Proteasome malfunction increases autophagosomes in cardiomyocytes and mouse hearts

Abstract

Background:Autophagy and the ubiquitin‐proteasome system (UPS) are two main routes for intracellular protein degradation. The effect of proteasome malfunction on autophagy in cardiomyocytes and the heart remains to be demonstrated.Methods & results:Proteasomal function in cultured neonatal ventricular myocytes (NRVMs) or mouse hearts is manipulated using pharmacological and/or genetic means. The resultant effects on autophagy are assessed by two methods: LC3‐II/LC‐I ratio measured by western blot analysis; autophagy index measured via confocal microscopic analysis of GFP‐LC3 distribution. Cardiac‐restricted expression of a 7‐amino‐acid deletion mutation of the desmin gene (D7‐des) has been linked to human desminopathy which develops aberrant desmin aggregation and severe proteasome malfunction in heart muscle cells. D7‐des increases LC3‐II/LC3‐I ratio and punctate GFP‐LC3 in mouse heart as early as 2 month. The increased autophagosomes is also detected in mouse heart 12 hours after an intravenous injection of proteasome inhibitor MG262 (5umol/kg). In NRVMs, proteasome inhibition by MG132 (1uM) increases the LC3‐II/LC3‐I ratio and autophagic flux measured as the difference of autophagosomes in the absence and presence of lysosomal protease inhibitors.Conclusions:Pharmacological and/or genetic proteasome inhibition increase autophagosomes in both NRVMs and mouse hearts.