A Two-state Model for the Diffusion of the A(2A) Adenosine Receptor in Hippocampal Neurons: AGONIST-INDUCED SWITCH TO SLOW MOBILITY IS MODIFIED BY SYNAPSE-ASSOCIATED PROTEIN 102 (SAP102)

The A(2A) receptor is a class A/rhodopsin-like G protein-coupled receptor. Coupling to its cognate protein, G(s), occurs via restricted collision coupling and is contingent on the presence of cholesterol. Agonist activation slows diffusion of the A(2A) adenosine receptor in the lipid bilayer. We explored the contribution of the hydrophobic core and of the extended C terminus by examining diffusion of quantum dot-labeled receptor variants in dissociated hippocampal neurons. Single particle tracking of the A(2A) receptor(1–311), which lacks the last 101 residues, revealed that agonist-induced confinement was abolished and that the agonist-induced decrease in diffusivity was reduced substantially. A fragment comprising the SH3 domain and the guanylate kinase domain of synapse-associated protein 102 (SAP102) was identified as a candidate interactor that bound to the A(2A) receptor C terminus. Complex formation between the A(2A) receptor and SAP102 was verified by coimmunoprecipitation and by tracking its impact on receptor diffusion. An analysis of all trajectories by a hidden Markov model was consistent with two diffusion states where agonist activation reduced the transition between the two states and, thus, promoted the accumulation of the A(2A) receptor in the compartment with slow mobility. Overexpression of SAP102 precluded the access of the A(2A) receptor to a compartment with restricted mobility. In contrast, a mutated A(2A) receptor (with (383)DVELL(387) replaced by RVRAA) was insensitive to the action of SAP102. These observations show that the hydrophobic core per se does not fully account for the agonist-promoted change in mobility of the A(2A) receptor. The extended carboxyl terminus allows for regulatory input by scaffolding molecules such as SAP102.