title: Role of AKAP79/150 protein in β1-adrenergic receptor trafficking and signaling in mammalian cells.
abstract: Protein kinase A-anchoring proteins (AKAPs) participate in the formation of macromolecular signaling complexes that include protein kinases, ion channels, effector enzymes, and G-protein-coupled receptors. We examined the role of AKAP79/150 (AKAP5) in trafficking and signaling of the β1-adrenergic receptor (β1-AR). shRNA-mediated down-regulation of AKAP5 in HEK-293 cells inhibited the recycling of the β1-AR. Recycling of the β1-AR in AKAP5 knockdown cells was rescued by shRNA-resistant AKAP5. However, truncated mutants of AKAP5 with deletions in the domains involved in membrane targeting or in binding to calcineurin or PKA failed to restore the recycling of the β1-AR, indicating that full-length AKAP5 was required. Furthermore, recycling of the β1-AR in rat neonatal cardiac myocytes was dependent on targeting the AKAP5-PKA complex to the C-terminal tail of the β1-AR. To analyze the role of AKAP5 more directly, recycling of the β1-AR was determined in ventricular myocytes from AKAP5(-/-) mice. In AKAP5(-/-) myocytes, the agonist-internalized β1-AR did not recycle, except when full-length AKAP5 was reintroduced. These data indicate that AKAP5 exerted specific and profound effects on β1-AR recycling in mammalian cells. Biochemical or real time FRET-based imaging of cyclic AMP revealed that deletion of AKAP5 sensitized the cardiac β1-AR signaling pathway to isoproterenol. Moreover, isoproterenol-mediated increase in contraction rate, surface area, or expression of β-myosin heavy chains was significantly greater in AKAP5(-/-) myocytes than in AKAP5(+/+) myocytes. These results indicate a significant role for the AKAP5 scaffold in signaling and trafficking of the β1-AR in cardiac myocytes and mammalian cells.
authors.affiliation: Department of Pharmacology, University of Tennessee Health Sciences Center, Memphis, Tennessee 38163.
authors.forename: Xin
authors.initials: X
authors.lastname: Li
authors.affiliation: Department of Pharmacology, University of Tennessee Health Sciences Center, Memphis, Tennessee 38163.
authors.forename: Mohammed M
authors.initials: MM
authors.lastname: Nooh
authors.affiliation: Department of Pharmacology, University of Tennessee Health Sciences Center, Memphis, Tennessee 38163. Electronic address: sbahouth@uthsc.edu.
authors.forename: Suleiman W
authors.initials: SW
authors.lastname: Bahouth
mesh_terms: D054758:A Kinase Anchor Proteins / Q000235:genetics / Q000378:metabolism*; D058665:Adrenergic beta-1 Receptor Agonists / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D051379:Mice; D018345:Mice, Knockout; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D051381:Rats; D018342:Receptors, Adrenergic, beta-1 / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*
chemical_list: D054758:A Kinase Anchor Proteins; C487775:AKAP5 protein, human; D058665:Adrenergic beta-1 Receptor Agonists; C520665:Akap5 protein, mouse; C415703:Akap5 protein, rat; D018342:Receptors, Adrenergic, beta-1; D017868:Cyclic AMP-Dependent Protein Kinases; D018995:Myosin Heavy Chains; D007545:Isoproterenol
keywords: AKAP; Adrenergic Receptor; Cyclic AMP (cAMP); Fluorescence Resonance Energy Transfer (FRET); G-protein-coupled Receptors (GPCR); Protein Kinase A (PKA); Receptor Recycling; Trafficking