GRK5‐mediated inhibitory phosphorylation is essential for inverse agonism at the cardiac mineralocorticoid receptor

Abstract

BackgroundIn the heart, aldosterone binds the mineralocorticoid receptor (MR) to exert damaging effects, e.g. fibrosis, apoptosis, oxidative stress, etc. GRK2 and GRK5 are the most abundant cardiac G protein‐coupled receptor (GPCR)‐kinases (GRKs). Both phosphorylate GPCRs but also non‐GPCR substrates. We recently showed that GRK5 blocks the cardio‐toxic MR‐dependent effects of aldosterone in the heart by directly phosphorylating the cardiac MR and inhibiting its transcriptional activity. MR antagonist (MRA) drugs are beneifical in human CHF by blocking the cardiac MR. Novel non‐steroidal MRAs, such as finerenone, may provide better cardio‐protection against aldosterone's cardio‐toxic actions than the classic steroidal MRAs, such as sprironolactone and eplerenone. Indeed, finerenone was very recently shown to be a more potent and efficacious inverse agonist at the MR than eplerenone in terms of cardiac fibrosis/adverse remodeling attenuation. Thus, in the present study, we sought to investigate potential differences between finerenone and eplerenone at engaging GRK5‐dependent cardiac MR phosphorylation and subsequent blockade.MethodsWe used the cardiomyocyte cell line H9c2 and neonatal rat ventricular myocytes (NRVMs). We measured MR phosphorylation via phospho‐serine immunoblotting & transcriptional activity via the luciferase reporter assay.ResultsGRK5 phosphorylates the MR in H9c2 cardiomyocytes in response to finerenone but not to eplerenone. Contrary to eplerenone, finerenone alone potently and efficiently suppresses MR transcriptional activity in H9c2 cardiac myocytes. GRK5‐dependent phosphorylation is necessary for the robust inverse agonism displayed by finerenone at the cardiac MR, since CRISPR‐mediated GRK5 gene deletion renders finerenone incapable of blocking cardiac MR transcriptional activity. Conversely, this non‐steroidal MRA fully suppresses cardiac MR transcriptional activity upon GRK5 overexpression in H9c2 cardiomyocytes. Eplerenone alone cannot suppress cardiac MR basal activity upon GRK5 overexpression or knockout. Finally, finerenone blocks aldosterone's effects more efficiently than eplerenone in control and GRK5‐overexpressing NRVMs.ConclusionsFinerenone appears a more efficacious cardiac MR blocker than eplerenone, partly via GRK5‐dependent cardiac MR inhibition, which eplerenone does not induce. This non‐canonical effect of GRK5 on the MR is essential for the efficient blockade of aldosterone's deleterious actions in the heart.Support or Funding Information1) American Heart Association 2) NSU President's Faculty Research & Development GrantThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.