin polymerization. Toca-1 overexpression has an effect on the chromatin state of transplanted nuclei, including the enhanced binding of nuclear actin to gene regulatory regions. This is the first report showing that naturally stored actin in an oocyte nucleus helps transcriptional reprogramming in a polymerization-dependent manner.
  • Identification and characterization of an oocyte factor required for development of porcine nuclear transfer embryos, Kei Miyamoto, Kouhei Nagai, Naoya Kitamura, Tomoaki Nishikawa, Haruka Ikegami, Nguyen T. Binh, Satoshi Tsukamoto, Mai Matsumoto, Tomoyuki Tsukiyama, Naojiro Minami, Masayasu Yamada, Hiroyoshi Ariga, Masashi Miyake, Tatsuo Kawarasaki, Kazuya Matsumoto, Hiroshi Imai, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 108(17), 7040 - 7045, Apr. 2011 , Refereed
    Summary:Nuclear reprogramming of differentiated cells can be induced by oocyte factors. Despite numerous attempts, these factors and mechanisms responsible for successful reprogramming remain elusive. Here, we identify one such factor, necessary for the development of nuclear transfer embryos, using porcine oocyte extracts in which some reprogramming events are recapitulated. After incubating somatic nuclei in oocyte extracts from the metaphase II stage, the oocyte proteins that were specifically and abundantly incorporated into the nuclei were identified by mass spectrometry. Among 25 identified proteins, we especially focused on a multifunctional protein, DJ-1. DJ-1 is present at a high concentration in oocytes from the germinal vesicle stage until embryos at the four-cell stage. Inhibition of DJ-1 function compromises the development of nuclear transfer embryos but not that of fertilized embryos. Microarray analysis of nuclear transfer embryos in which DJ-1 function is inhibited shows perturbed expression of P53 pathway components. In addition, embryonic arrest of nuclear transfer embryos injected with anti-DJ-1 antibody is rescued by P53 inhibition. We conclude that DJ-1 is an oocyte factor that is required for development of nuclear transfer embryos. This study presents a means for identifying natural reprogramming factors in mammalian oocytes and a unique insight into the mechanisms underlying reprogramming by nuclear transfer.
  • Mammalian nuclear transplantation to Germinal Vesicle stage Xenopus oocytes - A method for quantitative transcriptional reprogramming, R. P. Halley-Stott, V. Pasque, C. Astrand, K. Miyamoto, I. Simeoni, J. Jullien, J. B. Gurdon, METHODS, METHODS, 51(1), 56 - 65, May 2010 , Refereed
    Summary:Full-grown Xenopus oocytes in first meiotic prophase contain an immensely enlarged nucleus, the Germinal Vesicle (GV), that can be injected with several hundred somatic cell nuclei. When the nuclei of mammalian somatic cells or cultured cell lines are injected into a GV, a wide range of genes that are not transcribed in the donor cells, including pluripotency genes, start to be transcriptionally activated, and synthesize primary transcripts continuously for several days. Because of the large size and abundance of Xenopus laevis oocytes, this experimental system offers an opportunity to understand the mechanisms by which somatic cell nuclei can be reprogrammed to transcribe genes characteristic of oocytes and early embryos. The use of mammalian nuclei ensures that there is no background of endogenous maternal transcripts of the kind that are induced. The induced gene transcription takes place in the absence of cell division or DNA synthesis and does not require protein synthesis. Here we summarize new as well as established results that characterize this experimental system. In particular, we describe optimal conditions for transplanting somatic nuclei to oocytes and for the efficient activation of transcription by transplanted nuclei. We make a quantitative determination of transcript numbers for pluripotency and housekeeping genes, comparing cultured somatic cell nuclei with those of embryonic stem cells. Surprisingly we find that the transcriptional activation of somatic nuclei differs substantially from one donor cell-type to another and in respect of different pluripotency genes. We also determine the efficiency of an injected mRNA translation into protein. (C) 2010 Elsevier Inc. All rights reserved.
  • Cell-Free Extracts from Mammalian Oocytes Partially Induce Nuclear Reprogramming in Somatic Cells, Kei Miyamoto, Tomoyuki Tsukiyama, Yang Yang, Ning Li, Naojiro Minami, Masayasu Yamada, Hiroshi Imai, BIOLOGY OF REPRODUCTION, BIOLOGY OF REPRODUCTION, 80(5), 935 - 943, May 2009 , Refereed
    Summary:Nuclear transfer has been regarded as the only reliable tool for studying nuclear reprogramming of mammalian somatic cells by oocytes. However, nuclear transfer is not well suited for biochemical analyses of the molecular mechanisms of reprogramming. A cell-free system from oocytes is an attractive alternative way to mimic reprogramming in vitro, since a large number of cells can be treated and analyzed. Nevertheless, a cell-free system using oocytes has not been developed in mammals. Here, cell extracts from porcine oocytes were prepared and their ability to induce nuclear reprogramming was evaluated. Extracts from metaphase II (MII) oocytes erased the machinery for regulating gene expression in reversibly permeabilized somatic cells. For example, the extracts caused histone deacetylation and the disappearance of TATA box-binding protein from the nuclei. However, MII-extract-treated cells did not show any obvious changes after cell culture. In contrast, extracts from germinal vesicle (GV) oocytes activated pluripotent marker genes, especially NANOG, and induced partial dedifferentiation after cell culture. The activation of pluripotent marker genes by GV extracts was associated with histone acetylation that was induced during extract treatment. These results indicate that GV- and MII-oocyte extracts have different roles on nuclear reprogramming. Furthermore, both oocyte extracts induced site-specific demethylation in the upstream region of NANOG. These results indicate that cell-free extracts derived from GV- and MII-oocytes could be useful for studying the mechanisms involved in nuclear reprogramming.
