Functional Impact of the G279S Substitution in the Adenosine A 1 -Receptor (A 1 R-G279S 7.44 ), a Mutation Associated with Parkinson's Disease.

In medium-size, spiny striatal neurons of the direct pathway, dopamine D- and adenosine A-receptors are coexpressed and are mutually antagonistic. Recently, a mutation in the gene encoding the A-receptor (AR), AR-G279S, was identified in an Iranian family: two affected offspring suffered from early-onset l-DOPA-responsive Parkinson's disease. The link between the mutation and the phenotype is unclear. Here, we explored the functional consequence of the G279S substitution on the activity of the A-receptor after heterologous expression in HEK293 cells. The mutation did not affect surface expression and ligand binding but changed the susceptibility to heat denaturation: the thermodynamic stability of AR-G279S was enhanced by about 2 and 8 K when compared with wild-type A-receptor and AR-Y288A (a folding-deficient variant used as a reference), respectively. In contrast, the kinetic stability was reduced, indicating a lower energy barrier for conformational transitions in AR-G279S (73 ± 23 kJ/mol) than in wild-type AR (135 ± 4 kJ/mol) or in AR-Y288A (184 ± 24 kJ/mol). Consistent with this lower energy barrier, AR-G279S was more effective in promoting guanine nucleotide exchange than wild-type AR. We detected similar levels of complexes formed between D-receptors and wild-type AR or AR-G279S by coimmunoprecipitation and bioluminescence resonance energy transfer. However, lower concentrations of agonist were required for half-maximum inhibition of dopamine-induced cAMP accumulation in cells coexpressing D-receptor and AR-G279S than in those coexpressing wild-type AR. These observations predict enhanced inhibition of dopaminergic signaling by AR-G279S in vivo, consistent with a pathogenic role in Parkinson's disease. SIGNIFICANCE STATEMENT: Parkinson's disease is caused by a loss of dopaminergic input from the substantia nigra to the caudate nucleus and the putamen. Activation of the adenosine A-receptor antagonizes responses elicited by dopamine D-receptor. We show that this activity is more pronounced in a mutant version of the A-receptor (AR-G279S), which was identified in individuals suffering from early-onset Parkinson's disease.