Protein Quality Control Disruption by PKCbII in Heart Failure; Rescue by the Selective PKCbII Inhibitor, bIIV5-3 Julio C. B. Ferreira1,2, Berta Napchan Boer3, Max Grinberg3, Patricia Chakur Brum2, Daria Mochly- Rosen1* 1 Department of Chemical and Systems Biology, Stanford University School of Medicine, Stanford, California, United States of America, 2 School of Physical Education and Sport, University of Sa˜o Paulo, Sa˜o Paulo, Brazil, 3 Heart Institute (InCor), University of Sa˜o Paulo, Medical School, Sa˜o Paulo, Brazil Abstract Myocardial remodeling and heart failure (HF) are common sequelae of many forms of cardiovascular disease and a leading cause of mortality worldwide. Accumulation of damaged cardiac proteins in heart failure has been described. However, how protein quality control (PQC) is regulated and its contribution to HF development are not known. Here, we describe a novel role for activated protein kinase C isoform bII (PKCbII) in disrupting PQC. We show that active PKCbII directly phosphorylated the proteasome and inhibited proteasomal activity in vitro and in cultured neonatal cardiomyocytes. Importantly, inhibition of PKCbII, using a selective PKCbII peptide inhibitor (bIIV5-3), improved proteasomal activity and conferred protection in cultured neonatal cardiomyocytes. We also show that sustained inhibition of PKCbII increased proteasomal activity, decreased accumulation of damaged and misfolded proteins and increased animal survival in two rat models of HF. Interestingly, bIIV5-3-mediated protection was blunted by sustained proteasomal inhibition in HF. Finally, increased cardiac PKCbII activity and accumulation of misfolded proteins associated with decreased proteasomal function were found also in remodeled and failing human hearts, indicating a potential clinical relevance of our findings. Together, our data highlights PKCbII as a novel inhibitor of proteasomal function. PQC disruption by increased PKCbII activity in vivo appears to contribute to the pathophysiology of heart failure, suggesting that PKCbII inhibition may benefit patients with heart failure. (218 words) Citation: Ferreira JCB, Boer BN, Grinberg M, Brum PC, Mochly-Rosen D (2012) Protein Quality Control Disruption by PKCbII in Heart Failure; Rescue by the Selective PKCbII Inhibitor, bIIV5-3. PLoS ONE 7(3): e33175. doi:10.1371/journal.pone.0033175 Editor: Niels Olsen Saraiva Caˆmara, Universidade de Sao Paulo, Brazil Received October 31, 2011; Accepted February 5, 2012; Published March 30, 2012 Copyright: 2012 Ferreira et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This study was supported by National Institute of Health Grant HL076675 to DMR. JCBF held a post-doctoral fellowship from Fundac¸a˜o de Amparo a Pesquisa do Estado de Sa˜o Paulo (FAPESP 2009/03143-1) and and Coordenac¸a˜o de Aperfeic¸oamento de Pessoal de Ensino Superior – Programa de Doutorado no Pa´ıs com Esta´gio no Exterior (CAPES-PDEE, 2177-07-2) – Brasil. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: Daria Mochly-Rosen is the founder of KAI Pharmaceuticals, Inc, a company that plans to bring PKC regulators to the clinic. However, none of the work described in this study is based on or supported by the company. The other authors have no disclosure. This does not alter the authors’ adherence to all the PLoS ONE policies on sharing data and materials. * E-mail: mochly@stanford.edu Introduction Maintenance of blood circulation during continual stress, such as hypertension or following cardiac ischemic events and infarction, contributes to cardiac wear and tear and results in accumulation of damaged cardiac proteins leading to cell death and further deterioration of cardiac functions. The cellular protein quality control (PQC) can detect, repair and dispose of cytotoxic damaged proteins using multiple control mechanisms, which include chaperone proteins, the ubiquitin-proteasome system (UPS) and autophagy [1]. The UPS is the primary effector of the PQC process, protecting long-lived cells, such as neurons and cardiomyocytes, from accumulation of aberrant and misfolded proteins [2]. The pathophysiological role of the PQC machinery in the heart emerged from studies showing accumulation of damaged proteins in humans and in animal models with cardiac diseases as well as cardiac mutations in PQC components [3,4]. There is also up-regulation of proteins involved in UPS and elevated levels of ubiquitinated proteins in hearts of human dilated cardiomyopathy [5]. Some studies found an overall decrease in proteasomal activity associated with and probably contributing to the increased steady state level of ubiquitinated proteins and cell death [5,6]. However, others reported that several components of the ubiquitin-protein system and/or its overall activity are increased in experimental compensated cardiac hypertrophy and heart failure [7]. There- fore, it remains to be determined whether dysfunction of specific PQC components, such as the UPS, contribute to the develop- ment of end-stage heart failure and which signaling events regulate them. Numerous studies have focused on identifying intracellular nodes where signals converge and serve as multi-effector brakes to suppress or reverse heart failure. We and others have identified PKCbII, which is over activated in failing hearts of humans [8] and in animal models [8,9,10,11], as a potential key player in heart failure. However, the molecular targets of PKCbII are still unknown. Using human remodeled and failing hearts with different etiologies and two different heart failure models in rats (myocardial infarction-induced and hypertension-induced heart failure; HF), we found a pronounced decline in components of the PQC machinery. Furthermore, we show for the first time that PKCbII, which is over-activated in HF both in humans [8] and in animal PLoS ONE | www.plosone.org 1 March 2012 | Volume 7 | Issue 3 | e33175