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The Role of Mitochondrial GRK2 in the Pathogenesis and Progression of Heart Failure
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Thesis/Dissertation
Date
2022
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Molecular & Cellular Biosciences
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http://dx.doi.org/10.34944/dspace/8299
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.
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