Proteasome functional insufficiency in cardiac pathogenesis

Abstract

The ubiquitin-proteasome system (UPS) is responsible for the degradation of most cellular proteins. Alterations in cardiac UPS, including changes in the degradation of regulatory proteins and proteasome functional insufficiency, are observed in many forms of heart disease and have been shown to play an important role in cardiac pathogenesis. In the past several years, remarkable progress in understanding the mechanisms that regulate UPS-mediated protein degradation has been achieved. A transgenic mouse model of benign enhancement of cardiac proteasome proteolytic function has been created. This has led to the first demonstration of the necessity of proteasome functional insufficiency in the genesis of important pathological processes. Cardiomyocyte-restricted enhancement of proteasome proteolytic function by overexpression of proteasome activator 28α protects against cardiac proteinopathy and myocardial ischemia-reperfusion injury. Additionally, exciting advances have recently been achieved in the search for a pharmacological agent to activate the proteasome. These breakthroughs are expected to serve as an impetus to further investigation into the involvement of UPS dysfunction in molecular pathogenesis and to the development of new therapeutic strategies for combating heart disease. An interplay between the UPS and macroautophagy is increasingly suggested in noncardiac systems but is not well understood in the cardiac system. Further investigations into the interplay are expected to provide a more comprehensive picture of cardiac protein quality control and degradation.

Fig. 1.

A transgenic mouse model expressing carboxyl fusion of degron CL1 to green fluorescent protein (GFPdgn) as a surrogate misfolded protein. A: an illustration of the design of GFPdgn transgene and the sequence of degron CL1. GFPdgn is a fusion protein derived from carboxyl fusion of an enhanced GFP with degron CL1, which contains the depicted 16 amino acid residues. B: sequence and helical wheel representation of degron CL1 [adopted from Gilon et al. with permission (37)]. The hydrophobic amino acid residues are boxed. Note that in its helical structure, degron CL1 contains surface exposure of a long patch of hydrophobic amino acid residues. C: representative images of Western blot analysis of the indicated proteins in cardiac and hepatic tissues from adult GFPdgn transgenic mice (line 3) 20 h after an intraperitoneal injection of proteasome inhibitor MG-262 (5 μmol/kg) or the vehicle control. At the baseline, GFPdgn is expressed at a higher level in the heart than in the liver. MG-262 treatment increases ubiquitinated proteins, the precursor of the β₅-subunit of the 20S proteasome (pre-β5) and GFPdgn proteins in both organs shown here. Ub, ubiquitin; NS, not significant.

Fig. 2.

A schematic depiction of proteasome complexes expressed in mammalian cells. The 20S proteasome harbors all the proteolytic activities of the proteasome. To degrade proteins, the 20S needs to team up with the 19S and/or the 11S proteasome activator.

Fig. 3.

Use of GFPdgn reporter mice reveals proteasome functional insufficiency (PFI, A) and proteasome functional enhancement (B) in the heart. A: representative image of Western blot analyses for GFPdgn in the GFPdgn/CryAB^R120G double transgenic mouse heart, compared with littermate GFPdgn single transgenic mouse hearts. The soluble fraction of myocardial protein extracts from 1-mo-old mice of the indicated genotypes (top labels) were used for the analyses. PFI in the CryAB^R120G-based desmin-related cardiomyopathy mouse heart is revealed by the increased GFPdgn protein levels in the double transgenic mouse hearts. B: representative image of Western blot analyses for GFPdgn and proteasome activator 28α (PA28α) in the heart. Total proteins were extracted from ventricular myocardium of littermate mice of the indicated genotypes (labels at the top of the panel) at 8 wk of age and used for Western blot analyses for the indicated proteins. Note that GFPdgn protein levels were markedly decreased in PA28α overexpression hearts, indicating that proteasome proteolytic function in the heart is significantly enhanced by cardiomyocyte-restricted overexpression of PA28α. LC, loading control; tTA, tetracycline-controlled transactivator.

Fig. 4.

An illustration of a working hypothesis on the interplay between proteasome-mediated proteolysis and the selective autophagy. On the one hand, PFI accumulates ubiquitin conjugates which, through interaction, retain p62 proteins in the cell. PFI may also activate the nuclear factor erythroid-derived 2-related factor-1 (Nrf2)-mediated transcriptional program to activate p62 synthesis. The binding of p62 to ubiquitin conjugates in protein aggregates may recruit autophagosomes to and remove the aggregates, thereby increasing selective autophagy. On the other hand, chronic autophagy inhibition/impairment can accumulate p62, which is a substrate of autophagy. The accumulated p62 can bind to ubiquitinated proteins formed during normal ubiquitination and hinder them from being delivered to the proteasome for degradation; therefore, autophagy impairment may indirectly induce a PFI-like phenotype, although proteasome assembly and activities may not be decreased.