Abstract 13101: Mst1-Mediated Phosphorylation of Beclin1 in its BH3 Domain Promotes Beclin1-Bcl-2 Interaction, Thereby Negatively Regulating Autophagy in the Heart
Inhibition of autophagy below physiological levels deteriorates protein quality control (QC) and cellular function. Beclin1, a key mediator of autophagy, is negatively regulated by Bcl-2. Here we show that the Beclin1-Bcl-2 interaction is regulated by mammalian Ste20-like kinase 1 (Mst1) in the heart. In Mst1 transgenic mice (Tg-Mst1), a model of dilated cardiomyopathy, both aggresomes and p62, a protein degraded by autophagy, were accumulated and co-localized, and autophagosome formation, as evaluated by GFP-LC3 puncta, was suppressed in cardiomyocytes (CMs). Mst1 activity was suppressed by starvation in CMs, and restoration of Mst1 activity abolished starvation-induced autophagy. Mst1 inhibited the Beclin1-Vps34 complex kinase activity. Treatment with ABT-737, a BH3 mimetic compound, or knockdown of Bcl-2 reversed the Mst1-induced suppression of autophagy, suggesting that suppression of autophagy by Mst1 is both BH3 domain- and Bcl-2-dependent. Mst1 physically interacts with Beclin1 and enhances the Beclin1-Bcl-2 interaction in CMs. Mass spectrometry analysis revealed that Mst1 phosphorylates Beclin1 at Thr108 located in its BH3 domain, a binding site of Bcl-2. Transduction with adenovirus harboring Beclin1 Thr108Asp (Ad-T108D) attenuated starvation-induced autophagy in CMs, and injection with adenovirus harboring Beclin1 Thr108Ala (Ad-T108A), but not Ad-T108D, restored starvation-induced LV dysfunction in beclin1-/+ mice. The role of Mst1-Beclin1 interaction in response to chronic myocardial infarction (CMI) was evaluated with Tg-DN-Mst1. Inhibition of Mst1 attenuated LV dysfunction during CMI (%FS; Tg-DN-Mst1: 21.6±0.7*, NTg: 15.6±0.6, *p<0.05). The amount of phosphorylated Beclin1 and co-localization of aggresomes and p62 were significantly decreased, and the number of LC3 puncta was significantly increased in Tg-DN-Mst1 compared to NTg (Counts/HPF, 188±13*, 79±5, *p<0.05). Collectively, these results suggest that Mst1 inhibits autophagy through phosphorylation of Beclin1, enhancement of Beclin1-Bcl-2 interaction and suppression of Vps34. Activation of Mst1 by stress suppresses autophagy below physiological levels and inhibits protein QC, which in turn contributes to cardiac dysfunction.
- © 2011 by American Heart Association, Inc.