  • Reversible Membrane Permeabilization of Mammalian Cells Treated with Digitonin and Its Use for Inducing Nuclear Reprogramming by Xenopus Egg Extracts, Kei Miyamoto, Teruyoshi Yamashita, Tomoyuki Tsukiyama, Naoya Kitamura, Naojiro Minami, Masayasu Yamada, Hiroshi Imai, CLONING AND STEM CELLS, CLONING AND STEM CELLS, 10(4), 535 - 542, Dec. 2008 , Refereed
    Summary:Plasma membranes can be reversibly permeabilized by Streptolysin O. The permeabilized cells can be reprogrammed and partially dedifferentiated in the cell-free system from egg extracts. However, the permeabilizing activity of Streptolysin 0 is not stable, and therefore it is difficult to control its activity. An alternative method for reversible permeabilization is useful for establishing a cell-free system. Here, we used a nonionic detergent, digitonin, for permeabilization. A low concentration of digitonin induced reversible permeabilization of the plasma membrane in bovine, mouse, and porcine somatic cells. The permeabilized cells were treated with Xenoptis laevis egg extracts. The treated cells showed exchange of nuclear proteins from extracts such as incorporation of Xenopus-specific histone B4 and Lamin LIII into nuclei. After resealing of the membrane, the cells showed upregulation of OCT4, SOX2, and NANOG expression. Our results suggest that reversible permeabilization with digitonin can be used to induce nuclear reprogramming and to activate pluripotent genes by a cell-free system.
  • Reprogramming events of mammalian somatic cells induced by Xenopus laevis egg extracts, Kei Miyamoto, Tadashi Furusawa, Mari Ohnuki, Sandeep Goel, Tomoyuki Tokunaga, Naojiro Minami, Masayasu Yamada, Keita Ohsumi, Hiroshi Imai, MOLECULAR REPRODUCTION AND DEVELOPMENT, MOLECULAR REPRODUCTION AND DEVELOPMENT, 74(10), 1268 - 1277, Oct. 2007 , Refereed
    Summary:It is known that differentiated cells can be reprogrammed to an undifferentiated state in oocyte cytoplasm after nuclear transfer. Recently, some reports suggested that Xenopus egg extracts have the ability to reprogram mammalian somatic cells. Reprogramming events of mammalian cells after Xenopus egg extract treatment and after cell culture of extract-treated cells have not been elucidated. In this experiment, we examined reprogramming events in reversibly permeabilized or nonpermeabilized porcine fibroblast cells after Xenopus egg extract treatment. The Xenopus egg-specific histone 134 was assembled on porcine chromatin and nuclear lamin LIII was incorporated into nuclei. Deacetylation of histone H3 at lysine 9 in extract-treated cells was detected in nonpermeabilized cells, suggesting that a part of reprogramming may be induced even in nonpermeabilized cells. Following culture of extract-treated cells, the cells began to express the pluripotent marker genes such as POU5F1 (OCT4) and SOX2 and to form colonies. Reactivation of the OCT4 gene in extract-treated cells was also confirmed in bovine fibroblasts transformed with an OCT4-EGFP construct. These results suggest that nuclei of mammalian cells can be partially reprogrammed to an embryonic state by Xenopus egg extracts and the remodeled cells partly dedifferentiate after cell culture. A system using egg extracts may be useful for understanding the mechanisms and processes of dedifferentiation and reprogramming of mammalian somatic cells after nuclear transfer.
