Expression and growth-promoting function of the IL-8 receptor beta in human melanoma cells.

Abstract

The chemokine GRO alpha is an autocrine growth factor for melanoma cells. Although GRO alpha has been identified as a high affinity ligand for the IL-8 receptor beta (IL-8R beta) in recombinant systems, the receptor mediating its action in melanoma cells has been a matter of debate. Here, we show by reverse transcription and PCR expression of IL-8R beta, mRNA transcripts in different melanoma cell lines and in normal human melanocytes. To characterize the role of the IL-8R beta in melanoma cells, antiserum was raised in rabbits against a fusion protein containing the NH2-terminal portion of the receptor. Its specificity was shown by flow cytometry with IL-8R beta-transfected HL60 cells. A specific epitope could be mapped with IL-8R beta mutants to the peptide sequence between ASP-4 and ASP-14 of this receptor. Binding studies with [125I]GRO alpha in IL-8R beta transfectants indicated ligand antagonistic properties of this Ab. Expression of IL-8R beta protein at the cell surface of various melanoma cell lines could be shown by flow cytometry with F(ab')2 fragments of the IL-8R beta antiserum. Moreover, anti-IL-8R beta Ab partially blocked specific binding of [125I]GRO alpha in various melanoma cell lines. Addition of F(ab')2 fragments of the IL-8R beta antiserum or of neutralizing anti-GRO alpha mAb to different melanoma cell lines identified this GRO alpha-IL-8R beta interaction as a major component required for serum-independent melanoma cell growth.