Protein Kinase G Positively Regulates Proteasome-Mediated Degradation of Misfolded Proteins

Abstract

Background Proteasome functional insufficiency is implicated in a large subset of cardiovascular diseases and may play an important role in their pathogenesis. The regulation of proteasome function is poorly understood, hindering the development of effective strategies to improve proteasome function. Methods and Results Protein kinase G (PKG) was manipulated genetically and pharmacologically in cultured cardiomyocytes. Activation of PKG increased proteasome peptidase activities, facilitated proteasome-mediated degradation of surrogate (GFPu) and bona fide misfolded proteins (CryABR120G), and attenuated CryABR120G overexpression-induced accumulation of ubiquitinated proteins and cellular injury. PKG inhibition elicited the opposite responses. Differences in the abundance of the key 26S proteasome subunits Rpt6 and □5 between PKG manipulated and the control groups were not statistically significant, but the isoelectric points of were shifted by PKG activation. In transgenic mice expressing a surrogate substrate (GFPdgn), PKG activation by sildenafil increased myocardial proteasome activities and significantly decreased myocardial GFPdgn protein levels. Sildenafil treatment significantly increased myocardial PKG activity and significantly reduced myocardial accumulation of CryABR120G, ubiquitin conjugates, and aberrant protein aggregates in mice with CryABR120G-based desmin-related cardiomyopathy. No discernible effect on bona fide native substrates of the ubiquitin-proteasome system was observed from PKG manipulation in vitro or in vivo. Conclusions PKG positively regulates proteasome activities and proteasome-mediated degradation of misfolded proteins likely through posttranslational modifications to proteasome subunits. This may be a new mechanism underlying the benefit of PKG stimulation in treating cardiac diseases. Stimulation of PKG by measures such as sildenafil administration is potentially a new therapeutic strategy to treat cardiac proteinopathies.

Proteostasis in cardiac health and disease

“…Augmentation of PQC might, there fore, have beneficial effects in these forms of cardio myopathies. Indeed, experimental studies have already revealed that induction of proteasomal or autophagic degradation delays the onset of cardiomyopathy in mice with the Cryab R120G mutation [100][101][102] .…”

“…Prevention of cardiac remodelling by activ ation of cGMP dependent protein kinase by sildenafil treatment in aortic banded mice 186 was subsequently demonstrated to be mediated by augmentation of the UPS in the desmin related cardiomyopathy of Cryab R120G mice 101 . Conversely, proteasome inhibition was reported to prevent and regress ventricular hypertrophy during or after chronic administration of isoprenaline in mice 187 .…”

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

“…In a model of desmin-related cardiomyopathy, PA28a overexpression enhances lifespan and reduces hypertrophy, abundance of mutated CryAB aggregates, as well as ubiquitinated proteins (132). Similarly, decreased GFP/CL1 abundance after PKG activation via sildenafil is associated with reduced hypertrophy, less aggregation of mutated CryAB, and preserved cardiac function in the same DRC model (173). To see this illustration in color, the reader is referred to the web version of this article at www.liebertpub.com/ars (91).…”

“…Additional kinases affecting proteasome activities in the heart are PKG and PKN (Fig. 4) (173,197). In contrast to the stimulating effect of protein phosphorylation on proteasome activities, oxidative modification of proteasome subunits, for example, by 4-hydroxy-2-nonenal (a byproduct of lipid peroxidation), seems to be associated with…”

PKG Primes the Proteasome

“…The process of protein degradation by the 20S proteasome is dependent on ATP hydrolysis and appears to couple translocation, deubiquitination, and protease activity. 12 In this issue of Circulation, Ranek et al 13 build on earlier findings by this group that enhancement of proteasomal activities can protect the heart in the CryAB R120G model of desminrelated cardiomyopathy. In earlier work, these investigators used cardiomyocyte-specific overexpression of the proteasome subunit 28a (PA28a) to stimulate proteasomal activity.…”