Research activity information

• Dec. 2006 ハート昆虫研究奨励基金, 優秀賞

  光延 仁志

• Development of a highly efficient base editing system for Lactobacilli to improve probiotics and dissect essential functions

  Hitoshi Mitsunobu, Yudai Kita, Yumiko Nambu-Nishida, Shoko Miyazaki, Kensuke Nakajima, Ken-ichiro Taoka, Akihiko Kondo, Keiji Nishida

  Abstract Lactobacilli play essential roles in the food industry and have a significant potential as probiotics and therapeutic agents. Genomic and genetic information has increasingly accumulated and been linked to their various functions, to which transgenic approaches are being performed to verify crucial genes. In order to reasonably develop more useful strains, beneficial traits need to be introduced into any given strains and enhanced or combined based on such genotype characterization. However, for practical use as probiotics or foods, organisms with transgene are hardly acceptable. Here, we have introduced the base editing Target-AID system specifically for Lactobacilli, enabling precise installation of point mutations without donor DNA and at multiple genomic loci simultaneously. Lactiplantibacillus plantarum has been successfully engineered to reduce production of imidazole propionate, which has been reported to be associated with type 2 diabetes by impairing glucose tolerance and insulin signaling. Additionally, this system enabled transient knock-out of an essential gene, such as one involved in cell division, resulting in severe filamentous cell phenotype. This demonstrates Target-AID is a promising genetic tool for Lactobacilli and can accelerate both applied and fundamental research. Key points • Efficient and multiplexable cytosine base editing established in Lactobacilli. • Edited Lactobacillus reducing imidazole propionate associated with the risk of type 2 diabetes. • Transient knock-out and dissection of an essential gene function.

  Lead, Springer Science and Business Media LLC, Apr. 2025, Applied Microbiology and Biotechnology, 109(1) (1)

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Mechanism of Annealing of Complementary DNA Strands by the Single-Stranded DNA Binding Protein of Bacteriophage T7

  Seung-Joo Lee, Charlotte Ferguson, Sebastian Urbano, Jaehun Lee, Peter Jeong, Meghana Cheela, Hitoshi Mitsunobu, Bin Zhu, Ashmita Prajapati, Charles C. Richardson, Alfredo J. Hernandez

  American Chemical Society (ACS), Mar. 2025, Biochemistry, 64(7) (7), 1550 - 1559

  [Refereed]

  Scientific journal


• Cytosine base editing systems with minimized off-target effect and molecular size

  Ang Li, Hitoshi Mitsunobu, Shin Yoshioka, Takahisa Suzuki, Akihiko Kondo, Keiji Nishida

  Abstract Cytosine base editing enables the installation of specific point mutations without double-strand breaks in DNA and is advantageous for various applications such as gene therapy, but further reduction of off-target risk and development of efficient delivery methods are desired. Here we show structure-based rational engineering of the cytosine base editing system Target-AID to minimize its off-target effect and molecular size. By intensive and careful truncation, DNA-binding domain of its deaminase PmCDA1 is eliminated and additional mutations are introduced to restore enzyme function. The resulting tCDA1EQ is effective in N-terminal fusion (AID-2S) or inlaid architecture (AID-3S) with Cas9, showing minimized RNA-mediated editing and gRNA-dependent/independent DNA off-targets, as assessed in human cells. Combining with the smaller Cas9 ortholog system (SaCas9), a cytosine base editing system is created that is within the size limit of AAV vector.

  Springer Science and Business Media LLC, Aug. 2022, Nature Communications, 13(1) (1), 4531

  [Refereed]

  Scientific journal

  Link to Kobe Univ. Repository (Kernel)


• Mammalian synthetic biology by CRISPRs engineering and applications

  Kenta Katayama, Hitoshi Mitsunobu, Keiji Nishida

  Oct. 2019, Current Opinion in Chemical Biology, 52, 79 - 84, English

  [Refereed][Invited]

  Scientific journal


• Deaminase-mediated multiplex genome editing in Escherichia coli

  Satomi Banno, Keiji Nishida, Takayuki Arazoe, Hitoshi Mitsunobu, Akihiko Kondo

  Nature Publishing Group, Apr. 2018, Nature Microbiology, 3(4) (4), 423 - 429, English

  [Refereed]

  Scientific journal


• Beyond Native Cas9: Manipulating Genomic Information and Function

  Hitoshi Mitsunobu, Jun Teramoto, Keiji Nishida, Akihiko Kondo

  Oct. 2017, TRENDS IN BIOTECHNOLOGY, 35(10) (10), 983 - 996, English

  [Refereed]

  Scientific journal


• Deep-sea vent phage DNA polymerase specifically initiates DNA synthesis in the absence of primers

