TY - JOUR T1 - Identification and Characterization of Distinct C-terminal Domains of the Human Hydroxy- Carboxylic Acid Receptor-2 that are Essential for Receptor Export, Constitutive Activity, Desensitization and Internalization JF - Molecular Pharmacology JO - Mol Pharmacol DO - 10.1124/mol.112.081307 SP - mol.112.081307 AU - Guo Li AU - Qi Zhou AU - Yena Yu AU - Linjie Chen AU - Ying Shi AU - Jiansong Luo AU - Jeffrey L. Benovic AU - Jianxin Lu AU - Naiming Zhou Y1 - 2012/09/07 UR - http://molpharm.aspetjournals.org/content/early/2012/09/07/mol.112.081307.abstract N2 - The human hydroxy-carboxylic acid receptor 2 (HCA2), also known as GPR109A and HM74a, was first identified as a niacin receptor and has recently drawn significant attention because of its potential to clinically modify plasma lipids in a favorable manner. Our recent studies have demonstrated that the niacin-induced internalization of HCA2 receptors is regulated by GRK2 and arrestin3 and that internalized receptors rapidly recycle back to the cell surface. The investigation presented here used a combination of amino acid deletion and site-directed mutagenesis to identify structural and functional domains within the HCA2 C-terminus and explore their potential roles in receptor phosphorylation, desensitization and internalization. We first constructed four mutants with deletions of 10 to 15 amino acids each that were distinct from truncated mutants. We successfully identified different domains responsible for receptor export, constitutive activity, desensitization, phosphorylation and internalization. We also generated a comprehensive series of alanine substitution mutants, replacing conserved serine and threonine residues in the C-terminus with alanine residues, to pinpoint the key residues that are essential for GRK2-mediated phosphorylation and arrestin3 association. Moreover, we found that a sequence from residues 329 to 343 in the C-terminal tail of HCA2 plays a crucial role in keeping HCA2 in an inactive conformation. These data demonstrate the importance of distinct domains within the C-terminus of HCA2 for receptor cell surface expression, desensitization and internalization, phosphorylation and stabilization of an inactive receptor conformation. ER -