β-Arrestin Differentially Regulates the Chemokine Receptor CXCR4-mediated Signaling and Receptor Internalization, and This Implicates Multiple Interaction Sites between β-Arrestin and CXCR4

Abstract The chemokine receptor CXCR4 has recently been shown to be a co-receptor involved in the entry of human immunodeficiency virus type 1 into target cells. This study shows that coexpression of β-arrestin with CXCR4 in human embryonic kidney 293 cells attenuated chemokine-stimulated G protein activation and inhibition of cAMP production. Truncation of the C-terminal 34 amino acids of CXCR4 (CXCR4-T) abolished the effects of β-arrestin on CXCR4/G protein signaling, indicating the functional interaction of the receptor C terminus with β-arrestin. On the other hand, receptor internalization and the subsequent activation of extracellular signal-regulated kinases were significantly promoted by coexpression of β-arrestin with CXCR4, whereas the C-terminal truncation of CXCR4 did not affect this regulation of β-arrestin, suggesting that β-arrestin can functionally interact with CXCR4 with or without the C terminus. Moreover, β2V54D, the dominant inhibitory mutant of β-arrestin 2, exerted no effects on CXCR4/G protein signaling, but strongly influenced receptor internalization and extracellular signal-regulated kinase activation. Further cross-linking experiments demonstrated that β-arrestin as well as β2V54D could physically contact both CXCR4 and CXCR4-T. Glutathione S-transferase pull-down assay showed that β-arrestin was able to bind efficiently in vitro to both the third intracellular loop and the 34-amino acid C terminus of CXCR4. Taken together, our data clearly establish that β-arrestin can effectively regulate different functions of CXCR4 and that this is mediated through its distinct interactions with the C terminus and other regions including the third loop of CXCR4.