  • Effects of synchronization of donor cell cycle on embryonic development and DNA synthesis in porcine nuclear transfer embryos, Kei Miyamoto, Yoichiro Hoshino, Naojiro Minami, Masayasu Yamada, Hiroshi Imai, JOURNAL OF REPRODUCTION AND DEVELOPMENT, JOURNAL OF REPRODUCTION AND DEVELOPMENT, 53(2), 237 - 246, Apr. 2007 , Refereed
    Summary:The relationship between donor cell cycle and the developmental ability of somatic cell nuclear transfer (SCNT) embryos has not fully been elucidated. Donor cells that are usually prepared by serum starvation or confluent-cell culture for SCNT represent a heterogeneous population that includes mainly G0 phase cells, other cells in different phases of the cell cycle and apoptotic cells. In this study, we compared the developmental ability of porcine SCNT embryos reconstructed from G0 phase cells (GO-SCNT embryos) and strictly synchronized-G1 phase cells (G1-SCNT embryos), and examined the developmental rates and timing of first DNA synthesis. The G0 phase cells were synchronized by confluent culture, and the G1 phase cells were prepared from actively dividing M phase cells. The G1-SCNT embryos showed a significantly higher (P<0.05) developmental rate to the blastocyst stage per cleaved embryo (59%) than the G0-SCNT embryos (43%). Moreover, initiation of first DNA synthesis and cleavage occurred significantly earlier in the G1-SCNT embryos than in the G0-SCNT embryos. Delay of initiation of first DNA synthesis in the SCNT embryos by aphidicolin resulted in decreased developmental rates to the blastocyst stage without any effect on cleavage rates. Our data demonstrates that synchronized-G1 phase cells can be used as donor cells for SCNT embryos and that earlier initiation of first DNA synthesis may be important for subsequent development of SCNT embryos. The SCNT system using G1-synchronized cells, in terms of their highly uniform and viable cell states, can be useful for studying the reprogramming processes and embryonic development of SCNT embryos.

Conference Activities & Talks

  • Functional analysis of genes responsible for acquiring totipotency by somatic cell nuclear transfer,   2019 11 21
  • Transcriptional reprogramming induced by a novel nuclear transfer system that does not require cell division and DNA replication,   2019 11 21
  • Proteomic analysis of muscle and bone marrow tissues obtained from a 28,000- year-old woolly mammoth,   2019 07 26
  • Comprehensive analysis of post-translational modifications on the proteins from a 28,000-year-old woolly mammoth,   2019 07 26
  • Actin polymerization in reprogramming nuclear structures, Kei Miyamoto, ASCB/EMBO 2018meeting,   2018 12 08 , 招待有り
  • Roles of actin family proteins in chromatin and nuclear functions, International Symposium on 3R and 3C,   2018 11 13
  • Efficient nuclear reprogramming of somatic cells towards totipotency is supported by synergistic effects of small molecules, K. Miyamoto, Y. Tajima, K. Yoshida, M. Oikawa, R. Azuma, M. Mori, Y. Imasato, G.E. Allen, T. Tsujikawa, T. Tsukaguchi, C.R. Bradshaw, J. Jullien, K. Yamagata, K. Matsumoto, M. Anzai, H. Imai, J.B. Gurdon, M. Yamada, The 50th annual meeting of Japan Society of Developmental Biologists,   2017 05 10 , 招待有り
  • Nuclear actin in the regulation of transcription and nuclear structure, Kei Miyamoto, Human Frontier Science Program Kick-off Symposium,   2017 01 26 , 招待有り
  • Mechanisms of nuclear reprogramming in eggs and oocytes, Kei Miyamoto, King Abdullah University of Science and Technology,   2016 11 30 , 招待有り
