全細胞オミクスによる骨格筋組織を構築するシステムの解明

2021 Fiscal Year Annual Research Report

• Project/Area Number: 20H00456
• Research Institution: Kyushu University
• Principal Investigator: 大川 恭行 九州大学, 生体防御医学研究所, 教授 (80448430)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Keywords: 骨格筋分化 / クロマチン構造 / 単一細胞解析 / 空間オミクス

Outline of Annual Research Achievements

マウス骨格筋組織のうち今回は前腓骨筋を対象とし、筋組織はカルディオトキシン注射により損傷させたのち、１,３,５,７,１４,２１日目の段階で組織切片を作成した。独自に開発したChILSeq法により、単一細胞レベルで、トランスクリプトーム、プロテオーム、エピゲノムデータをそれぞれ取得した。既に公開している単一細胞エピゲノム解析法ChILseqに、新たに開発した連続免疫染色法、空間トランスクリプトーム技術PIC(photo Isolation Chemistry)を組み合わせた新たなマルチオミクス解析を可能した。本解析では、全て組織切片上で行った。切片上に存在する細胞内に含まれるRNAを逆転写後dsDNAに転換し、更にin vitro transcription(IVT)により増幅させることで１細胞レベルでRNAの検出が可能である（単一細胞トランスクリプトーム検出）。同様に細胞に存在する分子をあらかじめ消去可能な蛍光を付加した特異的抗体と反応させ、連続的に反応させることで膨大な抗体の種類を同一検体上で解析することが可能である（単一細胞プロテオーム検出）。更に、既に発表しているChILSeq法(Nat Cell Biol., 2019)では、抗体が反応した周辺のゲノムDNAを転写させRNAとして検出することが可能である。従ってこれらトランスクリプトーム、エピゲノム、プロテオームの情報を一度のRNA解析で取得が可能である（論文発表準備中、特許申請中）。本手法では、プロテオーム、エピゲノムデータは用いる抗体の種類に依存する。そこで、本年度は、特にプロテオームデータは筋、免疫細胞、間質細胞の代表的な細胞表面抗原を検出する抗体による細胞集団の大まかな区分けを行ったうえで、代表的なシグナル伝達経路の活性化に絞って定量解析を行った。エピゲノムデータについてはクロマチン構造状態の計測を行った。骨格筋再生過程における細胞集団の同定に向けた包括的なデータ解析を進めている。

Current Status of Research Progress

1: Research has progressed more than it was originally planned.

