Molecular design of quadruplex-forming nculeic acids that inhibit the pathological conformational conversion of PrP and elucidation of its inhibitory mechanism

• Project/Area Number: 16K07269
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Structural biochemistry
• Research Institution: Kyoto University
• Principal Investigator: Mashima Tsukasa 京都大学, エネルギー理工学研究所, 助教 (20707426)
• Project Period (FY): 2016-04-01 – 2020-03-31
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥4,940,000 (Direct Cost: ¥3,800,000、Indirect Cost: ¥1,140,000); Fiscal Year 2018: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000); Fiscal Year 2017: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000); Fiscal Year 2016: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
• Keywords: NMR / アプタマー / 四重鎖核酸 / プリオン病 / 立体構造

Outline of Final Research Achievements

Prion diseases are neurodegenerative disorders caused by the conversion of cellular prion protein to pathological isoform. Previously, we obtained an RNA aptamer, R12, that possesses anti-prion activity. We also revealed that R12 folds into a unique G-quadruplex structure and two R12 molecules form a homodimer. Here, we developed new prion aptamers, R24 and R12-A-R12, based on the structure of R12 homodimer. The assay with prion-infected cells showed that both aptamers exhibited much higher anti-prion activity than R12. Our NMR studies showed that the structure of a single R12-A-R12 molecule resembled that of the R12 homodimer. R24 is supposed to unimolecularly form a similar structure of R12 homodimer because the sequence of R24 is almost the same as that of R12-A-R12. The quadruplex structure of either R24 or R12-A-R12 formed by one molecule could be stable when they are administered to a prion-infected cell culture. This may be the reason they possess high anti-prion activity.

Academic Significance and Societal Importance of the Research Achievements

プリオン病は重篤な神経変性疾患であるが、根治できる治療法がなく、効果的な治療薬の開発が望まれている。今回我々が開発したアプタマーは、ある種の抗体などを除き、これまでの抗プリオン物質の中で最も高い効率でプリオンタンパク質の異常化を抑制する。よって治療薬への応用が期待されるため、社会的意義がある。
核酸、特に四重鎖構造を有する核酸は一塩基を置換・付加しただけで大きくそのトポロジーが変わることがあり、その原理は不明である。本研究では新規アプタマーを設計し、その立体構造が意図した通りであるかを確認し、さらに活性を大きく向上させることに成功したため、学術的にも意義がある。