  • Mechanisms of nuclear reprogramming in eggs and oocytes, Kei Miyamoto, New York University Abu Dhabi,   2016 11 28 , 招待有り
  • The improved culture condition for mouse nuclear transfer embryos enables highly efficient nuclear reprogramming, K. Miyamoto, Y. Tajima, K. Yoshida, T. Tsukaguchi, C.R. Bradshaw, G.E. Allen, M. Mori, Y. Imazato, J. Jullien, K. Matsumoto, H. Imai, J.B. Gurdon, M. Yamada, The 3rd China-Japan-Korea reproduction meeting,   2016 08 19 , 招待有り
  • Reprogramming of somatic nuclei towards totipotency is greatly facilitated by small molecules through epigenetic alterations, K. Miyamoto, Y. Tajima, K. Yoshida, M. Oikawa, T. Tsukaguchi, C.R. Bradshaw, G.E. Allen, J. Jullien, K. Matsumoto, H. Imai, J.B. Gurdon, M. Yamada, International Symposium on Epigenome Dynamics and Regulation in Germ Cells,   2016 02 17 , 招待有り
  • Genome-wide analysis of transcription-associated open chromatin regions during embryonic development and nuclear reprogramming, K. Miyamoto, G.E. Allen, C.R. Bradshaw, J.B. Gurdon, International Symposium on Chromatin Structure, Dynamics and Function,   2015 08 24
  • Molecular mechanisms how mouse somatic nuclei acquire totipotency, Kei Miyamoto,   2015 08 18 , 招待有り
  • Genome-wide analysis of transcription-associated open chromatin regions during early embryonic development and nuclear reprogramming, Kei Miyamoto,   2015 07 15 , 招待有り
  • Genome-wide analysis of open chromatin regions during early embryonic development and nuclear reprogramming, K. Miyamoto, G.E. Allen, C.R. Bradshaw, J.B. Gurdon,   2015 04 26 , 招待有り
  • Mechanisms of nuclear reprogramming in eggs and oocytes, Kei Miyamoto, The 78th of Stem Cell Biology and Regenerative Medicine Forum,,   2015 03 04 , 招待有り
  • Genome-wide analysis of open chromatin regions during early embryonic development and nuclear reprogramming, K. Miyamoto, G.E. Allen, C.R. Bradshaw, M. Teperek, J.B. Gurdon, The 1st Gurdon Institute Post-doc Association symposium,   2014 11 27
  • Mechanisms of transcriptional reprogramming in oocytes and eggs; nuclear actin and actin-binding proteins as key players, Kei Miyamoto,   2014 11 06 , 招待有り
  • Mechanisms of nuclear reprogramming by eggs and oocytes, Kei Miyamoto,   2014 10 15 , 招待有り
  • Mechanisms of nuclear reprogramming by eggs and oocytes, Kei Miyamoto,   2014 10 10 , 招待有り
  • Transcriptional Reprogramming of Sperm and Somatic Nuclei in Xenopus Laevis Oocytes and Eggs, Kei Miyamoto,   2014 09 24 , 招待有り
  • Transcriptional reprogramming of mammalian nuclei by maternal factors, Kei Miyamoto,   2014 04 07 , 招待有り
  • Transcriptional reprogramming of mammalian nuclei by maternal factors, Kei Miyamoto,   2014 02 24 , 招待有り
  • Transcriptional reprogramming of mammalian nuclei by maternal factors, Kei Miyamoto, Hiroshima University,   2013 12 12 , 招待有り
  • Transcriptional reprogramming of mammalian nuclei by maternal factors, Kei Miyamoto, Nagoya University,   2013 12 09 , 招待有り
  • Transcriptional reprogramming of mammalian nuclei by maternal factors, Kei Miyamoto,   2013 12 07 , 招待有り
  • Nuclear actin and actin-binding proteins in nuclear reprogramming and embryonic development: their important roles in transcriptional regulation, Kei Miyamoto, The 36th Annual of the Molecular Biology Society of Japan,   2013 12 05 , 招待有り
  • Mechanisms of nuclear reprogramming by eggs and oocytes, Kei Miyamoto, RIKEN,   2012 12 28 , 招待有り
  • The mechanisms of nuclear reprogramming by eggs and oocytes, Kei Miyamoto,   2012 06 11 , 招待有り
