Profiling novel pharmacology of receptor complexes using Receptor-HIT

Johnstone, Elizabeth K. M.; Pfleger, Kevin D. G.

doi:10.1042/bst20201110

Cited by 2 publications

(1 citation statement)

References 64 publications

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“…This study aimed to provide further evidence for the existence of the AT 2 -B 2 heteromer in HEK293FT cells, using various bioluminescence resonance energy transfer (BRET)-based proximity assays including the Receptor-Heteromer Investigation Technology (Receptor-HIT) (25,26) and the NanoBRET ligand binding assay (27,28). Receptor-HIT, which has most commonly been applied to GPCRs (GPCR-HIT) (5,25,27,(29)(30)(31)(32), is an assay that enables detection and characterization of heteromers through ligand-dependent interaction with biomolecules (Figure 1).…”

Section: Introductionmentioning

confidence: 99%

Novel Pharmacology Following Heteromerization of the Angiotensin II Type 2 Receptor and the Bradykinin Type 2 Receptor

Johnstone

Ayoub²,

Hertzman³

et al. 2022

Front. Endocrinol.

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The angiotensin type 2 (AT2) receptor and the bradykinin type 2 (B2) receptor are G protein-coupled receptors (GPCRs) that have major roles in the cardiovascular system. The two receptors are known to functionally interact at various levels, and there is some evidence that the observed crosstalk may occur as a result of heteromerization. We investigated evidence for heteromerization of the AT2 receptor and the B2 receptor in HEK293FT cells using various bioluminescence resonance energy transfer (BRET)-proximity based assays, including the Receptor Heteromer Investigation Technology (Receptor-HIT) and the NanoBRET ligand-binding assay. The Receptor-HIT assay showed that Gαq, GRK2 and β-arrestin2 recruitment proximal to AT2 receptors only occurred upon B2 receptor coexpression and activation, all of which is indicative of AT2-B2 receptor heteromerization. Additionally, we also observed specific coupling of the B2 receptor with the Gαz protein, and this was found only in cells coexpressing both receptors and stimulated with bradykinin. The recruitment of Gαz, Gαq, GRK2 and β-arrestin2 was inhibited by B2 receptor but not AT2 receptor antagonism, indicating the importance of B2 receptor activation within AT2-B2 heteromers. The close proximity between the AT2 receptor and B2 receptor at the cell surface was also demonstrated with the NanoBRET ligand-binding assay. Together, our data demonstrate functional interaction between the AT2 receptor and B2 receptor in HEK293FT cells, resulting in novel pharmacology for both receptors with regard to Gαq/GRK2/β-arrestin2 recruitment (AT2 receptor) and Gαz protein coupling (B2 receptor). Our study has revealed a new mechanism for the enigmatic and poorly characterized AT2 receptor to be functionally active within cells, further illustrating the role of heteromerization in the diversity of GPCR pharmacology and signaling.

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Section: Introductionmentioning

confidence: 99%

Novel Pharmacology Following Heteromerization of the Angiotensin II Type 2 Receptor and the Bradykinin Type 2 Receptor

Johnstone

Ayoub²,

Hertzman³

et al. 2022

Front. Endocrinol.

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Synthesis and Pharmacological Characterization of Fluorescent Ligands Targeting the Angiotensin II Receptors Derived from Agonists, β-Arrestin-Biased Agonists, and Antagonists

Delaitre,

Boisbrun,

Acherar

et al. 2024

J. Med. Chem.

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Angiotensin II (AngII) regulates cerebral circulation and binds with a similar affinity to AT 1 and AT 2 receptors. Biased AT 1 agonists, such as TRV027, which are able to selectively activate β-arrestin while blocking the G q pathway, appear promising as new therapeutics. New pharmacological tools are needed to further explore the impact of biased AT 1 agonists on cells or tissues, such as the cerebral vessels. We designed and synthesized new fluorescent derivatives based on AngII, TRV027, or the AT 1 antagonist losartan. We conducted pharmacological characterization to determine their selectivity, potency, and ability to activate or not specific AT 1 transduction pathways in cells and cerebral arteries. We report the first highly AT 1 -selective fluorescent ligand, based on losartan, that retains its antagonist activity with high affinity. Fluorescent derivatives of TRV027 become AT 2 -selective and lose their AT 1 β-arrestin bias. These new ligands can be applied to trace AT 1 or AT 2 receptors in vitro and ex vivo.

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Profiling novel pharmacology of receptor complexes using Receptor-HIT

Cited by 2 publications

References 64 publications

Novel Pharmacology Following Heteromerization of the Angiotensin II Type 2 Receptor and the Bradykinin Type 2 Receptor

Novel Pharmacology Following Heteromerization of the Angiotensin II Type 2 Receptor and the Bradykinin Type 2 Receptor

Synthesis and Pharmacological Characterization of Fluorescent Ligands Targeting the Angiotensin II Receptors Derived from Agonists, β-Arrestin-Biased Agonists, and Antagonists

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