  Bin Zhu, Longfei Wang, Hitoshi Mitsunobu, Xueling Lu, Alfredo J. Hernandez, Yukari Yoshida-Takashima, Takuro Nunoura, Stanley Tabor, Charles C. Richardson

  Mar. 2017, PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 114(12) (12), E2310 - E2318, English

  [Refereed]

  Scientific journal


• Dynamics of polycomb proteins-mediated histone modifications during UV irradiation-induced DNA damage

  Zhiqing Li, Hiroaki Mon, Hitoshi Mitsunobu, Li Zhu, Jian Xu, Jae Man Lee, Takahiro Kusakabe

  Dec. 2014, INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY, 55, 9 - 18, English

  [Refereed]

  Scientific journal


• Roles of Piwi Proteins in Transcriptional Regulation Mediated by HP1s in Cultured Silkworm Cells

  Tsuneyuki Tatsuke, Li Zhu, Zhiqing Li, Hitoshi Mitsunobu, Kaito Yoshimura, Hiroaki Mon, Jae Man Lee, Takahiro Kusakabe

  Mar. 2014, PLOS ONE, 9(3) (3), e92313, English

  [Refereed]

  Scientific journal


• Flap endonuclease of bacteriophage T7: Possible roles in RNA primer removal, recombination and host DNA breakdown

  MITSUNOBU Hitoshi, ZHU Bin, LEE Seung-Joo, TABOR Stanley, RICHARDSON Charles C

  Mar. 2014, Bacteriophage, 4, e28507, English

  [Refereed]

  Scientific journal


• Flap Endonuclease Activity of Gene 6 Exonuclease of Bacteriophage T7

  Hitoshi Mitsunobu, Bin Zhu, Seung-Joo Lee, Stanley Tabor, Charles C. Richardson

  Feb. 2014, JOURNAL OF BIOLOGICAL CHEMISTRY, 289(9) (9), 5860 - 5875, English

  [Refereed]

  Scientific journal


• Establishment of a Bombyx mori nucleopolyhedrovirus (BmNPV) hyper-sensitive cell line from the silkworm e21 strain

  Jae Man Lee, Naoya Kawakami, Hiroaki Mon, Hitoshi Mitsunobu, Kazuhiro Iiyama, Satoshi Ninaki, Katsumi Maenaka, Enoch Y. Park, Takahiro Kusakabe

  Oct. 2012, BIOTECHNOLOGY LETTERS, 34(10) (10), 1773 - 1779, English

  [Refereed]

  Scientific journal


• Post-translational modifications of the N-terminal tail of histone H3 in holocentric chromosomes of Bombyx mori

  Hiroaki Mon, Makiko Izumi, Hitoshi Mitsunobu, Tsuneyuki Tatsuke, Kazuhiro Iiyama, Hiroyuki Jikuya, Jae Man Lee, Takahiro Kusakabe

  Nov. 2011, INSECT BIOCHEMISTRY AND MOLECULAR BIOLOGY, 41(11) (11), 902 - 908, English

  [Refereed]

  Scientific journal


• Molecular characterization of heterochromatin proteins 1a and 1b from the silkworm, Bombyx mori

  MITSUNOBU Hitoshi, IZUMI Makiko, MON Hiroaki, TATSUKE Tsuneyuki, LEE Jae Min, KASUKABE Takahiro

  Feb. 2011, Insect Molecular Biology, 21(2) (2), 9 - 20, English

  [Refereed]

  Scientific journal


• Efficient soluble protein production on transgenic silkworms expressing cytoplasmic chaperones

  Sun Mee Hong, Jun Yamashita, Hitoshi Mitsunobu, Keiro Uchino, Isao Kobayashi, Hideki Sezutsu, Toshiki Tamura, Hideki Nakajima, Yoshitaka Miyagawa, Jae Man Lee, Hiroaki Mon, Yoshihiko Miyata, Yutaka Kawaguchi, Takahiro Kusakabe

  Aug. 2010, APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 87(6) (6), 2147 - 2156, English

  [Refereed]

  Scientific journal


• Molecular characterization of core histones in the silkworm, Bombyx mori

  MITSUNOBU Hitoshi, IZUMI Makiko, IIYAMA Kazuhiro, JIKUYA Hiroyuki, LEE Jae Man, Mon Hiroaki, KAWAGUCHI Yutaka, KUSAKABE Takahiro

  2010, Journal of Insect Biotechnology and Sericology, 79, 75 - 83, English

  [Refereed]

  Scientific journal


• Molecular Characterization, Localization, and Distribution of Innexins in the Silkworm, Bombyx mori

  Sun Mee Hong, Si Kab Noh, Kyung A. Kim, Hitoshi Mitsunobu, Hiroaki Mon, Jae Man Lee, Yutaka Kawaguchi, Takahiro Kusakabe