  • Nuclear WAVE1 is necessary for transcriptional reprogramming by Xenopus eggs and oocytes, K. Miyamoto, J.B. Gurdon, International Society for Stem Cell Research (ISSCR) Meeting,   2012 06
  • Mechanisms of nuclear reprogramming by eggs and oocytes, Kei Miyamoto,   2012 06 , 招待有り
  • WAVE1 is required for transcriptional reprogramming by Xenopus eggs and oocytes, K. Miyamoto, J.B. Gurdon, British Society for Cell Biology (BSCB), British Society for Developmental Biology (BSDB) and Japanese Society for Developmental Biologists (JSDB) Joint Spring Meeting,   2012 04
  • Mechanisms of nuclear reprogramming by eggs and oocytes, Kei Miyamoto, Tohoku University,   2011 12 26 , 招待有り
  • Nuclear actin in transcriptional reprogramming by Xenopus laevis oocytes, Kei Miyamoto, The WGF symposium “Actin and actin-associated proteins from gene to polysomes”,   2011 09 , 招待有り
  • Nuclear actin polymerization is required for transcriptional reprogramming of Oct4 by oocytes, K. Miyamoto, V. Pasque, J. Jullien, J.B. Gurdon, 1st annual Cambridge Stem Cell international symposium: Pluripotency and Development,   2011 07
  • Identification of reprogramming factors in oocytes by a novel screening method, K. Miyamoto, J.B. Gurdon, BSCB and BSDB Joint Spring Meeting,   2010 04 13
  • Establishment of novel systems for unraveling reprogramming, Kei Miyamoto, Kinki University,   2010 02 , 招待有り
  • Reprogramming in the oocyte cell-free system, Kei Miyamoto, Kinki University,   2009 04 03 , 招待有り
  • Mammalian oocyte extracts induce chromatin remodeling and dedifferentiation of somatic cells, but do not global demethylation, K. Miyamoto, T. Tsukiyama, N. Minami, M. Yamada, H. Imai, International Congress on Animal Reproduction (ICAR),   2008 07 14
  • Reprogramming of somatic cells in cell-free extracts from mammalian oocytes and identification of reprogramming-related proteins, Kei Miyamoto, New Bolton Center; University of Pennsylvania,   2008 06 17 , 招待有り
  • Reversible permeabilization of mammalian somatic cells treated with digitonin for reprogramming and dedifferentiation by Xenopus cell-free system, K. Miyamoto, T. Yamashita, N. Minami, M. Yamada, H. Imai, ISSCR,   2008 06 11
  • DNA demethylation associated with nuclear reprogramming of the somatic cell genome in cell-free extracts from mammalian oocytes, K. Miyamoto, T. Tsukiyama, Y. Yang, N. Li, N. Minami, M. Yamada, H. Imai, Society for the Study of Reproduction (SSR),   2008 05 27
  • Differential nuclear reprogramming of somatic cells by porcine germinal vesicle and metaphase II oocyte extracts, K. Miyamoto, T. Tsukiyama, N. Minami, M. Yamada, H. Imai, Asian Reproductive Biotechnology Society (ARBS),   2007 11 25
  • Nuclear reprogramming of porcine cells and their use as donor cell for nuclear transfer after treatment in Xenopus egg extracts, K. Miyamoto, M. Ohnuki, N. Minami, M. Yamada, H. Imai, The International Embryo Transfer Society (IETS) Meeting,   2007 01 07
  • Reprogramming of intact and permeabilized mammalian somatic cells by Xenopus egg extract, K. Miyamoto, M. Ohnuki, N. Minami, M. Yamada, H. Imai, ARBS,   2006 11 30
  • Nuclear reprogramming of porcine fibroblast cells by Xenopus egg extracts, K. Miyamoto, Y. Nagao, N. Minami, M. Yamada, K. Ohsumi, H. Imai, IETS Meeting,   2006 01 08
  • Imai. Reprogramming events of porcine fibroblast cells in Xenopus egg extracts, K. Miyamoto, T. Furusawa, T. Tokunaga, Y. Nagao, N. Minami, M. Yamada, K. Ohsumi, H. Imai, International Symposium on Germ Cells, Epigenetics, Reprogramming and Embryonic Stem Cells,   2005 11 18
