GRANULOCYTE CHEMOTACTIC PROTEIN-2 ACTS VIA BOTH IL-8 RECEPTORS, CXCR1AND CXCR2

Interleukin-8 (lL-8) acts on human neutrophils via two receptors, CXCR 1 and CXCR2. It shares CXCR2 with all neutrophil-activating chemokines , which like IL-8 have a conserved Glu-Leu-Arg (ELR) N-terminal motif, but is generally considered to be the only relevant agonist for CXCR1 . IL-8 has a basic residue at the sixth position after the second cyst eine, which was suggested to contribute to CXCR1 specificity. Among th e other ELR chemokines, only granulocyte chemotactic protein 2 (GCP-2) has such a basic determinant. Using Jurkat cells that stably express either CXCR1 or CXCR2, we studied receptor activation by IL-8, GCP-2 e pithelial neutrophil-activating protein 2 (ENA-78) (which shares 77% i dentical amino acids with GCP-2) and growth-regulated oncogene alpha ( GRO alpha). At 10 nM and higher concentrations, GCP-2 and IL-8 induced significant activation of CXCR1-expressing cells, but no activity was found with GRO alpha and ENA-78. As expected, however, all four chemo kines had similar activities on CXCR2-expressing cells. A variant of G CP-2 in which the basic residue, Arg(20), was replaced by a glycine wa s synthesized. This derivative was ineffective on CXCR1, hut was as ac tive as wild-type GCP-2 in CXCR2-expressing cells. GCP-2 displaced rad iolabeled IL-8 from both receptors with low affinity, and in this resp ect resembled ENA-78 and GRO alpha. Our data show that GCP-2 acts via both IL-8 receptors and thus appears to be functionally more similar t o IL-8 than to the other ELR chemokines. Activation of CXCR1 appears t o depend significantly on the presence of a basic binding determinant close to the second cysteine.