Cholesterol is a major component of the lipid bilayer membrane, and is involved in membrane organization and function. G-Protein Coupled Receptors (GPCRs), such as the Adenosine A2a Receptor (A2aR), represent the largest superfamily of cell membrane receptors in the living world (Dodeyski et al., 2011). Ligand binding and activation of the receptor leads to a signal transduction pathway across the membrane, ending in a cellular response within the cell. This is achieved without the ligand entering the cell, making GPCRs ideal drug targets. The adenosine A2aR has been studied as a target for treatment of many diseases including Parkinson's disease (Vallano et al., 2011). Adenosine A2aR is responsible for activating adenylyl cyclase, which induces synthesis of cyclic AMP (cAMP).  The recently discovered high-resolution crystal structure of Adenosine A2aR reveals cholesterol-binding sites on the receptor. Additionally, simulations indicate that cholesterol must be bound to the receptor for the activation of adenylyl cyclase and in turn synthesis of cAMP (Lee et all, 2012). Our study seeks to further the understanding of the relationship between cholesterol and Adenosine A2aR in ligand binding and downstream expression of cyclic AMP in mammalian cells. We utilize three CHO M19 cell lines stably expressing human Adenosine A2aR. Two of the cell lines, SRD-12B and SRD-13A, are unable to synthesize cholesterol due to mutations in sterol regulatory element-binding proteins (Rawson et al., 1999). We hypothesize that cholesterol deficiency will play a role in ligand binding of Adenosine A2aR and downstream synthesis of cAMP in CHO M19 cells. We will describe our studies with these three cell lines by utilizing Adenosine A2aR expression studies, ligand binding assays and downstream signaling assays.

Role of membrane cholesterol in ligand binding and downstream signaling of human adenosine A2a receptor

247th National Meeting (2014)

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.