Reason

当初予定の開発を早めて、データ取得を進めており当初の計画以上に進展している。

Strategy for Future Research Activity

研究計画に従って進めていく。

Research Products

• [Journal Article] Uterus-specific transcriptional regulation underlies eggshell pigment production in Japanese quail. (2022)
  • Author(s): Satoshi Ishishita, Shumpei Kitahara, Mayuko Takahashi, Sakura Iwasaki, Shoji Tatsumoto, Izumi Hara, Yoshiki Kaneko, Keiji Kinoshita, Katsushi Yamaguchi, Akihito Harada, Yasushige Ohmori, Yasuyuki Ohkawa, Yasuhiro Go, Shuji Shigenobu, Yoichi Matsuda, Takayuki Suzuki.
  • Journal Title: PLoS One. Volume: 17 Pages: e0265008
  • DOI: 10.1371/journal.pone.0265008.
  • Peer Reviewed / Open Access
• [Journal Article] Direct Conversion of Human Endothelial Cells Into Liver Cancer-Forming Cells Using Nonintegrative Episomal Vectors. (2022)
  • Author(s): Takeshi Goya, Kenichi Horisawa, Miyako Udono, Yasuyuki Ohkawa, Yoshihiro Ogawa, Sayaka Sekiya, Atsushi Suzuki.
  • Journal Title: Hepatol Commun Volume: 6 Pages: 1725-1740
  • DOI: 10.1002/hep4.1911.
  • Peer Reviewed / Open Access
• [Journal Article] Uhrf1 governs the proliferation and differentiation of muscle satellite cells. (2022)
  • Author(s): Sakai H, Sawada Y, Tokunaga N, Tanaka K, Nakagawa S, Sakakibara I, Ono Y, Fukada SI, Ohkawa Y, Kikugawa T, Saika T, Imai Y.
  • Journal Title: iScience. Volume: 25 Pages: 103928
  • DOI: 10.1016/j.isci.2022.103928.
  • Peer Reviewed / Open Access
• [Journal Article] Unusual nucleosome formation and transcriptome influence by the histone H3mm18 variant. (2022)
  • Author(s): Hirai S, Tomimatsu K, Miyawaki-Kuwakado A, Takizawa Y, Komatsu T, Tachibana T, Fukushima Y, Takeda Y, Negishi L, Kujirai T, Koyama M, Ohkawa Y, Kurumizaka H.
  • Journal Title: Nucleic Acids Res. Volume: 50 Pages: 72-91
  • DOI: 10.1093/nar/gkab1137.
  • Peer Reviewed / Open Access
• [Journal Article] Live imaging of transcription sites using an elongating RNA polymerase II-specific probe. (2022)
  • Author(s): Uchino S, Ito Y, Sato Y, Handa T, Ohkawa Y, Tokunaga M, Kimura H.
  • Journal Title: J Cell Biol. Volume: 221 Pages: e202104134
  • DOI: 10.1083/jcb.202104134.
  • Peer Reviewed / Open Access
• [Journal Article] Relayed signaling between mesenchymal progenitors and muscle stem cells ensures adaptive stem cell response to increased mechanical load. (2022)
  • Author(s): Kaneshige A, Kaji T, Zhang L, Saito H, Nakamura A, Kurosawa T, Ikemoto-Uezumi M, Tsujikawa K, Seno S, Hori M, Saito Y, Matozaki T, Maehara K, Ohkawa Y, Potente M, Watanabe S, Braun T, Uezumi A, Fukada SI.
  • Journal Title: Cell Stem Cell. Volume: 29 Pages: 265-280
  • DOI: 10.1016/j.stem.2021.11.003.
  • Peer Reviewed / Open Access
• [Journal Article] Recent advances in single-cell epigenomics. (2021)
  • Author(s): Harada A, Kimura H, Ohkawa Y.
  • Journal Title: Curr Opin Struct Biol. Volume: 71 Pages: 116-122
  • DOI: 10.1016/j.sbi.
  • Peer Reviewed / Open Access
• [Journal Article] High-throughput single-cell epigenomic profiling by targeted insertion of promoters (TIP-seq). (2021)
  • Author(s): Bartlett DA, Dileep V, Handa T, Ohkawa Y, Kimura H, Henikoff S, Gilbert DM.
  • Journal Title: J Cell Biol. Volume: 220 Pages: e202103078
  • DOI: 10.1083/jcb.202103078.
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Chromatin loading of MCM hexamers is associated with di-/tri-methylation of histone H4K20 toward S phase entry. (2021)
  • Author(s): Hayashi-Takanaka Y, Hayashi Y, Hirano Y, Miyawaki-Kuwakado A, Ohkawa Y, Obuse C, Kimura H, Haraguchi T, Hiraoka Y.
  • Journal Title: Nucleic Acids Res. Volume: 49 Pages: 12152-12166
  • DOI: 10.1093/nar/gkab1068.
  • Peer Reviewed / Open Access
• [Journal Article] Discriminative feature of cells characterizes cell populations of interest by a small subset of genes. (2021)
  • Author(s): Fujii T, Maehara K, Fujita M, Ohkawa Y.
  • Journal Title: PLoS Comput Biol. Volume: 17 Pages: e1009579
  • DOI: 10.1371/journal.pcbi.1009579.
  • Peer Reviewed / Open Access
• [Journal Article] Sex differences in metabolic pathways are regulated by Pfkfb3 and Pdk4 expression in rodent muscle. (2021)
  • Author(s): Christianto A, Baba T, Takahashi F, Inui K, Inoue M, Suyama M, Ono Y, Ohkawa Y, Morohashi KI.
  • Journal Title: Commun Biol. Volume: 4 Pages: 1264
  • DOI: 10.1038/s42003-021-02790-y.
  • Peer Reviewed / Open Access
• [Journal Article] Neural stem/precursor cells dynamically change their epigenetic landscape to differentially respond to BMP signaling for fate switching during brain development. (2021)
  • Author(s): Katada S, Takouda J, Nakagawa T, Honda M, Igarashi K, Imamura T, Ohkawa Y, Sato S, Kurumizaka H, Nakashima K.
  • Journal Title: Genes Dev. Volume: 35 Pages: 1431-1444
