2022 Fiscal Year Annual Research Report
Directed Evolution of a Palette of Optogenetic and Chemi-Optogenetic Indicators for Multiplexed Imaging of Cellular Metabolism
| Project/Area Number |
19H05633
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| Research Institution | The University of Tokyo |
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Principal Investigator |
キャンベル ロバート.アール 東京大学, 大学院理学系研究科(理学部), 教授 (40831318)
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| Co-Investigator(Kenkyū-buntansha) |
合田 圭介 東京大学, 大学院理学系研究科(理学部), 教授 (70518696)
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| Project Period (FY) |
2019-06-26 – 2024-03-31
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| Keywords | Protein Engineering / Fluorescence / Microscopy / Cell Biology |
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| Outline of Annual Research Achievements |
In FY2022 we reported many of the high performance genetically encoded and chemigenetic biosensors that had been under development in our lab over the past few years. For example, improved green and red genetically encoded biosensors, optimized for imaging of intra- and extracellular lactate, were reported in a series of three preprints posted to bioRxiv (2022.12.27.522013; 2022.10.19.512892; and 2022.08.30.505811). To demonstrate the utility and importance of our new biosensors, we collaborated with biologists who used them to investigate the role of lactate in endoplasmic reticulum-mitochondria interactions (described in Cell Reports). In other work we described an improved genetically-encoded far-red fluorescent Ca2+ biosensor in Protein Science. This publications included several collaborative and innovative examples of applying the new biosensor to multiplexed imaging. In the area of chemigenetic indicators, we reported our chemigenetic indicators based on synthetic Ca2+ and Na+ chelators and green fluorescent protein in Nature Chemical Biology. Beyond the published work, we are continuing to explore and further optimize this new design. During FY2022, Campbell gave 19 invited seminars on KIBAN(S) research, including a keynote at SPIE Photonics West, a keynote at the Annual meeting of the Bioimaging society, a conference presentation at the FENS Forum in Paris, and a Departmental presentation at Oxford University. Trainees in the Campbell lab gave many additional poster and oral presentations on KIBAN(S) projects.
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| Current Status of Research Progress |
Current Status of Research Progress
1: Research has progressed more than it was originally planned.
Reason
During FY2022 we made tremendous progress in the area of genetically encoded biosensors for key metabolites. In a series of preprints now available on bioRxiv, we report a 1st generation red fluorescent biosensor for extracellular lactate, a 1st generation green fluorescent biosensor for intracellular lactate, a 1st generation red biosensor for intracellular lactate, and a 2nd generation biosensor for extracellular lactate. In unreported work we have made tremendous progress towards 2nd and 3rd generation biosensors.
In the area of genetically encoded Ca2+ biosensors, we reported a new far-red biosensor, developed a 3rd generation near-infrared (NIR) biosensor, and developed a novel mNeonGreen-based biosensor that can be used to image Ca2+ concentration changes at membrane-membrane contact sites.
In the area of chemigenetic biosensors, we made tremendous progress in FY2022 and successfully published our work in Nature Chemical Biology.
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| Strategy for Future Research Activity |
In FY2023 we will continue to work towards establishing our series of high performance green and red biosensors for intra- and extra-cellular lactate as the definitive biosensors in this area. We will work with more than 30 collaborators in more than 10 countries to apply these new lactate biosensors for multiplexed imaging of intra- and extra-cellular lactate concentrations in various model systems. Beyond lactate, we have now initiated work on a variety of additional metabolite targets include acetyl CoA, pyruvate, and fumarate. These efforts are currently at an early “advanced prototype” stage, but we expect to make substantial progress in FY2023. In the area of genetically encoded Ca2+ biosensors, we will continue to improve our 3rd generation NIR bios
ensor and work with collaborators to assesses its utility for in vivo imaging of neural activity. We have recently acquired a colony picking robot that will drastically increase our throughput and accelerate our progress.
In the area of chemigenetic biosensors, we will continue to build upon our success to date and develop a broader range of chemigenetic sensors. Specifically, we will work towards expanding the ion specificity and color palette of chemigenetic indicators bases on synthetic chelators and fluorescent proteins.
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[Presentation] Uncovering the long-term effects of mRNA COVID-19 vaccinations on platelets by high-speed optical imaging on a chip2023
Author(s)
Y. Zhou, M. Nishikawa, H. Kanno, R. Yang, Y. Ibayashi, T.-H. Xiao, W. Peterson, M. Herbig, N. Nitta, S. Miyata, Y. Kanthi, G. K. Rohde, K. Moriya, Y. Yatomi, K. Goda
Organizer
SPIE Photonics West 2023
Int'l Joint Research
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[Presentation] 血小板凝集塊の無標識画像ヒックテータによる COVID-19 と血栓症のリアルタイム AI 分類2023
Author(s)
張 晨祺, Herbig Maik, 周 雨奇, 西川 真子, Shifat-E-Rabbi Mohammad, 菅野 寛志 , 楊 若曦 , 伊林 侑真 , 肖 廷輝 , Rohde Gustavo, 佐藤 将敬 , 小寺 聡, 大門 雅夫, 矢冨 裕, 合田 圭介
Organizer
第70回 応用物理学会 春季学術講演会
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[Presentation] Machine learning-based sort-timing prediction for image-activated cell sorting2022
Author(s)
Yaqi Zhao, Akihiro Isozaki, Maik Herbig, Mika Hayashi, Kotaro Hiramatsu, Sota Yamazaki, Naoko Kondo, Shinsuke Ohnuki, Yoshikazu Ohya, Nao Nitta, and Keisuke Goda
Organizer
化学とマイクロ・ナノシステム学会第45回研究会
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[Presentation] Detection and characterization of circulating tumor cell clusters by high-throughput optomechanical imaging flow cytometry2022
Author(s)
Hiroki Matsumura, Larina Shen, Akihiro Isozaki, Hideharu Mikami, Dan Yuan, Maik Herbig, Yuto Kondo, Tomoko Mori, Yoshika Kusumoto, Masako Nishikawa, Yutaka Yatomi, Satoshi Matsusaka, Keisuke Goda
Organizer
CYTO 2022
Int'l Joint Research
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[Presentation] A standardized metric for throughput for cytometry2022
Author(s)
Maik Herbig, Akihiro Isozaki, Dino Di Carlo, Jochen Guck, Nao Nitta, Robert Damoiseaux, Shogo Kamikawaji, Eigo Suyama, Hirofumi Shintaku, Angela Ruohao Wu, Itoshi Nikaido, and Keisuke Goda
Organizer
CYTO 2022
Int'l Joint Research
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[Presentation] Resolution-enhanced imaging flow cytometry by deep learning2022
Author(s)
Maik Herbig, Kangrui Huang, Hiroki Matsumura, Yaqi Zhao, Dan Yuan, Yohei Mineharu, Jeffrey Harmon, Justin Findinier, Mai Yamagishi, Shinsuke Ohnuki, Nao Nitta, Arthur R. Grossman, Yoshikazu Ohya, Hideharu Mikami, Akihiro Isozaki, and Keisuke Goda
Organizer
CYTO 2022
Int'l Joint Research
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[Presentation] Uncovering the pathogenesis of microvascular thrombosis in COVID-19 via high-throughput microscopy on a chip2022
Author(s)
Y. Zhou, M. Nishikawa, H. Kanno, T.-H. Xiao, C. Zhang, M. Herbig, Y. Ibayashi, N. Nitta, S. Miyata, G. K. Rohde, Y. Yatomi, K. Goda
Organizer
CYTO 2022
Int'l Joint Research
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