MECHANISMS OF G PROTEIN-COUPLED RECEPTOR 2 REGULATION AND INHIBITION IN CARDIOVASCULAR DISEASE AND AGING

Abstract

G protein-coupled receptor kinase 2 (GRK2) has been a thriving therapeutic target for cardiovascular disease treatment since its discovery for desensitizing and downregulating b-adrenergic receptors that are vital to cardiac function. GRK2 inhibition through a variety of methods in animal models of cardiac ischemia and heart failure achieved improvements in cardiac function, hemodynamic function, cardiomyocyte apoptosis, and fibrotic scar size among many other observations. Although GRK2 has been used as a therapeutic tool in multiple studies, its mechanisms of regulation are necessary to understand its role in disease pathogenesis and therapeutic application. This dissertation comprises two projects (1) investigating the microRNA (miRNA) regulation of GRK2 and (2) investigating the impact of loss of dynamic regulation of GRK2 through S-nitrosylation. (1) Candidate miRNAs were selected from miRNA microarray analysis of miRNA differential expression data and bioinformatic prediction. In vitro validation of kshv-miR- K12-3-5p and hsa-miR-181a-5p have shown their ability to bind to the GRK2 3’UTR as well as significantly decrease GRK2 mRNA or protein. The successful regulation of GRK2 through these miRNAs warrant in vivo application and investigation as GRK2-targeting HF therapy in a mouse model of HF. (2) In order to determine the impact of chronic GRK2 overactivity, mice that contained a knock-in mutation of GRK2 Cys340àSer (GRK2- C340S), a site of dynamic inhibitory regulation by S-nitrosylation, were allowed to age >12 months. Loss of S-nitrosylation of GRK2 was sufficient to cause cardiovascular remodeling and dysfunction over time.