Abstract
Receptor heteromerization is the formation of a complex involving at least two different receptors with pharmacology that is distinct from that exhibited by its constituent receptor units. Detection of these complexes and monitoring their pharmacology is crucial for understanding how receptors function. The Receptor-Heteromer Investigation Technology (Receptor-HIT) utilizes ligand-dependent modulation of interactions between receptors and specific biomolecules for the detection and profiling of heteromer complexes. Previously, the interacting biomolecules used in Receptor-HIT assays have been intracellular proteins, however in this study we have for the first time used bioluminescence resonance energy transfer (BRET) with fluorescently-labeled ligands to investigate heteromerization of receptors on the cell surface. Using the Receptor-HIT ligand binding assay with NanoBRET, we have successfully investigated heteromers between the angiotensin II type 1 (AT1) receptor and the β2 adrenergic receptor (AT1-β2AR heteromer), as well as between the AT1 and angiotensin II type 2 receptor (AT1-AT2 heteromer).
Highlights
Receptor heteromerization is the formation of a complex composed of two functional receptor units
In order to address this issue, we have developed the Receptor-Heteromer Investigation Technology (ReceptorHIT) [2,3], which has been used to investigate several G protein-coupled receptor (GPCR)
We have demonstrated the Receptor-HIT assay utilizing NanoBRET ligand binding to investigate two GPCR heteromers: between the angiotensin II type 1 (AT1 ) receptor and the β2 adrenergic receptor (AT1 -β2 AR heteromer), as well as between the AT1 and angiotensin II type 2 receptor (AT1 -AT2 heteromer)
Summary
Receptor heteromerization is the formation of a complex composed of two (or more) functional receptor units. The constituent receptors are functional in their own right as monomers/homomers, the formation of a heteromer results in a complex that may have distinct biochemical properties [1]. This attainment of novel heteromer pharmacology expands the complexity of receptor signaling networks and adds selectivity and specificity to receptor signaling. This makes receptor heteromers exciting potential drug targets, to date only modest progress has been made towards therapeutically targeting these complexes. As we haveaspreviously viously discussed [17], Receptor-HIT can betoconfigured investigateatheteromers discussed [17], can be configured investigatetoheteromers their ex- at their extracellular the use of aligand fluorescent the labeled interacting tracellular surface, withsurface, the usewith of a fluorescent as theligand labeledasinteracting biomoleculebiomolecule (Figure 1b). (Figure 1b)
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