Site-specific O-Glycosylation by Polypeptide N-Acetylgalactosaminyltransferase 2 (GalNAc-transferase T2) Co-regulates ?₁-Adrenergic Receptor N-terminal Cleavage.

ABSTRACT: The ?₁-adrenergic receptor (?₁AR) is a G protein-coupled receptor (GPCR) and the predominant adrenergic receptor subtype in the heart, where it mediates cardiac contractility and the force of contraction. Although it is the most important target for ?-adrenergic antagonists, such as ?-blockers, relatively little is yet known about its regulation. We have shown previously that ?₁AR undergoes constitutive and regulated N-terminal cleavage participating in receptor down-regulation and, moreover, that the receptor is modified by O-glycosylation. Here we demonstrate that the polypeptide GalNAc-transferase 2 (GalNAc-T2) specifically O-glycosylates ?₁AR at five residues in the extracellular N terminus, including the Ser-49 residue at the location of the common S49G single-nucleotide polymorphism. Using in vitro O-glycosylation and proteolytic cleavage assays, a cell line deficient in O-glycosylation, GalNAc-T-edited cell line model systems, and a GalNAc-T2 knock-out rat model, we show that GalNAc-T2 co-regulates the metalloproteinase-mediated limited proteolysis of ?₁AR. Furthermore, we demonstrate that impaired O-glycosylation and enhanced proteolysis lead to attenuated receptor signaling, because the maximal response elicited by the ?AR agonist isoproterenol and its potency in a cAMP accumulation assay were decreased in HEK293 cells lacking GalNAc-T2. Our findings reveal, for the first time, a GPCR as a target for co-regulatory functions of site-specific O-glycosylation mediated by a unique GalNAc-T isoform. The results provide a new level of ?₁AR regulation that may open up possibilities for new therapeutic strategies for cardiovascular diseases.

SUBMITTER: Goth CK 

PROVIDER: S-EPMC5377785 | biostudies-literature | 2017 Mar

REPOSITORIES: biostudies-literature