Activation of muscarinic receptor 2 stimulates proteasome function in cardiomyocytes

Abstract

Ubiquitin‐proteasome system (UPS)‐mediated proteolysis plays a central role in protein quality control and degradation in the cell. UPS dysfunction was observed in various forms of heart disease and might play an important role in the development of congestive heart failure (CHF).The parasympathetic nervous system (PNS) activity is decreased during CHF. As an initial step to investigate if and how PNS regulates the UPS in the heart, we employed neonatal rat ventricular myocytes (NRVM) culture. To better monitor UPS activity, a validated UPS surrogate substrate (a modified Green Fluorescent Protein, GFPu and GFPdgn) and a red fluorescent protein (RFP, not a specific substrate of the UPS) were co‐expressed in cultured NRVMs via adenoviral gene delivery.We found that stimulation of the muscarinic 2 receptor with pilocarpine significantly reduced the GFPu/RFP ratio, which is reversed by inhibiting protein kinase G (PKG). We have further found that the phosphorylation status of the Rpt6 subunit of the 19S proteasome and the beta 5 subunit of the 20S proteasome was markedly altered by stimulating this receptor. These preliminary findings suggest that PNS may positively regulate cardiac proteasome through activating PKG. Supported by NIH grants HL072166; HL085629, and HL068936 and American Heart Association grant 0740025N.