Abstract 570: Activin A Regulates SERCA2a Expression in Cardiomyocytes Through Ubiquitin-proteasome Mediated Degradation

Abstract

Sarco/endoplasmic reticulum Ca 2+ -ATPase 2a (SERCA2a) is a critical regulator of cardiac function whose expression is decreased in aging and heart failure (HF). Activin-A, a member of the TGFβ superfamily, has been shown to be a potent paracrine inhibitor of SERCA2a expression in cardiomyocytes (CM). However, the mechanism by which this occurs remains unclear. Here, we confirm that Activin-A decreases SERCA2a protein expression in isolated CMs (reductions in SERCA2a expression at 25ng/ml: 35±5%, p=0.02; 50ng/ml: 55±14%, p= 0.003; 100ng/ml: 56±8%, p.= 0.001; n=2-3/group, three replicates). Moreover, we show that increasing circulating Activin-A levels in vivo also decreases cardiac SERCA2a protein (reductions in SERCA2a expression at 1-25ng/ml: 22±12%, p=0.27; 25-50ng/ml: 63±14%, p=0.0015; >50ng/ml: 79±2%, p= 0.0002; n=4 per group). Using systematic gene set enrichment analysis of RNAseq data from hearts and CMs exposed to increased Activin-A, we identify the ubiquitin-proteasome system (UPS) as one of the most highly up-regulated pathways induced by Activin-A. With ubiquitination prediction modeling revealing multiple ubiquitin binding sites on SERCA2a, we hypothesized that Activin-A regulates SERCA2a expression through modulation of its UPS-mediated degradation. Indeed, inhibiting the proteasome with MG132 attenuated the inhibitory effects of Activin-A on CM SERCA2a expression. Although Activin-A did not directly affect proteasome activity in CMs, we found that it increased ubiquitination of SERCA2a, resulting in increased proteasome-mediated degradation. Additionally, targeted inhibition of the Activin type II receptors (ActRII) with a monoclonal ActRII blocking antibody reversed the effects of Activin-A on SERCA2a protein expression in isolated CMs in a dose-dependent fashion. Importantly, these effects of Activin-A/ActRII blockade on SERCA2a expression were recapitulated in vivo in a transverse aortic constriction model of HF, and further led to improvements in cardiac function. Collectively, these data identify a novel UPS-mediated mechanism by which Activin-A regulates CM SERCA2a expression, which warrants further investigation as a potential therapeutic approach to augmenting SERCA2a in HF.