  • Cell cycle synchronization of donor cells at G1 phase and developmental ability of nuclear transfer embryos in miniature pigs, K. Miyamoto, Y. Hoshino, Y. Nagao, N. Minami, M. Yamada, H. Imai, IETS Meeting,   2005

Misc

  • A nuclear role for actin polymerization in early embryonic development,   2020 08
  • 核内アクチンタンパク質の重合化がマウス初期胚の細胞分裂に及ぼす影響, 坂本裕子, 奥野智美, LI Yang, 山本真理, 神谷拓磨, 越智浩介, 井橋俊哉, 辻本佳加理, 松橋珠子, 松本和也, GROSSE Robert, 宮本圭, Journal of Mammalian Ova Research, 36, 1, S49,   2019 04 22 , https://jglobal.jst.go.jp/detail?JGLOBAL_ID=201902228819978023
  • Screening and functional analysis of genes responsible for acquiring totipotency in somatic cell nuclear transfer embryos, IHASHI Shunya, OCHI Kousuke, SAKAMOTO Yuko, TSUJIMOTO Kagari, KASAHARA Yoshito, MATSUHASHI Tamako, MATSUMOTO Kazuya, MIYAMOTO Kei, MORI Miki, IMASATO Yuma, KUSAKABE Haruna, KAJIKURI Naoaki, MATSUZAWA Yuka, KAMIYA Takuma, OKUNO Tomomi, YAMAMOTO Mari, The Journal of Reproduction and Development Supplement, 112, 0, P, 98-P-98,   2019 , http://ci.nii.ac.jp/naid/130007719383
    Summary:<p>雌性生殖細胞である卵子は精子と受精することにより分化全能性を獲得する。一方,除核した卵子に体細胞核を移植すると,体細胞核が卵子内因子の働きにより初期化され核移植胚が発生を開始する。しかし,体細胞核移植胚の発生率は依然として低い。また,核移植胚の低発生率に関与する特定の遺伝子は明らかになっていない。我々の研究グループは近年,マウス核移植胚をTricostatin A(TSA)で処理後に連続してVitamin C(VC)添加培地で一定時間培養することで,TSA単独の発生率よりも4倍近く高い発生能を示し,約15%の核移植胚が産仔にまで発生することを発見した。そこで本研究では,核移植胚の発生率が大幅に向上する条件下で発現変動を示す遺伝子群に着目し,核移植胚の発生に関与する特定の遺伝子の同定を目指す。まず,RNAシークエンシングを用いてTSAとVCで処理した核移植胚と通常核移植胚のトランスクリプトームを比較し,2細胞期胚において通常核移植胚より発現が上昇している遺伝子141個を同定した。それら141個の遺伝子の内,受精卵と比較して通常の核移植胚で発現の低下する遺伝子を16個にまで絞り込んだ。これらの遺伝子は,核移植胚の発生率が向上した際に発現上昇するが,通常の核移植胚では受精卵と比較して発現抑制されている。同定した16個の遺伝子の胚発生における役割を調べるため,マウス受精卵へsiRNAインジェクションを行い,各遺伝子を発現抑制して胚盤胞期胚までの発生率への影響を調べた。その結果,16個の候補遺伝子のうち2つの遺伝子で,siRNAインジェクションにより胚盤胞期までの発生率が有意に低下した。この2つの遺伝子はいずれも核内因子であり,初期胚の核内での局在を確認している。今後,本実験で同定した遺伝子が体細胞核移植胚の全能性獲得に関与しているか調べるために,機能解析を進めていく。</p>
  • 初期胚特異的レトロトランスポゾンMERVLの活性化機構の解析, 奥野智美, 樋口智香, 神谷拓磨, 山本真理, 越智浩介, 西野亜理紗, 井橋俊哉, 辻本佳加理, 松橋珠子, 安齋政幸, 黒坂哲, 三谷匡, 山縣一夫, 細井美彦, 松本和也, 宮本圭, Journal of Reproduction and Development, 64, Suppl Japanese Issue, j125, 70-P-70,   2018 09 05 , https://jglobal.jst.go.jp/detail?JGLOBAL_ID=201802285393389434
    Summary:<p>【目的】哺乳類ゲノムの約半分は,トランスポゾン由来の配列で構成されている。特に,Murine Endogenous RetroVirus-L(MERVL)と呼ばれるレトロトランスポゾンは,マウス2細胞期胚にかけてその発現が一過的に上昇する独特な発現様式を示し,2細胞期胚特異的に発現する遺伝子群の転写活性を制御している。また,マウスES細胞においてもMERVLを発現する細胞群が存在する。しかし,MERVLの活性化機構の全容は未だ明らかにされていない。そこで本研究では,マウスES細胞を用いて,MERVLの遺伝子発現調節領域(LTR領域)の活性化が誘導される条件を検討した。【方法】マウスES細胞E14Tg2a株を用い,ヒストン脱アセチル化酵素阻害剤であるTrichostatin A(TSA)及びクロマチンのメチル化状態を変化させるVitamin C(VC)を培地に添加した。まず,10 nM TSA添加区,10 ng/µl VC添加区,10 nM TSA及び10 ng/µl VC添加区において,RT-qPCRを用いて2細胞期胚で強発現する遺伝子の転写量を比較した。さらに,2C::td Tomatoベクターをゲノム上に組み込み,MERVL-LTR領域の活性化を可視化できるES細胞を用いて,各培養条件下におけるMERVL-LTR領域活性化細胞数の割合を調べた。【結果】7日間TSAを添加した培養条件下において,2細胞期胚特異的遺伝子群の転写量が有意に上昇していた。また,TSA処理継続時間に比例してMERVL-LTR領域活性化細胞数の割合が約5%まで増加することがわかった。以上の結果より,ES細胞においてMERVL-LTR領域を活性化する培養条件を示した。また,TSA処理によるMERVL活性化促進の結果より,ゲノムワイドなクロマチン弛緩がMERVL-LTR領域の活性化につながる可能性が示唆された。現在,クロマチン脱凝縮を誘導する因子の過剰発現がMERVL活性化を促すか検討を進めている。</p>
  • 細胞分裂及びDNA複製非依存的リプログラミングシステムの構築, 神谷拓磨, 本上遥, 久米健太, 樋口智香, 奥野智美, 山本真理, 越智浩介, 井橋俊哉, 辻本佳加理, 松橋珠子, 細井美彦, 松本和也, 宮本圭, Journal of Reproduction and Development, 64, Suppl Japanese Issue, j117, 54-P-54,   2018 09 05 , https://jglobal.jst.go.jp/detail?JGLOBAL_ID=201802285849451967
