title: Proteomic analysis of beta1-adrenergic receptor interactions with PDZ scaffold proteins.
abstract: Many G protein-coupled receptors possess carboxyl-terminal motifs ideal for interaction with PDZ scaffold proteins, which can control receptor trafficking and signaling in a cell-specific manner. To gain a panoramic view of beta1-adrenergic receptor (beta AR) interactions with PDZ scaffolds, the beta1AR carboxyl terminus was screened against a newly developed proteomic array of PDZ domains. These screens confirmed beta1AR associations with several previously identified PDZ partners, such as PSD-95, MAGI-2, GIPC, and CAL. Moreover, two novel beta1AR-interacting proteins, SAP97 and MAGI-3, were also identified. The beta1AR carboxyl terminus was found to bind specifically to the first PDZ domain of MAGI-3, with the last four amino acids (E-S-K-V) of beta1AR being the key determinants of the interaction. Full-length beta1AR robustly associated with full-length MAGI-3 in cells, and this association was abolished by mutation of the beta1AR terminal valine residue to alanine (V477A), as determined by co-immunoprecipitation experiments and immunofluorescence co-localization studies. MAGI-3 co-expression with beta1AR profoundly impaired beta1AR-mediated ERK1/2 activation but had no apparent effect on beta1AR-mediated cyclic AMP generation or agonist-promoted beta1AR internalization. These findings revealed that the interaction of MAGI-3 with beta1AR can selectively regulate specific aspects of receptor signaling. Moreover, the screens of the PDZ domain proteomic array provide a comprehensive view of beta1AR interactions with PDZ scaffolds, thereby shedding light on the molecular mechanisms by which beta1 AR signaling and trafficking can be regulated in a cell-specific manner.
authors.affiliation: Department of Biochemistry and Molecular Biology, Capital University of Medical Sciences, Beijing 100054, China.
authors.forename: Junqi
authors.initials: J
authors.lastname: He
authors.forename: Michele
authors.initials: M
authors.lastname: Bellini
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Inuzuka
authors.forename: Jianguo
authors.initials: J
authors.lastname: Xu
authors.forename: Ying
authors.initials: Y
authors.lastname: Xiong
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Yang
authors.forename: Amanda M
authors.initials: AM
authors.lastname: Castleberry
authors.forename: Randy A
authors.initials: RA
authors.lastname: Hall
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D002352:Carrier Proteins / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D011485:Protein Binding; D040901:Proteomics / Q000379:methods*; D018342:Receptors, Adrenergic, beta-1 / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D002352:Carrier Proteins; C496469:MAGI3 protein, human; D008565:Membrane Proteins; D018342:Receptors, Adrenergic, beta-1; D043562:Receptors, G-Protein-Coupled; D000242:Cyclic AMP; D020928:Mitogen-Activated Protein Kinases