Proteome Dynamics and Proteome Function of Cardiac 19S Proteasomes

Abstract

Myocardial proteasomes are comprised of 20S core particles and 19S regulatory particles, which together carry out targeted degradation of cardiac proteins. The 19S complex is unique among the regulators of proteasomes in that it affects both the capacity and specificity of protein degradation. However, a comprehensive molecular characterization of cardiac 19S complexes is lacking. In this investigation, we tailored a multidimensional chromatography-based purification strategy to isolate structurally intact and functionally viable 19S complexes from murine hearts. Two distinct subpopulations of 19S complexes were isolated based upon (1) potency of activating 20S proteolytic activity, and (2) molecular composition using a combination of immuno-detection, two-dimensional-differential gel electrophoresis, and MS-based approaches. Heat shock protein 90 (Hsp90) was identified to be characteristic to 19S subpopulation I. The physical interaction of Hsp90 with 19S complexes was demonstrated via multiple approaches. Inhibition of Hsp90 activity using geldanamycin or BIIB021 potentiated the ability of subpopulation I to activate 20S proteasomes in the murine heart, thus demonstrating functional specificity of Hsp90 in subpopulation I. This investigation has advanced our understanding of the molecular heterogeneity of cardiac proteasomes by identifying molecularly and functionally distinct cardiac 19S complexes. The preferential association of Hsp90 with 19S subpopulation

The ubiquitin proteasome system in human cardiomyopathies and heart failure

“…Two isoforms of regulatory particle non-ATPase 10 (multi-ubiquitin chain binding subunit) due to alternative splicing have been identified in the mouse heart (28). More recently, two distinct subpopulations of 19S complexes that differ in composition and proteolytic activity were isolated from mouse hearts (90). Finally, the 20S proteasome can complex with the 11S regulatory particle or in a hybrid formation with a 19S and 11S on either end (86).…”

High-Resolution Separation of Cardiovascular Proteomes

“…75 The selective incorporation rate of proteasome subunits in the pathogenesis of cardiac disease is of particular interest, because the myocardium harbors a heterogenic group of proteasome subpopulations. 76,77 The discovery of cardiac proteasome heterogeneity under nondenaturing conditions was enabled by multidimensional LC supported by 2-DE (Table) as well as a novel approach for isoelectric focusing of multiprotein complexes in a laminar flow (free flow electrophoresis). Throughout nondenaturing sample preparation, function and assembly are the most suitable parameters for optimization and standardization.…”

Targeting the Ubiquitin-Proteasome System in Heart Disease: The Basis for New Therapeutic Strategies

“…Functional studies on PA200 and 11S in cardiac tissue have not been performed in detail, but both generally enhance proteasome activities. Two distinct forms of 19S complexes observed in hearts contribute to additional heterogeneity due to an interaction with HSP90 (213). Again, their roles in healthy and diseased myocardium have yet to be elucidated.…”

“…Similar to other tissues and cells, cardiac 19S proteasome complexes consist of at least 19 distinct subunits (74,213). It should be noted that any 20S proteasome subpopulation can be assembled with 19S proteasome complexes.…”