(542.3) Mitochondrial GRK2 is a Novel Regulator of Cardiac Energetics

Poster Board Number: B159

Kimberly Ferrero (Lewis Katz School of Medicine at Temple University), Kurt Chuprun (Lewis Katz School of Medicine at Temple University), Jessica Pfleger (Virginia Tech University), Douglas Tilley (Lewis Katz School of Medicine at Temple University), Erhe Gao (Lewis Katz School of Medicine at Temple University), Walter Koch (Lewis Katz School of Medicine at Temple University)

G protein-coupled receptor (GPCR) kinase 2 (GRK2) is highly expressed in the heart, where during injury or heart failure (HF), both its levels and activity increase. GRK2 is canonically studied in the context of GPCR phosphorylation; however, noncanonical activities of GRK2 have emerged and it is now appreciated that GRK2 has a large non-GPCR interactome. For example, in cardiac myocytes, GRK2 translocates from the cytosol to mitochondria (mtGRK2) following oxidative stress or ischemia injury, and this pool of mtGRK2 is associated with negative effects on metabolism and also induces myocyte cell death. However, the mechanisms by which mtGRK2 contributes to cardiac dysfunction and HF are not fully understood. We hypothesized that mtGRK2 could have novel substrates and phosphorylate proteins involved in mitochondrial bioenergetics, thus contributing to our previously established post-injury phenotype. Stress-induced mitochondrial translocation of cytosolic GRK2 was validated in cell and animal models and the mtGRK2 interactome was identified using liquid chromatography-mass spectroscopy (LCMS). Proteomics analysis identified mtGRK2 interacting proteins which were involved in mitochondrial dysfunction, bioenergetics, and OXPHOS, particularly complexes I, II, IV and V of the electron transport chain (ETC). Specifically, mtGRK2 interactions with Complex V (ATP synthase) subunits were particularly increased following stress. We established that mtGRK2 phosphorylates ATP synthase on the F1 catalytic barrel, which is critical for oxidative phosphorylation and ATP production. We have also determined that alterations in either the levels or activity of GRK2 appear to alter ATP synthase enzymatic activity in vitro. Excitingly, in vivo data suggest that reducing levels of GRK2 in a mouse model of myocardial infarction prevents the post-injury reduction in ATP synthesis. We are currently assessing the ability of the SSRI drug paroxetine, a GRK2 inhibitor, to preserve mitochondrial bioenergetics in a transgenic GRK2 mouse model. Thus, phosphorylation of the ATP synthesis machinery by mtGRK2 may contribute to the impaired mitochondrial phenotype observed in injured or failing hearts such as reduced fatty acid metabolism and substrate utilization. These data uncover a druggable, novel target for rescuing cardiac function in patients with injured and/or failing hearts.

NIH R01 HL061690, NIH P01 HL075443, AHA 18MERIT33900036 to WJK.
NIH T32 HL091804, NIH 1F31 HL154567 to KMF.





Fig.1: Visual abstract for submission, "Mitochondrial GRK2 is a Novel Regulator of Cardiac Energetics"; Fig.2: Representative data. A) LCMS results for mitochondrial dysfunction; B) Electron transport chain hits in GRK2 interactome; C) Co-IP of ATP synthase subunits with GRK2; D) In-vivo injury model; E) mtGRK2 validation in vivo; F) Knockdown of GRK2 is energetically protective after myocardial infarction; G) mtGRK2 validation in vitro; H) GRK2 phosphorylates subunits of the F1 ATP synthase barrel; I) Identification of GRK2 target amino acids on alpha/beta F1 subunits.