RT Journal Article SR Electronic T1 Mammalian Tolloid Alters Subcellular Localization, Internalization, and Signaling of α1a-Adrenergic Receptors JF Molecular Pharmacology JO Mol Pharmacol FD American Society for Pharmacology and Experimental Therapeutics SP 532 OP 541 DO 10.1124/mol.105.016451 VO 70 IS 2 A1 Qi Xu A1 Ning Xu A1 Tan Zhang A1 Hui Zhang A1 Zijian Li A1 Feng Yin A1 Zhizhen Lu A1 Qide Han A1 Youyi Zhang YR 2006 UL http://molpharm.aspetjournals.org/content/70/2/532.abstract AB In the present study, we identified the CUB5 domain of mammalian Tolloid (mTLD) as a novel protein binding to α1A-adrenergic receptor (AR) using the yeast two-hybrid system. Whereas CUB5 did not couple to either α1B-AR or α1D-AR. It was determined that amino acids 322 to 359 of α1A-AR were the major binding region for CUB5. The direct interaction between α1A-AR cytoplasmic tail and CUB5 was discovered by glutathione S-transferase pull-down assay. We confirmed the interaction of mTLD with α1A-AR in human embryonic kidney (HEK) 293 cells by immunoprecipitation, immunofluorescence, and fluorescence resonance energy transfer. Although mTLD did not affect the density and affinity of receptors in crudely prepared membranes from HEK293 cells stably expressing α1A-AR, it significantly altered the subcellular localization of the receptors. Moreover, mTLD reduced the level of cell surface α1A-ARs, delayed the initial rate of agonist-induced receptor internalization, and facilitated agonist-induced calcium transient. We have demonstrated that mTLD interacts with α1A-AR directly, alters the subcellular localization of receptor, and influences agonist-induced α1A-AR internalization and calcium signaling.