  Sep. 2009, MOLECULAR BIOTECHNOLOGY, 43(1) (1), 52 - 58, English

  [Refereed]

  Scientific journal


• Construction of gene expression systems in insect cell lines using promoters from the silkworm, Bombyx mori

  Jae Man Lee, Masateru Takahashi, Hiroaki Mon, Hitoshi Mitsunobu, Katsumi Koga, Yutaka Kawaguchi, Yumiko Nakajima, Takahiro Kusakabe

  Jan. 2008, JOURNAL OF BIOTECHNOLOGY, 133(1) (1), 9 - 17, English

  [Refereed]

  Scientific journal


• Erratum: "Screening of high-permissive silkworm strains for efficient recombinant protein production in Autographa californica nuclear polyhedrosis virus (AcNPV)" (Journal of Insect Biotechnology and Sericology vol. 76 (2) (101-105))

  Jae Man Lee, Hiroaki Mon, Chisa Yasunaga-Aoki, Masateru Takahashi, Naoya Kawakami, Hitoshi Mitsunobu, Yutaka Banno, Katsumi Koga, Keiro Uchino, Yutaka Kawaguchi, Takahiro Kusakabe

  Oct. 2007, Journal of Insect Biotechnology and Sericology, 76(3) (3), 169

  Scientific journal


• Molecular cloning of silkworm Cdc37 and its interaction with Hsp90 chaperon

  YAMASHITA Jun, MIYAGAWA Yoshitaka, SUGAHARA Ryohei, MON Hiroaki, MITSUNOBU Hitoshi, LEE Jae Man, KAWAGUCHI Yutaka, KUSAKABE Takahiro

  Oct. 2007, Journal of Insect Biotechnology and Sericology, 76, 137 - 143, English

  [Refereed]

  Scientific journal


• Heterotrimeric complex of replication protein A, a single-stranded DNA binding protein, from the silkworm, Bombyx mori

  SUGAHARA Ryohei, MON Hiroaki, YAMASHITA Jun, MITSUNOBU Hitoshi, LEE Jae Man, KAWAGUCHI Yutaka, KOGA Katsumi, KUSAKABE Takahiro

  Jun. 2007, Journal of Insect Biotechnology and Sericology, 76, 129 - 135, English

  [Refereed]

  Scientific journal


• Construction of gateway-based destination vectors for detecting subcellular localization of proteins in the silkworm, Bombyx mori

  MITSUNOBU Hitoshi, SAKASHITA Kosuke, MON Hiroaki, YOSHIDA Hiroko, LEE Jae Man, KAWAGUCHI Yutaka, KOGA Katsumi, KUSAKABE Takahiro

  Japanese Society of Sericultural Science, Oct. 2006, Journal of Insect Biotechnology and Sericology, 75(3) (3), 141 - 145, English

  [Refereed]

  Scientific journal

• 乳酸菌の遺伝子情報を精密に改変可能なゲノム編集技術

  光延 仁志, 西田 敬二

  Lead, 一般財団法人 食品産業センター, Sep. 2025, 明日の食品産業, 2025(9) (9), 48 - 53

  [Invited]


• 塩基編集ツールの開発と応用—特集 ゲノム編集医療 : 技術開発・治療応用戦略を中心に

  光延 仁志, 西田 敬二

  Lead, 大阪 : メディカルドゥ, Oct. 2020, 遺伝子医学 = Gene & medicine, 10(4) (4), 22 - 27, Japanese


• カイコ・ホルモンレセプターに対するBmHSP90コシャペロンの機能解析

  山下隼, 門宏明, 光延仁志, 沢津橋俊, 加藤茂明, LEE J.Man, 河口豊, 日下部宣宏

  2009, 日本分子生物学会年会講演要旨集, 32nd(Vol.3) (Vol.3)


• 哺乳類MAPK及び昆虫ホルモンレセプターに対するカイコHSP90コシャペロンの相互作用

  山下隼, 門宏明, 宮川世志幸, 光延仁志, 李在萬, 河口豊, 日下部宜宏

  2009, 日本蚕糸学会大会・蚕糸・昆虫機能学術講演会講演要旨集, 79th


• カイコシャペロン過剰発現のもたらすターゲットタンパク質への影響

  山下隼, 山下隼, 宮川世志幸, 門宏明, 光延仁志, 光延仁志, 久保雄二, 沢津橋俊, 加藤茂明, LEE Jaeman, 河口豊, 日下部宜宏

  2008, 生化学


• Screening of high-permissive silkworm strains for efficient recombinant protein production in Autographa californica nuclear polyhedrosis virus (AcNPV)