  • DOI: 10.1101/gad.348797.
  • Peer Reviewed / Open Access
• [Journal Article] Modeling population size independent tissue epigenomes by ChIL-seq with single thin sections. (2021)
  • Author(s): Maehara K, Tomimatsu K, Harada A, Tanaka K, Sato S, Fukuoka M, Okada S, Handa T, Kurumizaka H, Saitoh N, Kimura H, Ohkawa Y.
  • Journal Title: Mol Syst Biol. Volume: 17 Pages: e10323
  • DOI: 10.15252/msb.
  • Peer Reviewed / Open Access
• [Journal Article] Genome-wide analysis of chromatin structure changes upon MyoD binding in proliferative myoblasts during the cell cycle. (2021)
  • Author(s): Wu Q, Fujii T, Harada A, Tomimatsu K, Miyawaki-Kuwakado A, Fujita M, Maehara K, Ohkawa Y.
  • Journal Title: J Biochem. Volume: 169 Pages: 653-661
  • DOI: 10.1093/jb/mvab001.
  • Peer Reviewed / Open Access
• [Journal Article] Integrated Analysis of Transcriptome and Histone Modifications in Granulosa Cells During Ovulation in Female Mice. (2021)
  • Author(s): Shirafuta Y, Tamura I, Ohkawa Y, Maekawa R, Doi-Tanaka Y, Takagi H, Mihara Y, Shinagawa M, Taketani T, Sato S, Tamura H, Sugino N.
  • Journal Title: Endocrinology. Volume: 162 Pages: bqab128
  • DOI: 10.1210/endocr/bqab128.PMID: 34171084.
  • Peer Reviewed / Open Access
• [Journal Article] An extensive and dynamic trans-omic network illustrating prominent regulatory mechanisms in response to insulin in the liver. (2021)
  • Author(s): Matsuzaki F, Uda S, Yamauchi Y, Matsumoto M, Soga T, Maehara K, Ohkawa Y, Nakayama KI, Kuroda S, Kubota H.
  • Journal Title: Cell Rep. Volume: 36 Pages: 109569
  • DOI: 10.1016/j.celrep.2021.
  • Peer Reviewed / Open Access
• [Journal Article] Targeted inhibition of EPAS1-driven IL-31 production by a small-molecule compound. (2021)
  • Author(s): Kamikaseda Y, Uruno T, Kunimura K, Harada A, Saiki K, Oisaki K, Sakata D, Nakahara T, Kido-Nakahara M, Kanai M, Nakamura S, Ohkawa Y, Furue M, Fukui Y.
  • Journal Title: J Allergy Clin Immunol. Volume: 148 Pages: 633-638
  • DOI: 10.1016/j.jaci.2021.03.029.
  • Peer Reviewed / Open Access
• [Journal Article] High-depth spatial transcriptome analysis by photo-isolation chemistry. (2021)
  • Author(s): Honda M, Oki S, Kimura R, Harada A, Maehara K, Tanaka K, Meno C, Ohkawa Y.
  • Journal Title: Nat Commun. Volume: 12 Pages: 4416
  • DOI: 10.1038/s41467-021-24691-8.
  • Peer Reviewed / Open Access
• [Journal Article] Transcriptome analysis of gene expression changes upon enzymatic dissociation in skeletal myoblasts. (2021)
  • Author(s): Miyawaki-Kuwakado A, Wu Q, Harada A, Tomimatsu K, Fujii T, Maehara K, Ohkawa Y.
  • Journal Title: Genes Cells. Volume: 26 Pages: 530-540
  • DOI: 10.1111/gtc.12870. Epub 2021 Jun 8.
  • Peer Reviewed / Open Access
• [Journal Article] Hoxa10 mediates positional memory to govern stem cell function in adult skeletal muscle. (2021)
  • Author(s): Yoshioka K, Nagahisa H, Miura F, Araki H, Kamei Y, Kitajima Y, Seko D, Nogami J, Tsuchiya Y, Okazaki N, Yonekura A, Ohba S, Sumita Y, Chiba K, Ito K, Asahina I, Ogawa Y, Ito T, Ohkawa Y, Ono Y.
  • Journal Title: Sci Adv. Volume: 7 Pages: eabd7924
  • DOI: 10.1126/sciadv.abd7924.
  • Peer Reviewed / Open Access
• [Journal Article] Totipotency of mouse zygotes extends to single blastomeres of embryos at the four-cell stage. (2021)
  • Author(s): Maemura M, Taketsuru H, Nakajima Y, Shao R, Kakihara A, Nogami J, Ohkawa Y, Tsukada YI.
  • Journal Title: Sci Rep. Volume: 11 Pages: 11167
  • DOI: 10.1038/s41598-021-90653-1.
  • Peer Reviewed / Open Access
• [Journal Article] Chromatin structure-dependent histone incorporation revealed by a genome-wide deposition assay. (2021)
  • Author(s): Tachiwana H, Dacher M, Maehara K, Harada A, Seto Y, Katayama R, Ohkawa Y, Kimura H, Kurumizaka H, Saitoh N.
  • Journal Title: Elife Volume: 10 Pages: e66290
  • DOI: 10.7554/eLife.66290.
  • Peer Reviewed / Open Access
• [Presentation] 骨格筋再性能を制御するクロマチン構造等その破綻 (2021)
  • Author(s): 大川恭行
  • Organizer: 第94回日本生化学会大会
  • Invited
• [Presentation] 空間マルチオミクスの展開～エピゲノム, トランスクリプトームからプロテオームの空間計測の試み～ (2021)
  • Author(s): 大川恭行
  • Organizer: がん研セミナー
  • Invited
• [Presentation] Identification and Analysis of Minor Histone Variants (2021)
  • Author(s): Yasuyuki Ohkawa
  • Organizer: EMBO Workshop Physiology and function of histone variants
• [Book] 少数細胞エピゲノム解析技術の開発 (2021)
  • Author(s): 原田 哲仁，大川 恭行
  • Total Pages: -
  • Publisher: 日本生化学会
• [Book] 実験医学2021年9月号 (2021)
  • Author(s): 沖 真弥、大川 恭行
  • Total Pages: 131
  • Publisher: 羊土社
  • ISBN: 978-4-7581-2547-5
• [Book] 遺伝子医学MOOK36 (2021)
  • Author(s): 富松航佑, 大川 恭行
  • Total Pages: 218
  • Publisher: 株式会社メディカルドゥ
• [Remarks] 九州大学 トランスクリプトミクス分野ホームページ
  • URL: https://tx.bioreg.kyushu-u.ac.jp/