Study On Interaction Of Human Chemokine Receptor Ccr3 And Beta-Arrestin

G protein-coupled receptors (GPCRs) mediate different cell transmembrane signal transduction, and are important drug targets. The beta-arrestin mediated pathway is one of the important ways for GPCRs to play their function, which owns important significance for regulation of the function of GPCRs. However, until now it is still not clear how beta-arrestin interacts with GPCRs and mediates their trans-membraned signal transduction pathway. To address this issue,the CC chemokine receptor 3 (CCR3) was selected to study the interaction between beta-arrestin and GPCRs. Firstly, a co-expression system of beta-arrestin and CCR3 was constructed, and the interaction between beta-arrestin and CCR3 in living cells was analyzed using laser confocal fluorescence microscopy and fluorescence resonance energy transfer techniques. And the regulation effect of beta-arrestin on the chemotaxis of CCR3 stably transfected cells was also studied by RNAi and chemotaxis experiments. In addition, the interaction between beta-arrestin mutant (R169E) and CCR3 was further confirmed using QCM technology in vitro, and their binding constant was also determined. As a result, upon the stimulation of CCL11 (chemokine C-C motif ligand 11), the intracellular distance between beta-arrestin and CCR3 was significantly changed, and beta-arrestin protein was recruited to the cell membrane, which suggests that beta-arrestin could interact with CCR3 and involve in the CCR3-mediated signal transduction process. After silencing beta-arrestin by transfection with beta-arrestin-siRNA, the migration of CCR3 stablely transfected cells induced by CCL11 and CCL24 was significantly decreased, while the migration rate induced by CCL5 was not obviously changed. These results indicated that different chemokines shows different regulatory effects on the interaction between CCR3 and beta-arrestin. In vitro binding experiments further confirmed the interaction between beta-arrestin and CCR3, and the binding constant K-D between beta-arrestin mutant and CCR3 was determined as 1.35 x 10(-7). In conclusion, beta-arrestin can interact with CCR3 in living cells, and plays an important role in CCR3-mediated cell transmembrane signal transduction.