Directed Evolution of a Palette of Optogenetic and Chemi-Optogenetic Indicators for Multiplexed Imaging of Cellular Metabolism

用于细胞代谢多重成像的光遗传学和化学光遗传学指标调色板的定向进化

• 批准号: 19H05633
• 负责人: キャンベル ロバート.アール
• 金额: $ 128.96万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (S)
• 财政年份: 2019
• 资助国家: 日本
• 起止时间: 2019-06-26 至 2024-03-31
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-19H05633/
• 关键词: Protein Engineering; Fluorescence; Cell signalling; Microscopy; Metabolism; Cell Biology

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.

于2022财政年度，我们报告了过去几年在我们实验室开发的许多高性能基因编码和化学遗传生物传感器。例如，在发布到bioRxiv的一系列三个预印本（2022.12.27.522013; 2022.10.19.512892;和2022.08.30.505811）中报告了改进的绿色和红色遗传编码生物传感器，其针对细胞内和细胞外乳酸盐的成像进行了优化。为了证明我们的新生物传感器的实用性和重要性，我们与生物学家合作，他们使用它们来研究乳酸在内质网-线粒体相互作用中的作用（在Cell Reports中描述）。在其他工作中，我们描述了一个改进的基因编码的远红外荧光钙离子生物传感器在蛋白质科学。该出版物包括将新生物传感器应用于多重成像的几个合作和创新的例子。在化学指示剂方面，我们在《自然化学生物学》上报道了我们基于人工合成的Ca 2+和Na+螯合剂和绿色荧光蛋白的化学指示剂。除了已发表的作品，我们还在继续探索和进一步优化这种新设计。于2022财政年度，坎贝尔就KIBAN（S）研究举办了19场特邀研讨会，包括SPIE Photonics West的主题演讲、生物成像学会年会的主题演讲、巴黎FENS论坛的会议演讲，以及牛津大学的部门演讲。坎贝尔实验室的受训人员就KIBAN（S）项目做了许多额外的海报和口头报告。

项目成果

Towards the development of novel single fluorescent protein-based biosensors for sugar phosphates

致力于开发新型基于单一荧光蛋白的磷酸糖生物传感器

• 发表时间: 2022
• 作者: Karl Matthew Lin;Fu Chai;Yusuke Nasu;Takuya Terai;Robert Earl Campbell
• 通讯作者: Robert Earl Campbell

GFP と合成キレーターを組み合わせた ケミジェネティックカルシウムセンサーの親和性向上

提高结合 GFP 和合成螯合剂的化学遗传学钙传感器的亲和力

• 发表时间: 2022
• 作者: 今井渉世;Wenchao Zhu;寺井琢也;Robert E. Campbell
• 通讯作者: Robert E. Campbell

乳酸バイオセンサーの開発

乳酸生物传感器的研制

• 发表时间: 2022
• 作者: 那須雄介;Giang Le;針尾紗彩;上條由貴;Robert E. Campbell
• 通讯作者: Robert E. Campbell

Evolutionary molecular engineering for the discovery of substrate peptides for E3 ligases and the development of an ubiquitin biosensor

用于发现 E3 连接酶底物肽和开发泛素生物传感器的进化分子工程

• 发表时间: 2022
• 作者: Kenwa Tamagawa;Takuya Terai;Robert E. Campbell
• 通讯作者: Robert E. Campbell

Intelligent image-activated cell sorting based on complex intracellular morphology

基于复杂细胞内形态的智能图像激活细胞分选

• 发表时间: 2022
• 作者: Jeffrey Harmon;Justin Findinier;Natsumi Tiffany Ishii;Maik Herbig;Akihiro Isozaki;Arthur R. Grossman;and Keisuke Goda
• 通讯作者: and Keisuke Goda