Proteomic Analysis of β1-Adrenergic Receptor Interactions with PDZ Scaffold Proteins

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 β 1 -adrenergic receptor (β AR) interactions with PDZ scaffolds, the β 1 AR carboxyl terminus was screened against a newly developed proteomic array of PDZ domains. These screens confirmed β 1 AR associations with several previously identified PDZ partners, such as PSD-95, MAGI-2, GIPC, and CAL. Moreover, two novel β 1 AR-interacting proteins, SAP97 and MAGI-3, were also identified. The β 1 AR 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 β 1 AR being the key determinants of the interaction. Full-length β 1 AR robustly associated with full-length MAGI-3 in cells, and this association was abolished by mutation of the β 1 AR terminal valine residue to alanine (V477A), as determined by co-immunoprecipitation experiments and immunofluorescence co-localization studies. MAGI-3 co-expression with β 1 AR profoundly impaired β 1 AR-mediated ERK1/2 activation but had no apparent effect on β 1 AR-mediated cyclic AMP generation or agonist-promoted β 1 AR internalization. These findings revealed that the interaction of MAGI-3 with β 1 AR can selectively regulate specific aspects of receptor signaling. Moreover, the screens of the PDZ domain proteomic array provide a comprehensive view of β 1 AR interactions with PDZ scaffolds, thereby shedding light on the molecular mechanisms by which β 1 AR signaling and trafficking can be regulated in a cell-specific manner.