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

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 Gao, Wei Zheng, Haining Zhong, Shangbang Gao, Zhonghua LuPei Gen Ren, Ho Leung Ng, Jie He, Shoudeng Chen, Min Xu, Yulong Li, Jun Chu

Research output: Contribution to journalArticlepeer-review

11 Scopus citations

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.

Original languageEnglish (US)
Article number5363
JournalNature communications
Volume13
Issue number1
DOIs
StatePublished - Dec 2022

ASJC Scopus subject areas

  • General Physics and Astronomy
  • General Chemistry
  • General Biochemistry, Genetics and Molecular Biology

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