title: beta 1-adrenergic receptor association with PSD-95. Inhibition of receptor internalization and facilitation of beta 1-adrenergic receptor interaction with N-methyl-D-aspartate receptors.
abstract: The beta(1)-adrenergic receptor (beta(1)AR) is the most abundant subtype of beta-adrenergic receptor in the mammalian brain and is known to potently regulate synaptic plasticity. To search for potential neuronal beta(1)AR-interacting proteins, we screened a rat brain cDNA library using the beta(1)AR carboxyl terminus (beta(1)AR-CT) as bait in the yeast two-hybrid system. These screens identified PSD-95, a multiple PDZ domain-containing scaffolding protein, as a specific binding partner of the beta(1)AR-CT. This interaction was confirmed by in vitro fusion protein pull-down and blot overlay experiments, which demonstrated that the beta(1)AR-CT binds specifically to the third PDZ domain of PSD-95. Furthermore, the full-length beta(1)AR associates with PSD-95 in cells, as determined by co-immunoprecipitation experiments and immunofluorescence co-localization studies. The interaction between beta(1)AR and PSD-95 is mediated by the last few amino acids of the beta(1)AR, and mutation of the beta(1)AR carboxyl terminus eliminated the binding and disrupted the co-localization of the beta(1)AR and PSD-95 in cells. Agonist-induced internalization of the beta(1)AR in HEK-293 cells was markedly attenuated by PSD-95 co-expression, whereas co-expression of PSD-95 has no significant effect on either desensitization of the beta(1)AR or beta(1)AR-induced cAMP accumulation. Furthermore, PSD-95 facilitated the formation of a complex between the beta(1)AR and N-methyl-d-aspartate receptors, as assessed by co-immunoprecipitation. These data reveal that PSD-95 is a specific beta(1)AR binding partner that modulates beta(1)AR function and facilitates physical association of the beta(1)AR with synaptic proteins, such as the N-methyl-d-aspartate receptors, which are known to be regulated by beta(1)AR stimulation.
authors.affiliation: Howard Hughes Medical Institute, Departments of Medicine and Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: L A
authors.initials: LA
authors.lastname: Hu
authors.forename: Y
authors.initials: Y
authors.lastname: Tang
authors.forename: W E
authors.initials: WE
authors.lastname: Miller
authors.forename: M
authors.initials: M
authors.lastname: Cong
authors.forename: A G
authors.initials: AG
authors.lastname: Lau
authors.forename: R J
authors.initials: RJ
authors.lastname: Lefkowitz
authors.forename: R A
authors.initials: RA
authors.lastname: Hall
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D000076122:Disks Large Homolog 4 Protein; D015723:Gene Library; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D051381:Rats; D018342:Receptors, Adrenergic, beta-1 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016194:Receptors, N-Methyl-D-Aspartate / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D014162:Transfection
chemical_list: D000076122:Disks Large Homolog 4 Protein; C501449:Dlg4 protein, rat; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010446:Peptide Fragments; D018342:Receptors, Adrenergic, beta-1; D016194:Receptors, N-Methyl-D-Aspartate; D011993:Recombinant Fusion Proteins; C033652:postsynaptic density proteins