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A high-performance genetically encoded fluorescent indicator for in vivo cAMP imaging

Author

Listed:
  • Liang Wang

    (Chinese Academy of Sciences)

  • Chunling Wu

    (PKU-IDG–McGovern Institute for Brain Research
    Chinese Academy of Sciences)

  • Wanling Peng

    (Chinese Academy of Sciences)

  • Ziliang Zhou

    (Sun Yat-sen University
    Affiliated Stomatology Hospital of Guangzhou Medical University)

  • Jianzhi Zeng

    (PKU-IDG–McGovern Institute for Brain Research)

  • Xuelin Li

    (PKU-IDG–McGovern Institute for Brain Research)

  • Yini Yang

    (PKU-IDG–McGovern Institute for Brain Research)

  • Shuguang Yu

    (Chinese Academy of Sciences)

  • Ye Zou

    (Kansas State University)

  • Mian Huang

    (Kansas State University)

  • Chang Liu

    (Chinese Academy of Sciences)

  • Yefei Chen

    (Chinese Academy of Sciences)

  • Yi Li

    (Huazhong University of Science and Technology)

  • Panpan Ti

    (Huazhong University of Science and Technology)

  • Wenfeng Liu

    (Chinese Academy of Sciences)

  • Yufeng Gao

    (Chinese Academy of Sciences)

  • Wei Zheng

    (Chinese Academy of Sciences)

  • Haining Zhong

    (Oregon Health and Science University)

  • Shangbang Gao

    (Huazhong University of Science and Technology)

  • Zhonghua Lu

    (Chinese Academy of Sciences)

  • Pei-Gen Ren

    (Chinese Academy of Sciences)

  • Ho Leung Ng

    (Kansas State University)

  • Jie He

    (Chinese Academy of Sciences)

  • Shoudeng Chen

    (Sun Yat-sen University
    Sun Yat-sen University)

  • Min Xu

    (Chinese Academy of Sciences)

  • Yulong Li

    (PKU-IDG–McGovern Institute for Brain Research)

  • Jun Chu

    (Chinese Academy of Sciences
    and Shenzhen Institute of Synthetic Biology
    Chinese Academy of Sciences)

Abstract

cAMP is a key second messenger that regulates diverse cellular functions including neural plasticity. However, the spatiotemporal dynamics of intracellular cAMP in intact organisms are largely unknown due to low sensitivity and/or brightness of current genetically encoded fluorescent cAMP indicators. Here, we report the development of the new circularly permuted GFP (cpGFP)-based cAMP indicator G-Flamp1, which exhibits a large fluorescence increase (a maximum ΔF/F0 of 1100% in HEK293T cells), decent brightness, appropriate affinity (a Kd of 2.17 μM) and fast response kinetics (an association and dissociation half-time of 0.20 and 0.087 s, respectively). Furthermore, the crystal structure of the cAMP-bound G-Flamp1 reveals one linker connecting the cAMP-binding domain to cpGFP adopts a distorted β-strand conformation that may serve as a fluorescence modulation switch. We demonstrate that G-Flamp1 enables sensitive monitoring of endogenous cAMP signals in brain regions that are implicated in learning and motor control in living organisms such as fruit flies and mice.

Suggested Citation

  • Liang Wang & Chunling Wu & Wanling Peng & Ziliang Zhou & Jianzhi Zeng & Xuelin Li & Yini Yang & Shuguang Yu & Ye Zou & Mian Huang & Chang Liu & Yefei Chen & Yi Li & Panpan Ti & Wenfeng Liu & Yufeng Ga, 2022. "A high-performance genetically encoded fluorescent indicator for in vivo cAMP imaging," Nature Communications, Nature, vol. 13(1), pages 1-16, December.
  • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-32994-7
    DOI: 10.1038/s41467-022-32994-7
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    References listed on IDEAS

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    1. Stephen X. Zhang & Andrew Lutas & Shang Yang & Adriana Diaz & Hugo Fluhr & Georg Nagel & Shiqiang Gao & Mark L. Andermann, 2021. "Hypothalamic dopamine neurons motivate mating through persistent cAMP signalling," Nature, Nature, vol. 597(7875), pages 245-249, September.
    2. Yuki Oe & Xiaowen Wang & Tommaso Patriarchi & Ayumu Konno & Katsuya Ozawa & Kazuko Yahagi & Hirokazu Hirai & Takashi Tsuboi & Tetsuya Kitaguchi & Lin Tian & Thomas J. McHugh & Hajime Hirase, 2020. "Author Correction: Distinct temporal integration of noradrenaline signaling by astrocytic second messengers during vigilance," Nature Communications, Nature, vol. 11(1), pages 1-1, December.
    3. Yuki Oe & Xiaowen Wang & Tommaso Patriarchi & Ayumu Konno & Katsuya Ozawa & Kazuko Yahagi & Hirokazu Hirai & Takashi Tsuboi & Tetsuya Kitaguchi & Lin Tian & Thomas J. McHugh & Hajime Hirase, 2020. "Distinct temporal integration of noradrenaline signaling by astrocytic second messengers during vigilance," Nature Communications, Nature, vol. 11(1), pages 1-15, December.
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