Research Products

• [Int'l Joint Research] Gyeongsang National University(韓国)
• [Journal Article] Development and structural determination of an anti-PrPC aptamer that blocks pathological conformational conversion of prion protein (2020)
  • Author(s): Mashima Tsukasa、Lee Joon-Hwa、Kamatari Yuji O.、Hayashi Tomohiko、Nagata Takashi、Nishikawa Fumiko、Nishikawa Satoshi、Kinoshita Masahiro、Kuwata Kazuo、Katahira Masato
  • Journal Title: Scientific Reports Volume: 10 Issue: 1 Pages: 4934-4934
  • DOI: 10.1038/s41598-020-61966-4
  • NAID: 120006813315
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] RNA sequence and length contribute to RNA-induced conformational change of TLS/FUS (2020)
  • Author(s): Hamad Nesreen、Mashima Tsukasa、Yamaoki Yudai、Kondo Keiko、Yoneda Ryoma、Oyoshi Takanori、Kurokawa Riki、Nagata Takashi、Katahira Masato
  • Journal Title: Scientific Reports Volume: 10 Issue: 1 Pages: 2629-2629
  • DOI: 10.1038/s41598-020-59496-0
  • Peer Reviewed / Open Access
• [Journal Article] The anti‐prion RNA aptamer R12 disrupts the Alzheimer's disease‐related complex between prion and amyloid β (2019)
  • Author(s): Iida Mamiko、Mashima Tsukasa、Yamaoki Yudai、So Masatomo、Nagata Takashi、Katahira Masato
  • Journal Title: The FEBS Journal Volume: 印刷中 Issue: 12 Pages: 2355-2365
  • DOI: 10.1111/febs.14819
  • Peer Reviewed / Open Access
• [Journal Article] Significance of a histone-like protein with its native structure for the diagnosis of asymptomatic tuberculosis (2018)
  • Author(s): Ohara Yukiko、Ozeki Yuriko、Tateishi Yoshitaka、Mashima Tsukasa、Arisaka Fumio、Tsunaka Yasuo、Fujiwara Yoshie、Nishiyama Akihito、Yoshida Yutaka、Kitadokoro Kengo、Kobayashi Haruka、Kaneko Yukihiro、Nakagawa Ichiro、Maekura Ryoji、Yamamoto Saburo、Katahira Masato、Matsumoto Sohkichi
  • Journal Title: PLOS ONE Volume: 13 Issue: 10 Pages: 0204160-0204160
  • DOI: 10.1371/journal.pone.0204160
  • NAID: 120006534592
  • Peer Reviewed / Open Access
• [Journal Article] Plastic roles of phenylalanine and tyrosine residues of TLS/FUS in complex formation with the G-quadruplexes of telomeric DNA and TERRA (2018)
  • Author(s): Kondo Keiko、Mashima Tsukasa、Oyoshi Takanori、Yagi Ryota、Kurokawa Riki、Kobayashi Naohiro、Nagata Takashi、Katahira Masato
  • Journal Title: Scientific Reports Volume: 8 Issue: 1 Pages: 2864-2864
  • DOI: 10.1038/s41598-018-21142-1
  • Peer Reviewed / Open Access
• [Journal Article] DNA G-wire formation using an artificial peptide is controlled by protease activity (2017)
  • Author(s): Usui, K., Okada, A., Sakashita, S., Shimooka, M., Tsuruoka, T., Nakano, S., Miyoshi, D., Mashima, T., Katahira, M., Hamada, Y.
  • Journal Title: Molecules Volume: 22 Issue: 11 Pages: 1991-1991
  • DOI: 10.3390/molecules22111991
  • NAID: 120006501324
  • Peer Reviewed / Open Access
• [Journal Article] Development of an RNA aptamer that acquires binding capacity against HIV-1 Tat protein via G-quadruplex formation in response to potassium ions (2017)
  • Author(s): Yamaoki, Y., Nagata, T., Mashima, T., Katahira, M.
  • Journal Title: Chemical communications Volume: 53 Issue: 52 Pages: 7056-7059
  • DOI: 10.1039/c7cc03312e
  • Peer Reviewed / Open Access
• [Presentation] Development of anti-prion RNA aptamers and destruction of the Alzheimer's disease-related complex (2019)
  • Author(s): Mashima, T., Iida, M., Lee, J., Yamaoki, Y., So, M., Kamatari, Y. O., Hayashi, T., Kinoshita, M., Kuwata, K., Nagata, T. and Katahira, M.
  • Organizer: The 46th International Symposium on Nucleic Acids Chemistry
  • Int'l Joint Research
• [Presentation] Disruption of the Alzheimer’s disease-related complex between prion and amyloid β by an anti-prion aptamer (2019)