  Jae Man Lee, Hiroaki Mon, Masateru Takahashi, Naoya Kawakami, Hitoshi Mitsunobu, Yutaka Banno, Katsumi Koga, Keiro Uchino, Yutaka Kawaguchi, Takahiro Kusakabe

  01 Jun. 2007, Journal of Insect Biotechnology and Sericology, 76, 101 - 105


• カイコTPRドメインタンパク質BmCprA1,BmFKBP59,BmHOPとBmHsp90との相互作用

  山下隼, 宮川世志幸, 門宏明, 光延仁志, LEE Jae Man, 河口豊, 日下部宜宏

  2007, 生化学


• Screening of high-permissive strains for efficient recombinant protein production in Autographa californica nuclear polyhedrosis virus (AcNPV)

  Lee J.-M, Mon H, Aoki-Yasunaga, C.Takahashi M, Kawakami N, Mitsunobu H, Banno Y, Koga K, Uchino K, Kawaguchi Y, Kusakabe T

  2007, Journal of Insect Biotechnology and Sericology, 76(2) (2), 101 - 105

• Bio Clinica The basics and clinical applications of genome editing

  光延 仁志, 西田 敬二

  塩基編集の基礎と応用, 北隆館, Jul. 2025


• 最新のゲノム編集技術と用途展開 第5章 DNA二重鎖切断を伴わない塩基編集技術

  大西 美和, 光延 仁志, 西田 敬二

  シーエムシー出版, Feb. 2021, Japanese, ISBN: 9784781315867


• ゲノム編集技術を応用した製品開発とその実用化 3-5 デアミナーゼを用いたゲノム編集の開発とそのオフターゲット効果の評価

  Li Ang, 光延 仁志, 西田 敬二

  第3章 5節 デアミナーゼを用いたゲノム編集の開発とそのオフターゲット効果の評価, 技術情報協会, Feb. 2021, Japanese, ISBN: 9784861048272


• Gene & Medicine

  メディカルドゥ, Oct. 2020, Japanese, ISBN: 9784909508096

• Target-AID enables efficient point mutagenesis in Lactobacilli

  光延 仁志

  第30回腸内細菌学会学術集会, Jun. 2026

  Oral presentation


• Target-AID base editing enables precise genetic manipulation in lactic acid bacteria

  光延 仁志, 喜多 雄大, 南部 由美子, 宮崎 翔子, 中島 健介, 西田 敬二

  The 47th Annual Meeting of the Molecular Biology Society of Japan, Nov. 2024

  Oral presentation


• 乳酸菌の高効率ゲノム改変を実現する塩基編集技術 Target-AID

  光延 仁志, 喜多 雄大, 南部 由美子, 宮崎 翔子, 中島 健介, 西田 敬二

  第４回先端バイオ工学研究センター成果発表会, Sep. 2024


• Development of DNA base editing technology and its applications

  光延 仁志

  システムバイオロジー研究セミナ 〜UC San Diego & Kobe Univ. STIN〜, 公益財団法人神戸医療産業都市推進機構, 神戸大学統合究拠点

  [Invited]

  Invited oral presentation

• カイコにおけるジーンターゲティングの分子機構とその効率向上

  光延 仁志

  日本学術振興会, 科学研究費助成事業, 特別研究員奨励費, 九州大学, 2007 - 2009

  前年度のカイコにおいてクロマチン制御機構が他生物と異なる可能性が示唆されたため,本年度はカイコHP1アイソフォームの機能的な相違に着目しながら,カイコにおけるクロマチン制御機構についてより詳細に解析を試みた. 前年度構築した転写抑制効果測定系においてレポーター遺伝子のプロモーターより活性の強いものに変えた場合における転写抑制効果を検証した.HP1αにおいては転写抑制効果が認められなかったが,HP1βにおいては明らかな転写抑制が観察された.前年度の結果と一致するものであり,カイコのHP1はアイソフォームにより局所的な転写抑制活性が異なることが明らかとなった.HP1αは局所的な転写抑制活性は弱いものの,ヘテロクロマチンの拡張に関わるタンパク質との強い相互作用を有しており,構成的なヘテロクロマチン形成に関わると考えられる.両者は互いに相互作用することから,制御領域における両者の相対比による転写抑制活性を変化させ,様々な遺伝子の転写レベルを調整していることが推測された.今回の研究により,HP1アイソフォームの機能的な相違が明らかとなり,昆虫におけるヘテロクロマチン形成機構の基礎的理解が進んだと考えられる.人為的に局所的ヘテロクロマチン形成の阻害あるいはユークロマチンへの誘導を促進することが可能となれば,組換え効率の向上が期待でき,ひいてはジーンターゲティングの効率を向上させることが可能になると考えられる.