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Visualization of the activation of the histamine H3 receptor (H3R) using novel fluorescence resonance energy transfer biosensors and their potential application to the study of H3R pharmacology
Author(s) -
Liu Ying,
Zeng Hong,
Pediani John D.,
Ward Richard J.,
Chen LuYao,
Wu Nan,
Ma Li,
Tang Mei,
Yang Yang,
An Su,
Guo XiaoXi,
Hao Qian,
Xu TianRui
Publication year - 2018
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.14484
Subject(s) - förster resonance energy transfer , yellow fluorescent protein , histamine h3 receptor , agonist , chemistry , biophysics , receptor , biology , fluorescence , biochemistry , physics , quantum mechanics , gene
Activation of the histamine‐3 receptor (H3R) is involved in memory processes and cognitive action, while blocking H3R activation can slow the progression of neurological disorders, such as Alzheimer's disease, schizophrenia and narcolepsy. To date, however, no direct way to examine the activation of H3R has been utilized. Here, we describe a novel biosensor that can visualize the activation of H3R through an intramolecular fluorescence resonance energy transfer ( FRET ) signal. To achieve this, we constructed an intramolecular H3R FRET sensor with cyan fluorescent protein ( CFP ) attached at the C terminus and yellow fluorescent protein ( YFP ) inserted into the third intracellular loop. The sensor was found to internalize normally on agonist treatment. We measured FRET signals between the donor CFP and the acceptor YFP in living cells in real time, the results of which indicated that H3R agonist treatment (imetit or histamine) increases the FRET signal in a time‐ and concentration‐dependent manner with Kon and Koff values consistent with published data and which maybe correlated with decreasing cAMP levels and the promotion of ERK 1/2 phosphorylation. The FRET signal was inhibited by H3R antagonists, and the introduction of mutations at F419A, F423A, L426A and L427A, once again, the promotion of ERK 1/2 phosphorylation, was diminished. Thus, we have built a H3R biosensor which can visualize the activation of receptor through real‐time structure changes and which can obtain pharmacological kinetic data at the same time. The FRET signals may allow the sensor to become a useful tool for screening compounds and optimizing useful ligands.