  • Author(s): Mashima, T., Iida, M., Yamaoki, Y., So, M., Nagata, T. and Katahira, M.
  • Organizer: Commemorative International Symposium of Nucleic Acid Chemistry
  • Int'l Joint Research
• [Presentation] 非常に高い活性を有する抗プリオンRNAアプタマーの構造機能相関 (2019)
  • Author(s): 真嶋 司、Lee, Joon-Hwa、鎌足 雄司、林 智彦、木下 正弘、桑田 一夫、永田 崇、片平 正人
  • Organizer: 第58回NMR討論会
• [Presentation] プリオンタンパク質の構造変換を抑制するRNAアプタマーの開発と構造決定 (2019)
  • Author(s): 真嶋 司、Lee, Joon-Hwa、鎌足 雄司、林 智彦、永田 崇、西川 富美子、西川 諭、木下 正弘、桑田 一夫、片平 正人
  • Organizer: 第21回日本RNA学会年会
• [Presentation] Development of RNA Aptamer That Has High Anti-prion Activity and Its Structural Basis (2018)
  • Author(s): Mashima, T., Lee, J., Kamatari, Y. O., Hayashi, T., Nishikawa, F., Nagata, T., Nishikawa, S., Kinoshita, M., Kuwata, K. and Katahira, M.
  • Organizer: The 45th International Symposium on Nucleic Acids Chemistry
  • Int'l Joint Research
• [Presentation] 抗プリオン活性を有する四重鎖RNAの構造決定と高い活性の発現機構の解明 (2018)
  • Author(s): 真嶋司、Lee Joon-Hwa、鎌足雄司、林智彦、西川富美子、永田崇、西川諭、木下正弘、桑田一夫、片平正人
  • Organizer: 第57回NMR討論会
• [Presentation] An RNA aptamer disrupts the interaction of prion protein with Amyloid β (2018)
  • Author(s): Iida, M., Mashima, T., Yamaoki, Y., So, M., Nagata, T. and Katahira, M.
  • Organizer: 第56回日本生物物理学会年会
• [Presentation] The effect of different target nucleic acids on the structural change of TLS (2017)
  • Author(s): Hamad, N., Mashima, T., Watanabe, H., Uchihashi, T., Kurokawa, R., Nagata, T. Katahira, M.
  • Organizer: 2017年度生命科学系学会合同年次大会
• [Presentation] Interaction between non-coding RNA and TLS/FUS protein, and structural change (2017)
  • Author(s): Mashima, T., Nagata, T., Hamad, N., Ozawa, S., Yamaoki, Y., Kondo, K., Watanabe, H., Yoneda, R., Uchihashi, T., Kurokawa, R. Katahira, M.
  • Organizer: The 44th International Symposium on Nucleic Acids Chemistry
  • Int'l Joint Research
• [Presentation] 非コードRNAとTLS/FUSタンパク質の相互作用様式と結合に伴うTLS/FUSの大きな構造変化 (2017)
  • Author(s): 真嶋司，ハマッド ネスリン，小澤駿介，渡辺大輝，内橋貴之，米田竜馬，黒川理樹，永田崇，片平正人
  • Organizer: 第56回NMR討論会
• [Presentation] テロメア長短縮をもたらすTLS/FUS蛋白質とテロメアDNAおよびTERRAのグアニン四重鎖との複合体に関するNMR解析 (2017)
  • Author(s): 近藤敬子, 真嶋司, 大吉崇文, 黒川理樹, 小林直宏, 永田崇, 片平正人
  • Organizer: 第55回日本生物物理学会年会
• [Presentation] プリオンアプタマーはアルツハイマー病に関与するプリオン蛋白質と Aβ オリゴマーの複合体の形成を阻害する (2017)
  • Author(s): 飯田真美子, 真嶋司, 山置佑大, 永田崇, 片平正人
  • Organizer: 第55回日本生物物理学会年会
• [Presentation] TLS/FUS の非コード RNA の認識機構の構造学的研究 (2017)
  • Author(s): 真嶋司，小澤 駿介, N. Hamad, 米田 竜馬, 黒川理樹，永田崇，片平正人
  • Organizer: 第19回日本RNA学会年会
• [Presentation] Conformational change detection of translocated in liposarcoma, TLS, upon binding to various RNAs (2017)
  • Author(s): Hamad, N., Mashima, T., Yamaoki, Y., Kondo, K., Kurokawa, R., Nagata, T. Katahira, M.
  • Organizer: 第19回日本RNA学会年会
• [Presentation] TLS/FUS のRNA認識機構と結合に伴うコンフォメーション変化の研究 (2017)
  • Author(s): 真嶋司，小澤 駿介, N. Hamad, 米田 竜馬, 黒川理樹，永田崇，片平正人
  • Organizer: 第17回日本蛋白質科学会年会
• [Presentation] Invention of RNA aptamer and ribozyme whose activities switch on in response to potassium ion via G-quadruplex formation (2017)
  • Author(s): Yamaoki, Y., Mashima, T., Nagata, T. Katahira, M.
  • Organizer: 6th International Meeting on Quadruplex Nucleic Acids
  • Int'l Joint Research
• [Presentation] 三者複合体形成によってテロメア短縮をもたらすTLS/FUSのテロメアDNAおよびTERRA認識機構に関するNMR解析 (2016)
  • Author(s): 近藤敬子, 真嶋司, 大吉崇文, 永田崇, 片平正人
  • Organizer: 第39回日本分子生物学会年会
  • Place of Presentation: パシフィコ横浜、神奈川県
  • Year and Date: 2016-11-30
• [Presentation] 非コードRNAと結合してcyclin D1転写を抑制するTLS/FUSのRNA認識機構の構造学的基盤 (2016)
  • Author(s): 真嶋司, 小澤駿介, 近藤敬子, Nesreen Hamad, 米田竜馬, 黒川理樹, 永田崇, 片平正人
  • Organizer: 第39回日本分子生物学会年会
  • Place of Presentation: パシフィコ横浜、神奈川県
