Confinement of β1- and β2-adrenergic receptors in the plasma membrane of cardiomyocyte-like H9c2 cells is mediated by selective interactions with PDZ domain and A-kinase anchoring proteins but not caveolae

The sympathetic nervous system regulates cardiac output by activating adrenergic receptors (ARs) in cardiac myocytes. The predominant cardiac ARs, beta(1)- and beta(2)AR, are structurally similar but mediate distinct signaling responses. Scaffold protein-mediated compartmentalization of ARs into discrete, multiprotein complexes has been proposed to dictate differential signaling responses. To test the hypothesis that beta ARs integrate into complexes in live cells, we measured receptor diffusion and interactions by single-particle tracking. Unstimulated beta 1- and beta AR were highly confined in the membrane of H9c2 cardiomyocyte-like cells, indicating that receptors are tethered and presumably integrated into protein complexes. Selective disruption of interactions with postsynaptic density protein 95/disks large/zonula occludens-1 (PDZ)-domain proteins and A-kinase anchoring proteins (AKAPs) increased receptor diffusion, indicating that these scaffold proteins participate in receptor confinement. In contrast, modulation of interactions between the putative scaffold caveolae and beta(2)AR did not alter receptor dynamics, suggesting that these membrane domains are not involved in beta(2)AR confinement. For both beta(1)- and beta(2)AR, the receptor carboxy-terminus was uniquely responsible for scaffold interactions. Our data formally demonstrate that distinct and stable protein complexes containing beta(1)- or beta(2)AR are formed in the plasma membrane of cardiomyocyte-like cells and that selective PDZ and AKAP interactions are responsible for the integration of receptors into complexes.