Tissue-specific pre-mRNA processing in living animals

• Project/Area Number: 20H03181
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Review Section: Basic Section 43010:Molecular biology-related
• Research Institution: University of the Ryukyus
• Principal Investigator: KUROYANAGI HIDEHITO 琉球大学, 医学(系)研究科(研究院), 教授 (30323702)
• Project Period (FY): 2020-04-01 – 2023-03-31
• Project Status: Completed (Fiscal Year 2023)
• Budget Amount: ¥17,810,000 (Direct Cost: ¥13,700,000、Indirect Cost: ¥4,110,000); Fiscal Year 2022: ¥5,590,000 (Direct Cost: ¥4,300,000、Indirect Cost: ¥1,290,000); Fiscal Year 2021: ¥5,590,000 (Direct Cost: ¥4,300,000、Indirect Cost: ¥1,290,000); Fiscal Year 2020: ¥6,630,000 (Direct Cost: ¥5,100,000、Indirect Cost: ¥1,530,000)
• Keywords: 遺伝子発現制御 / 新生RNA / 線虫 / スプライシング / 転写後プロセシング / 塩基修飾 / 代謝標識 / 長鎖シーケンシング解析 / RNAプロセシング / mRNA前駆体 / 遺伝子発現 / 組織特異性

Outline of Research at the Start

真核生物のmRNA前駆体の選択的プロセシングはタンパク質の多様性を創出する重要な遺伝子発現機構であり、多細胞生物の多くの遺伝子で組織特異的な制御が見られる。本研究では、線虫の生体で組織特異的にRNAを濃縮する手法を開発し、組織特異的選択的プロセシングの概要を明らかにする。さらに、組織特異的制御因子の変異体を用いて同様の解析を行い、選択的mRNAプロセシングの制御機構を体系的に理解することを目指す。

Outline of Final Research Achievements

We developed new methods to label nascent precursor messenger RNAs (pre-mRNAs) in vivo and successfully purified them by using a nematode worm C. elegans as a model multicellular organism. With the methods, we purified pre-mRNAs from a mutant worms strain defective in mRNA surveillance system and subjected them to long-read cDNA sequencing. We successfully identified genes whose expression levels are regulated at the pre-mRNA processing level coupled with the surveillance system genome wide. Among them was a gene whose expression level was regulated by nucleotide modification at the very splice acceptor site.

Academic Significance and Societal Importance of the Research Achievements

mRNA前駆体のプロセシング動態および品質管理機構と共役する遺伝子発現制御が多細胞生物の個体レベルで明らかになるのは初めてである。哺乳類では個体レベルで同様の方法を適用しても同様の結果を得ることは実質的に困難である。また、転写後の塩基修飾が直接的にスプライシングに影響を与える例を始めて見出したものである。本研究成果は、哺乳類を含む他の多細胞動物における転写と共役したmRNAの転写後プロセシングによる遺伝子発現制御機構を考察する上で基礎となる重要な知見を与えるものである。

