The Role of Mitochondrial GRK2 in the Pathogenesis and Progression of Heart Failure

Abstract

Rationale: G protein-coupled receptor (GPCR) kinases (GRKs) are important regulators of cardiac function whose primary role is the phosphorylation of GPCRs and attenuation of downstream signaling, and GRK2 is upregulated after cardiac stress, injury, and during heart failure (HF). Recent studies have identified novel non-GPCR roles for GRKs such as cytoskeletal assembly, insulin signaling, fibrosis and pro-death signaling. One of the most compelling discoveries regarding novel roles of GRK2 in HF pathogenesis is its mitochondrial translocation (mtGRK2) following cardiac injury, which is associated with increased ROS production, decreased FA metabolic oxygen consumption, and pro-death signaling. Moreover, this localization is dependent on phosphorylation of serine 670 (S670) by kinases such as ERK1/2. Of note, expression of a GRK2 C-terminus peptide, βARKct, which contains S670 but no kinase domain or activity, is cardioprotective following injury in part due to inhibition of endogenous mtGRK2. This is significant because growing evidence demonstrates that cardiomyocyte metabolism is regulated by levels and activities of individual mitochondrial proteins Objective: This study sought to identify mitochondrial proteins which GRK2 interacts with either basally or after injury- or stress-induced translocation, and to determine whether functional regulation of mitochondrial function via phosphorylation of these proteins contributes to the phenomenon of bioenergetic defects during the development of HF. Methods and Results: Co-immunoprecipitation of GRK2 in vitro from primary ventricular cardiomyocytes and a human transformed cardiomyocyte-derived cell line was followed by liquid-chromatography mass-spectroscopy identification of all GRK2- interacting proteins. We followed this with proteomics analysis using DAVID and iv Ingenuity software to identify main pathways altered by GRK2 during stress or injury, focusing on mitochondrial dysfunction which revealed that GRK2 interacts with all major components of the electron transport chain (ETC). Using recombinant proteins as well as in vitro and in vivo models of myocardial infarction (MI), we demonstrate that GRK2 is able to phosphorylate the catalytic barrel of mitochondrial ATP synthase (complex V). Moreover, reduction of GRK2 levels in vivo using a GRK2 knockdown mouse model appears to protect against injury-induced bioenergetic deficits, whereas increased GRK2 levels in a transgenic GRK2 overexpression mouse model reveals both baseline deficits in ATP production as well as worsened post-MI outcomes. Conclusions: These collective data highlight the significance of the mitochondrial GRK2 interactome as a driver of cardiac bioenergetic deficits, particularly as a response to MI injury which progresses to HF. Given the current lack of effective HF treatments, this highly novel mechanism of GRK2 regulation of the mitochondrial ETC emphasizes the need for GRK2-targeting therapies for treating HF.