    Summary:<p>【目的】分化した体細胞核を未受精卵子内に移植することにより,リプログラミングが誘導され,クローン動物の作出が可能となる。未受精卵を用いてクローン胚を作成する場合,細胞分裂やDNA複製を経て,移植された体細胞核から胚性遺伝子が発現を開始するため,細胞分裂やDNA複製は転写のリプログラミングに不可欠な要素の一つと考えられてきた。そこで本研究では,転写リプログラミングにおける細胞分裂やDNA複製の寄与を明らかにするため,マウス初期胚を用いて細胞分裂及びDNA複製非依存的に体細胞核の転写リプログラミングを誘導する核移植法の開発を目指す。【方法】C57BL/6雌マウスとDBA/2雄マウスを用いてIVFを行い,その後mKSOM培地で4細胞期胚まで培養した。4細胞期胚をDemecolcine添加培地に移し,細胞周期をG2/M期に停止した。G2/M期停止4細胞期胚にTransgeneによりトレース可能な細胞株を移植し,24時間後に免疫染色を行い,共焦点顕微鏡下で移植細胞核の構造的な変化を観察した。また,G2/M期停止4細胞期胚にC2C12筋芽細胞を核移植し,α-amanitinによって転写を阻害した区と非添加区に分け24時間培養後,RNA-seqによって遺伝子発現を調べた。【結果】免疫染色の結果,移植した細胞核が24時間以内に急速なリモデリングを受け,胚由来の核と似た構造を示すことが分かった。移植核中には2番目のセリンがリン酸化を受けたRNA PolIIが確認され,移植後の細胞核は転写活性を有することが分かった。次にRNA-seqの結果,初期胚で高発現する遺伝子の多くが核移植した4細胞期胚から新たに転写されることが分かった。さらに,Utf1やEsrrbなど4細胞期からES細胞にかけて発現の高い遺伝子の転写も確認した。以上の結果より,マウス4細胞期胚を用いた新規核移植法を示した。また,本実験で発展した核移植法により,細胞分裂及びDNA複製非依存的に体細胞核の転写リプログラミングが誘導できる可能性が示唆された。</p>
  • 畜産領域へのリキッドバイオプシーの展開 2:牛の枝肉成績を肥育中に予測する血清バイオマーカータンパク質の探索, 松橋珠子, 池上春香, 越智浩介, 大林賢伍, 佐野文美, 森隆史, 本廣多胤, 奥野智美, 神谷拓磨, 永井宏平, 宮本圭, 吉廣卓哉, 坂口慎一, 松本和也, 日本畜産学会大会講演要旨, 124th, 184,   2018 03 28 , https://jglobal.jst.go.jp/detail?JGLOBAL_ID=201802243958807831
  • レトロトランスポゾンMERVLの活性化機構の解析, 奥野智美, 山口壮輝, 樋口智香, 神谷拓磨, 山本真理, 越智浩介, 西野亜理紗, 井橋俊哉, 辻本佳加理, 坂本裕子, 松橋珠子, 安齋政幸, 黒坂哲, 三谷匡, 山縣一夫, 細井美彦, 松本和也, 宮本圭, 日本分子生物学会年会プログラム・要旨集(Web), 41st, ROMBUNNO.2P‐0413 (WEB ONLY),   2018 , https://jglobal.jst.go.jp/detail?JGLOBAL_ID=201802273880748976
  • Efficient nuclear reprogramming of somatic cells towards totipotency is supported by synergistic effects of small molecules., K. Miyamoto, Y. Tajima, K. Yoshida, M. Oikawa, R. Azuma, M. Mori, Y. Imasato, G.E. Allen, T. Tsujikawa, T. Tsukaguchi, C.R. Bradshaw, J. Jullien, K. Yamagata, K. Matsumoto, M. Anzai, H. Imai, J.B. Gurdon, M. Yamada, the 50th annual meeting of Japan Society of Developmental Biologists,   2017 05 , Refereed, 招待有り
  • 受精後のユビキチン・プロテアソーム系による母性タンパク質の分解はマウス初期胚発生に重要である, 樋口智香, 守田昂太郎, 山口壮輝, 松橋珠子, 永井宏平, 安齋政幸, 安齋政幸, 山縣一夫, 細井美彦, 宮本圭, 松本和也, 日本卵子学会誌, 2, 1, S48, S48,   2017 04 01 , http://jglobal.jst.go.jp/public/201702240011137048
  • マウス初期胚発生過程におけるH3R2me2sの役割, 守田昂太郎, 樋口智香, 山口壮輝, 松橋珠子, 松橋珠子, 永井宏平, 安齋政幸, 安齋政幸, 加藤博巳, 加藤博巳, 山縣一夫, 細井美彦, 宮本圭, 松本和也, 日本卵子学会誌, 2, 1, S47, S47,   2017 04 01 , http://jglobal.jst.go.jp/public/201702274754974238
  • The improved culture condition for mouse nuclear transfer embryos enables highly efficient nuclear reprogramming., K. Miyamoto, Y. Tajima, K. Yoshida, T. Tsukaguchi, C.R. Bradshaw, G.E. Allen, M. Mori, Y. Imazato, J. Jullien, K. Matsumoto, H. Imai, J.B. Gurdon, M. Yamada, The 3rd China-Japan-Korea reproduction meeting,   2016 08 , Refereed, 招待有り
  • Reprogramming of somatic nuclei towards totipotency is greatly facilitated by small molecules through epigenetic alterations, Kei Miyamoto, Tomomasa Tsukaguchi, Charles R Bradshaw, George E Allen, Jerome Jullien, Kazuya Matsumoto, J. B. Gurdon, Masayasu Yamada, International symposium on epigenome dynamics and regulation in germ cells,   2016 02 , Refereed, 招待有り
  • A transient inhibition of proteasome pathway entails a delay in the onset of ZGA and impairs full term development of mice, HIGUCHI Chika, SHIMIZU Natsumi, MORITA Kohtaro, UCHIBORI Sho, TSUKAGUCHI Tomomasa, NAGAI Kohei, ANZAI Masayuki, YAMAGATA Kazuo, HOSOI Yoshihiko, MIYAMOTO Kei, MATSUMOTO Kazuya, The Journal of Reproduction and Development Supplement, 108, 0, P, 12-P-12,   2015 , http://ci.nii.ac.jp/naid/130005492035
  • Involvement of Peroxiredoxin (Prdx) in reducing hydrogen peroxide in pronuclei of mouse zygotes., MORITA Kohtaro, TOKORO Mikiko, HIGUCHI Chika, UCHIBORI Sho, TSUKAGUCHI Tomomasa, NAGAI Kohei, ANZAI Masayuki, YAMAGATA Kazuo, HOSOI Yoshihiko, MIYAMOTO Kei, MATSUMOTO Kazuya, The Journal of Reproduction and Development Supplement, 108, 0, P, 15-P-15,   2015 , http://ci.nii.ac.jp/naid/130005492029
