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J Biol Chem, Vol. 274, Issue 23, 16287-16294, June 4, 1999
-Arrestins Regulate Interleukin-8-induced CXCR1
Internalization
§,§,,,§,,,¶,§
From the The functional role of neutrophils during acute
inflammatory responses is regulated by two high affinity interleukin-8
receptors (CXCR1 and CXCR2) that are rapidly desensitized and
internalized upon binding their cognate chemokine ligands. The
efficient re-expression of CXCR1 on the surface of neutrophils
following agonist-induced internalization suggests that CXCR1 surface
receptor turnover may involve regulatory pathways and intracellular
factors similar to those regulating Laboratory of Molecular Immunology and
Inflammation, John P. Robarts Research Institute, London, Ontario N6G
2V4, Canada, the § Department of Microbiology and
Immunology, The University of Western Ontario, London, Ontario N6A 5C1
Canada, and the ¶ Department of Physiology, Pharmacology and
Toxicology, The University of Western Ontario, and John P. Robarts
Research Institute, London, Ontario N6A 5K8, Canada
2-adrenergic
receptor internalization and re-expression. To examine the
internalization pathway utilized by ligand-activated CXCR1, a CXCR1-GFP
construct was transiently expressed in two different cell lines, HEK
293 and RBL-2H3 cells. While interleukin-8 stimulation promoted CXCR1
sequestration in RBL-2H3 cells, receptor internalization in HEK 293 cells required co-expression of G protein-coupled receptor kinase 2 and
-arrestin proteins. The importance of -arrestins in CXCR1
internalization was confirmed by the ability of a dominant negative
-arrestin 1-V53D mutant to block internalization of CXCR1 in RBL-2H3
cells. A role for dynamin was also demonstrated by the lack of CXCR1 internalization in dynamin I-K44A dominant negative mutant-transfected RBL-2H3 cells. Agonist-promoted co-localization of transferrin and
CXCR1-GFP in endosomes of RBL-2H3 cells confirmed that receptor internalization occurs via clathrin-coated vesicles. Our data provides
a direct link between agonist-induced internalization of CXCR1 and a
requirement for G protein-coupled receptor kinase 2, -arrestins, and
dynamin during this process.
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