A two-state model for the diffusion of the a2A adenosine receptor in hippocampal neurons: Agonist-induced switch to slow mobility is modified by synapse-associated protein 102 (SAP102)
J. Biol. Chem.. 2014-02-07; 289(13): 9263-9274
DOI: 10.1074/jbc.M113.505685
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The A2A receptor is a class A/rhodopsin-like G protein-coupled receptor. Coupling
to its cognate protein, Gs, occurs via restricted collision coupling and is
contingent on the presence of cholesterol. Agonist activation slows diffusion of
the A2A 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 A2A 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
A2A receptor C terminus. Complex formation between the A2A 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 A2A
receptor in the compartment with slow mobility. Overexpression of SAP102
precluded the access of the A2A receptor to a compartment with restricted
mobility. In contrast, a mutated A2A 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 A2A receptor. The extended carboxyl terminus allows for
regulatory input by scaffolding molecules such as SAP102.