  • Year and Date: 2016-11-30
• [Presentation] プリオンタンパク質とプリオンタンパク質を標的とする RNA 分子の Aβ線維化への影響 (2016)
  • Author(s): M. Iida, T. Mashima, Y. Yamaoki, T. Nagata
  • Organizer: 生物物理学会第54回年会
  • Place of Presentation: つくば国際会議場、茨城県
  • Year and Date: 2016-11-25
• [Presentation] Cyclin D1遺伝子転写抑制やテロメア長制御に関わる TLS/FUS タンパク質と非コード核酸間の相互作用のＮＭＲ法による解析 (2016)
  • Author(s): 近藤敬子, 真嶋司, 大吉崇文, 米田竜馬, 黒川理樹, 永田崇, 片平正人
  • Organizer: 第55回NMR討論会
  • Place of Presentation: 広島国際会議場、広島県
  • Year and Date: 2016-11-16
• [Presentation] C:G塩基対を選択的に認識する人工塩基を含む三重鎖DNAの構造解析 (2016)
  • Author(s): 真嶋司、岡村秀紀、王磊、谷口陽祐、佐々木茂貴、片平正人
  • Organizer: 第55回NMR討論会
  • Place of Presentation: 広島国際会議場、広島県
  • Year and Date: 2016-11-16
• [Presentation] リグノセルロースバイオリファイナリーのためのリグニン・ペプチド間相互作用解析 (2016)
  • Author(s): 山口亜佐子，末富高志，大城理志，徳永有希，西村裕司，永田崇，真嶋司，片平正人，礒﨑勝弘，高谷光，中村正治，渡辺隆司
  • Organizer: 第61回リグニン討論会
  • Place of Presentation: 京都大学、京都府
  • Year and Date: 2016-10-27
• [Presentation] Non-coding RNA/DNA recognition by TLS/FUS that causes repression of cyclin D1 transcription and telomere elongation (2016)
  • Author(s): K. Kondo, T. Mashima, T. Oyoshi, R. Yoneda, R. Kurokawa, T. Nagata, M. Katahira
  • Organizer: The 43rd International Symposium on Nucleic Acids Chemistry
  • Place of Presentation: 熊本大学、熊本県
  • Year and Date: 2016-09-27
  • Int'l Joint Research
• [Presentation] カリウムイオン応答性Tat結合RNAアプタマーおよびハンマーヘッドリボザイムの創製および in-cell NMR法による細胞内核酸の観測 (2016)
  • Author(s): 山置佑大, 清石彩華, 真嶋司, 加納ふみ, 村田昌之, 永田崇, 片平正人
  • Organizer: 第10回バイオ関連化学シンポジウム
  • Place of Presentation: 県立音楽堂、石川県
  • Year and Date: 2016-09-07
• [Presentation] NMR studies of non-coding RNA and DNA recognition by TLS/FUS which induces transcriptional repression of cyclin D1 and telomere shortening (2016)
  • Author(s): K. Kondo, T. Mashima, T. Oyoshi, R. Yoneda, R. Kurokawa, T. Nagata, M. Katahira
  • Organizer: The XXVIIth International Conference on Magnetic Resonance in Biological Systems
  • Place of Presentation: 京都国際会館、京都府
  • Year and Date: 2016-08-21
  • Int'l Joint Research
• [Presentation] K+-responsive activation of dual-quadruplex forming ribozyme system requiring no heating and cooling treatment (2016)
  • Author(s): Y. Yamaoki, T. Mashima, T. Nagata, M. Katahira
  • Organizer: The 21st Annual Meeting of the RNA Society
  • Place of Presentation: 京都国際会館、京都府
  • Year and Date: 2016-06-28
  • Int'l Joint Research
• [Presentation] NMR analysis of the recognition of non-coding RNA and DNA by TLS/FUS which causes transcriptional repression of cyclin D1 1 and telomere shorteningThe 21st Annual Meeting of the RNA Society (2016)
  • Author(s): T. Mashima. K. Kondo, T. Oyoshi, R. Yoneda, R. Kurokawa, T. Nagata, M. Katahira
  • Organizer: The 21st Annual Meeting of the RNA Society
  • Place of Presentation: 京都国際会館、京都府
  • Year and Date: 2016-06-28
  • Int'l Joint Research
• [Presentation] プリオン蛋白質とそれを標的とする RNA 分子の Aβ線維化への影響 (2016)
  • Author(s): 飯田真美子, 真嶋司, 山置佑大, 永田崇, 片平正人
  • Organizer: 第16回日本蛋白質科学会年会
  • Place of Presentation: 福岡国際会議場、福岡県
  • Year and Date: 2016-06-07
• [Presentation] Cyclin D1転写抑制を担う TLS/FUS タンパク質と非コードRNA の相互作用の NMR 法による解析 (2016)
  • Author(s): 真嶋司, 小澤駿介, 近藤敬子, 永田崇, 黒川理樹, 片平正人
  • Organizer: 第16回日本蛋白質科学会年会
  • Place of Presentation: 福岡国際会議場、福岡県
  • Year and Date: 2016-06-07
• [Presentation] K+-responsive activation of ribozyme system via structural transition from duplex to dual-quadruplex requiting no heating and cooling treatment (2016)
  • Author(s): Y. Yamaoki, T. Mashima, T. Nagata, M. Katahira
  • Organizer: The XXVIIth International Conference on Magnetic Resonance in Biological Systems
  • Place of Presentation: 京都国際会館、京都府
  • Int'l Joint Research