  • The effect of Coenzyme Q10 on in vitro fertilization and subsequent preimplantation development of post-ovulatory aged oocytes in mice, UCHIBORI Sho, HIGUCHI Chika, MORITA Kohtaro, TSUKAGUCHI Tomomasa, ANZAI Masayuki, YAMAGATA Kazuo, HOSOI Yoshihiko, MIYAMOTO Kei, MATSUMOTO Kazuya, The Journal of Reproduction and Development Supplement, 108, 0, P, 10-P-10,   2015 , http://ci.nii.ac.jp/naid/130005492071
  • 核内Wave1は卵母細胞における転写の初期化と正常な発生に必要である, Kei Miyamoto, Japanese Scientist in Science 2013, 60, 61,   2014 , Refereed, 招待有り
  • The roles of Hsp90 in reprogramming process in mouse preimplantation embryonic development, Luong PQ, Matsumoto M, Suzuki S, Kitamura N, Miyamoto K, Minami N, Yamada M, Imai H, CiRA International Symposium,   2012 , Refereed
  • Mammalian oocyte extracts induce chromatin remodeling and dedifferentiation of somatic cells, but do not global demethylation, K. Miyamoto, T. Tsukiyama, N. Minami, M. Yamada, H. Imai, REPRODUCTION IN DOMESTIC ANIMALS, 43, 194, 194,   2008 07 , Refereed
  • Reprogramming of mouse embryonic fibroblasts by cell-free extracts from nanog-overexpressing embryonic stem cells, Tomoyuki Tsukiyama, Kei Miyamoto, Naojiro Minami, Masayasu Yamada, Hiroshi Imai, BIOLOGY OF REPRODUCTION, 85, 85,   2008 , Refereed
  • DNA demethylation associated with nuclear reprogramming of the somatic cell genome in cell-free extracts from mammalian oocytes, Kei Miyamoto, Tomoyuki Tsukiyama, Yang Yang, Ning Li, Naojiro Minami, Masayasu Yamada, Hiroshi Imai, BIOLOGY OF REPRODUCTION, 55, 55,   2008 , Refereed
  • Differential nuclear reprogramming of somatic cells by porcine germinal vesicle and metaphase II oocyte extracts., Miyamoto K, Tsukiyama T, Minami N, Yamada M, Imai H, The 4th Asian Reproductive Biotechnology Conference,   2007 , Refereed
  • Nuclear reprogramming of porcine cells and their use as donor cell for nuclear transfer after treatment in Xenopus egg extracts, K. Miyamoto, M. Ohnuki, N. Minami, M. Yamada, H. Imai, REPRODUCTION FERTILITY AND DEVELOPMENT, 19, 1, 150, 150,   2007 , Refereed
  • Nuclear reprogramming of porcine fibroblast cells by Xenopus egg extracts., Miyamoto K, Nagao Y, Minami N, Yamada M, Ohsumi K, Imai H, 32nd International Embryo Transfer Society,   2006 , Refereed
  • Reprogramming of intact and permeabilized mammalian somatic cells by Xenopus egg extract, Miyamoto K, Ohnuki M, Minami N, Yamada M, Imai H, The 3rd Asian Reproductive Biotechnology Conference,   2006 , Refereed
  • Cell cycle synchronization of donor cells at G1 phase and developmental ability of nuclear transfer embryos in miniature pigs, Miyamoto K, Hoshino Y, Nagao Y, Minami N, Yamada M, Imai H, The Interational Embryo Transfer Society Meeting, Copenhagen, Denmark,   2005 01 08 , Refereed

Awards & Honors

  •   2018 09 , Society for Reproduction and Development, SRD Young Investigator Award
  •   2018 04 , The Minister of Education, Culture, Sports, Science and Technology, Young Scientists’ Prize
  •   2016 07 , Human Frontier Science Program, Human Frontier Project Grant, Nuclear actin assembly in chromatin structure and dynamics for cell cycle control and reprogramming
  •   2012 04 , Herchel Smith Postdoctoral Fellowship award

Research Grants & Projects

  • collaboration work with Central Institute for Experimental Animals on marmoset
  • Research Fellowship Program, The Grant from the Japan Society for the Promotion of Science