GENOMIC STRUCTURE, CHARACTERIZATION, AND IDENTIFICATION OF THE PROMOTER OF THE HUMAN IL-8 RECEPTOR-A GENE

Two unique but homologous receptors for the neutrophil chemoattractant , IL-8 have been cloned (designated IL-8RA and IL-8RB), each of which binds IL-8 with high affinity. IL-8RA mRNA expression was found to be regulated by granulocyte-CSF and LPS. in an attempt to understand the tissue-specific expression and to identify transcriptional regulatory elements, we have cloned, sequenced, and characterized the human IL-8R A gene. A h-DASH clone encoding the entire human IL-8RA gene was isola ted by screening a genomic library with a PCR-generated cDNA. After ma pping, subcloning, and sequencing several restriction fragments, a 9.2 -kb continuous DNA sequence was obtained. As the sizes of the publishe d cDNA (1.9 kb) and the mRNA determined by Northern blot analysis (2.1 kb) were not in agreement, a full-length cDNA was cloned by using a m odified rapid amplification of cDNA ends technique. We identified a 5' -untranslated region of 119 bp. After comparison with the genomic sequ ence, we found the gene consisted of two exons interrupted by an intro n of 1.7 kb. A 1050-bp ORF was encoded entirely in the second exon tog ether with a 834-bp 3'-untranslated region. The immediate GC-rich 5'-f lanking region upstream of exon 1 could serve as a constitutively acti ve promoter in chloramphenicol-acetyl-transferase-expression assays. E xpression analysis of additional upstream regions suggested the presen ce of silencer elements between positions -841 and -280. In conclusion , cloning a full-length cDNA permitted us to clone the human IL-8RA ge ne, identify the genomic structure, and characterize the promoter regi on.