Mapping the Interface of a GPCR Dimer: A Structural Model of the A2A Adenosine and D2 Dopamine Receptor Heteromer

Abstract

The A2A adenosine (A2AR) and D2 dopamine (D2R) receptors form oligomers in the cell membrane and allosteric interactions across the A2AR–D2R heteromer represent a target for development of drugs against central nervous system disorders. However, understanding of the molecular determinants of A2AR–D2R heteromerization and the allosteric antagonistic interactions between the receptor protomers is still limited. In this work, a structural model of the A2AR–D2R heterodimer was generated using a combined experimental and computational approach. Regions involved in the heteromer interface were modeled based on the effects of peptides derived from the transmembrane (TM) helices on A2AR–D2R receptor–receptor interactions in bioluminescence resonance energy transfer (BRET) and proximity ligation assays. Peptides corresponding to TM-IV and TM-V of the A2AR blocked heterodimer interactions and disrupted the allosteric effect of A2AR activation on D2R agonist binding. Protein–protein docking was used to construct a model of the A2AR–D2R heterodimer with a TM-IV/V interface, which was refined using molecular dynamics simulations. Mutations in the predicted interface reduced A2AR–D2R interactions in BRET experiments and altered the allosteric modulation. The heterodimer model provided insights into the structural basis of allosteric modulation and the technique developed to characterize the A2AR–D2R interface can be extended to study the many other G protein-coupled receptors that engage in heteroreceptor complexes.

Acute cocaine treatment enhances the antagonistic allosteric adenosine A2A-dopamine D2 receptor–receptor interactions in rat dorsal striatum without increasing significantly extracellular dopamine levels

“…It is established that there exist antagonistic adenosine A2A receptor (A2AR)-dopamine (DA) D2 receptor (D2R) interactions in higher-order A2AR-D2R heteroreceptor complexes in cellular models and in the rat dorsal striatum, as demonstrated with biochemical binding techniques using D2R radioligands and proximity ligation assay [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16]. This is also in agreement with early behavioral findings in hemiparkinsonian rats [17].…”

“…1 h after the acute cocaine or vehicle administration, the rats were sacrificed, and the dorsal striatum was dissected out and immediately frozen on dry ice and stored at − 80 °C. The brain membrane preparation (0.15 mg/ml) and [ 3 H]-raclopride competition assays with minor modifications were performed according to previously published methods [9,28].…”

Dimerization energetics of the G‐protein coupled bile acid receptor TGR5 from all‐atom simulations

“…[89][90][91]105,106] Furthermore, knowledge of hot spot residues that are specific for the interfaces could be utilized to disrupt the dimerization upon mutation to alanine to probe the physiological effect of TGR5 interactions via specific interfaces, as done before for other GPCRs. [106] Finally, we determined that Y111 3.51 plays an important role in the formation of the 4/5 interface. Y111 3.51 , which is part of the conserved (D/E)RY motif, [107] is involved markedly in the dimer formation.…”

Adenosine heteroreceptor complexes in the basal ganglia are implicated in Parkinson’s disease and its treatment

“…The structural determinants that decide if a receptor pair forms a heteromer or not exist in the receptor interface that mediates the allosteric receptor–receptor interaction (Woods et al 2005 ). A substantial amount of work has been devoted to identify the key residues involved, including the development of a model of the A2AR–D2R heterodimer (Borroto-Escuela et al 2018b ). It is accepted that the transmembrane helices in class A, B and C dimers play a significant role ( http://www.gpcr-hetnet.com ) (Borroto-Escuela et al 2014a ).…”

“…The formation of homo-and heteroreceptor complexes in a synaptic or extrasynaptic area of the plasma membrane is governed by several factors and especially by the density of the participating receptor protomers (Fuxe and Borroto-Escuela 2016 ). Another factor is the affinity of one receptor protomer for another protomer, which is related to the number of hot spots that can develop in the receptor interface (Borroto-Escuela et al 2018b ). The presence or absence of adapter proteins in the heteroreceptor complex can be a significant factor for determining the affinity that develops between two or more receptor protomers (Borroto-Escuela et al 2018b ).…”