GalNAc-transferase 2 deficiency protects the heart from sustained beta-1 adrenergic receptor stimulation

Abstract Introduction The β1-adrenergic receptor (β1AR) is a G protein-coupled receptor (GPCR) that regulates the sympathetic activation of the heart. We have previously demonstrated that the β1AR extracellular N-terminal domain is post-translationally modified by O-glycosylation and proteolytic cleavage both in heterologous expression systems as well as in the rat heart (1). We have also shown that one of the transferases that initiates O-glycosylation, N-acetylgalactosamine-transferase 2 (GalNAc-T2), specifically O-glycosylates β1AR (1). GalNAc-T2-mediated O-glycosylation was found to protect the receptor from proteolytic processing, whereas impaired O-glycosylation led to enhanced receptor cleavage that decreased β1AR signaling. Purpose The aim of this study was to investigate how GalNAc-T2-mediated O-glycosylation modulates the function of β1AR in the heart in vitro and in vivo. For this, we utilized a Galnt2 knockout (KO) rat model. We hypothesized that attenuated signaling of β1AR in the Galnt2 KO rats could diminish the pathological remodeling of the heart. Methods Isolated cardiomyocytes were used to study changes in β1AR signaling and contractile function in vitro, assessed with Western blot and cell shortening assay. In the in vivo experiments, male Galnt2 KO rats and their wild-type littermates, aged 9–11 weeks, were treated with βAR agonist isoproterenol (ISO) via osmotic minipumps at a dose of 5 mg/kg/day for 10 days. Assessment of cardiac function, hypertrophy and ventricular remodeling was performed using transthoracic echocardiography, staining of histological cardiac sections and qPCR analysis of left ventricular samples. Results A significant attenuation of ISO-mediated phosphorylation of troponin I was observed in the isolated cardiomyocytes of Galnt2 KO rats. In addition, Galnt2 KO cardiomyocytes showed attenuated contractility under ISO stimulation in a cell shortening assay. In line with these results, the echocardiography analysis demonstrated that Galnt2 deficiency significantly attenuated ISO-induced contractility and hypertrophy. This was associated with reduced hypertrophy and fibrosis in the histological analysis of cardiac sections as well as with the diminished expression of corresponding genes in the qPCR analysis. Conclusions The lack of GalNAc-T2-mediated O-glycosylation of β1AR and subsequent enhancement in receptor cleavage leads to impaired β1AR signaling in the rat heart, resulting in attenuated pathological remodeling under sympathetic cardiac stress. Targeting cardiac GalNAc-T2 may offer a way to protect the heart from sustained sympathetic stimulation in cardiac diseases, and thus this study can aid the development of pharmacological therapies.