Research Products

• [Int'l Joint Research] University of Pittsburgh Sch Medicine(米国)
• [Int'l Joint Research] University of Oslo(ノルウェー)
• [Journal Article] Structure of the<i>Caenorhabditis elegans</i>m6A methyltransferase METT10 that regulates SAM homeostasis (2023)
  • Author(s): Ju Jue、Aoyama Tomohiko、Yashiro Yuka、Yamashita Seisuke、Kuroyanagi Hidehito、Tomita Kozo
  • Journal Title: Nucleic Acids Research Volume: 51 Issue: 5 Pages: 2434-2446
  • DOI: 10.1093/nar/gkad081
  • Peer Reviewed / Open Access
• [Journal Article] Simultaneous measurement of nascent transcriptome and translatome using 4-thiouridine metabolic RNA labeling and translating ribosome affinity purification (2023)
  • Author(s): Imai Hirotatsu、Utsumi Daisuke、Torihara Hidetsugu、Takahashi Kenzo、Kuroyanagi Hidehito、Yamashita Akio
  • Journal Title: Nucleic Acids Research Volume: 51 Issue: 14 Pages: e76-e76
  • DOI: 10.1093/nar/gkad545
  • Peer Reviewed / Open Access
• [Journal Article] Alternative splicing of a single exon causes a major impact on the affinity of Caenorhabditis elegans tropomyosin isoforms for actin filaments (2023)
  • Author(s): Ono Shoichiro、Watabe Eichi、Morisaki Keita、Ono Kanako、Kuroyanagi Hidehito
  • Journal Title: Frontiers in Cell and Developmental Biology Volume: 11 Pages: 1208913-1208913
  • DOI: 10.3389/fcell.2023.1208913
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Comprehensive analysis of alternative splicing coupled with nonsense-mediated mRNA decay (AS-NMD) in C. elegans (2022)
  • Author(s): Hidehito Kuroyanagi
  • Journal Title: 生化学 Volume: 94 Issue: 6 Pages: 868-874
  • DOI: 10.14952/SEIKAGAKU.2022.940868
  • ISSN: 0037-1017
  • Year and Date: 2022-12-25
  • Peer Reviewed / Open Access
• [Journal Article] m ⁶ A‐mediated alternative splicing coupled with nonsense‐mediated mRNA decay regulates SAM synthetase homeostasis (2021)
  • Author(s): Watabe Eichi、Togo‐Ohno Marina、Ishigami Yuma、Wani Shotaro、Hirota Keiko、Kimura‐Asami Mariko、Hasan Sharmin、Takei Satomi、Fukamizu Akiyoshi、Suzuki Yutaka、Suzuki Tsutomu、Kuroyanagi Hidehito
  • Journal Title: The EMBO Journal Volume: 40 Issue: 14
  • DOI: 10.15252/embj.2020106434
  • Peer Reviewed / Open Access
• [Journal Article] Loss of <i>Caenorhabditis elegans</i> homologue of human MOB4 compromises life span, health life span and thermotolerance (2021)
  • Author(s): Jahan Momotaj、Iwasa Hiroaki、Kuroyanagi Hidehito、Hata Yutaka
  • Journal Title: Genes to Cells Volume: 26 Issue: 10 Pages: 798-806
  • DOI: 10.1111/gtc.12891
  • Peer Reviewed / Open Access
• [Presentation] Alternative splicing through m6A modification at a 3' splice site for SAM synthetase homeostasis (2021)
  • Author(s): Eichi Watabe, Marina Togo-Ohno, Yuma Ishigami, Keiko Hirota, Akiyoshi Fukamizu, Yutaka Suzuki, Tsutomu Suzuki and Hidehito Kuroyanagi
  • Organizer: 26th Annual Meeting of the RNA Society
  • Int'l Joint Research
• [Presentation] Alternative splicing through m6A modification at a 3’ splice site for SAM synthetase homeostasis (2021)
  • Author(s): Eichi Watabe, Marina Togo-Ohno, Yuma Ishigami, Keiko Hirota, Akiyoshi Fukamizu, Yutaka Suzuki, Tsutomu Suzuki and Hidehito Kuroyanagi
  • Organizer: 23rd International C. elegans Conference
  • Int'l Joint Research
• [Book] 遺伝学の百科事典 (2022)
  • Author(s): 日本遺伝学会 黒柳秀人（分担執筆）
  • Total Pages: 690
  • Publisher: 丸善出版
  • ISBN: 9784621306604
• [Remarks] 琉球大学大学院医学研究科生化学講座
  • URL: https://biochem.med.u-ryukyu.ac.jp/research/detail/120
• [Remarks] 東京医科歯科大学プレスリリース
  • URL: https://www.tmd.ac.jp/files/topics/55271_ext_04_2.pdf
• [Remarks] 琉球大学大学院医学研究科生化学講座
  • URL: http://biochem.med.u-ryukyu.ac.jp/news/detail/975