Cloning, mRNA distribution, and functional expression of an avian counterpart of the chemokine receptor/HIV coreceptor CXCR4

The chemokine signaling system, which coordinates the basal and emergency t rafficking of leukocytes, presumably coevolved with the hematopoietic syste m, To study its phylogenetic origins, we used the open reading frame (ORF) of the human chemokine receptor CXCR4 as a genomic probe, since in mammals it is the most highly conserved chemokine receptor known, CXCR4 cross-hybri dized to genomic DNA from mouse and chicken, but not zebrafish, Drosophila, or Caenorhabditis elegans. Accordingly, we cloned the corresponding chicke n cDNA. The ORF is 359 codons long versus 352 for human CXCR4, and encodes a protein 82% identical to human CXCR4. In a calcium flux assay of receptor function, CHO-K1 cells stably transfected with the chicken cDNA responded specifically to human SDF-1, the specific ligand for CXCR4, but not to a pa nel of other chemokines tested at 100 nM. SDF-1 activated the cells in a do se-dependent manner (EC50 similar to5 nM), whereas parental CHO-K1 cells di d not respond, The CHO-K1 cell transfectants also bound I-125-SDF-1 specifi cally. Leukocytes from chicken peripheral blood expressed chCXCR4 mRNA and responded to human SDF-1 in a calcium flux assay with an EC50 similar to th at for chCXCR4-transfected CHO cells, suggesting that this response is medi ated by native chCXCR4. Analysis of chicken genomic DNA with the chicken cD NA as probe revealed a pattern consistent with a single copy gene, and the absence of any closely related genes. mRNA was detected in brain, bursa, li ver, small and large intestine, embryonal fibroblasts, and blood leukocytes , but not in stomach or pancreas, These results, which identify the first f unctional non-viral, non-mammalian chemokine receptor, suggest that the ori gins of a functional chemokine system extend at least to birds and suggest that, as in mammals, CXCR4 functions in many avian tissues.