title: Confinement of β(1)- and β(2)-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.
abstract: The sympathetic nervous system regulates cardiac output by activating adrenergic receptors (ARs) in cardiac myocytes. The predominant cardiac ARs, β(1)- and β(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 βARs integrate into complexes in live cells, we measured receptor diffusion and interactions by single-particle tracking. Unstimulated β(1)- and β(2)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 β(2)AR did not alter receptor dynamics, suggesting that these membrane domains are not involved in β(2)AR confinement. For both β(1)- and β(2)AR, the receptor carboxy-terminus was uniquely responsible for scaffold interactions. Our data formally demonstrate that distinct and stable protein complexes containing β(1)- or β(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.
authors.affiliation: Division of Nephrology, Department of Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
authors.forename: Cathleen D
authors.initials: CD
authors.lastname: Valentine
authors.forename: Peter M
authors.initials: PM
authors.lastname: Haggie
mesh_terms: D054758:A Kinase Anchor Proteins / Q000378:metabolism*; D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000818:Animals; D021941:Caveolae / Q000378:metabolism*; D002460:Cell Line; D002522:Chlorocebus aethiops; D036681:Fluorescence Recovery After Photobleaching; D006801:Humans; D008856:Microscopy, Fluorescence; D032383:Myocytes, Cardiac / Q000378:metabolism*; D054731:PDZ Domains; D010446:Peptide Fragments / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D018342:Receptors, Adrenergic, beta-1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018343:Receptors, Adrenergic, beta-2 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D059010:Single-Cell Analysis; D059008:Time-Lapse Imaging
chemical_list: D054758:A Kinase Anchor Proteins; D000199:Actins; D010446:Peptide Fragments; D018342:Receptors, Adrenergic, beta-1; D018343:Receptors, Adrenergic, beta-2