Pseudoexons and regulatory elements in the genomic sequence of the beta-chemokine, alternative macrophage activation-associated CC-chemokine (AMAC)-1.
Recently, the authors reported the cloning of a novel human CC chemokine of alternatively activated macrophages (AMAC-1), whose expression is induced by Th2-associated cytokines such as interleukin 4 (IL-4), IL-13 and IL-10; vice versa, AMAC-1 expression is inhibited by Th1-associated cytokines such as interferon gamma (IFN-gamma). In order to study the genomic organization and transcriptional regulation of the AMAC-1 gene, genomic clones were isolated by screening a human lambda genomic library. Sequencing of a clone with a 1.7-kb insert gave a partial genomic sequence for the AMAC-1 gene. The complete AMAC-1 genomic sequence was obtained by bioinformational methods and the whole region spanning the AMAC-1 gene was verified by PCR amplification of subfragments and sequencing. The AMAC-1 gene consists of three exons. Whereas exons 2 and 3 were separated by a small intron of 411 bp, exon 1 and exon 2 were separated by 6 kb of non-translated genomic sequence containing two pseudoexons that are not expressed although they feature intact exon/intron boundaries and complete open reading frames. In order to allow a detailed analysis, a 2.7-kb fragment containing the promoter region and the first exon of AMAC-1 gene was cloned into a reporter gene construct. In the AMAC-1 promoter, two possible transcription start points were identified. In addition, several putative regulatory sequences for IL-4- and IFN-gamma-dependent transcriptional pathways were found including STAT6 and STAT1 binding sites as well as several AP-1 and C/EBP elements. Interestingly, a combined STAT6/STAT1 binding element is located in the direct vicinity of the first putative transcription start point. Competitive binding of IL-4-induced STAT6 versus IFN-gamma-induced STAT1 to this site may explain the antagonistic effects these cytokines exert on AMAC-1 expression.
Politz
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Klinik und Poliklinik für Dermatologie, Universitätsklinikum Benjamin Franklin, Freie Universität Berlin, Berlin, Germany.
Kodelja
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Guillot
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Orfanos
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Goerdt
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D000595:Amino Acid Sequence; D001483:Base Sequence; D019742:Chemokines, CC / Q000235:genetics*; D003001:Cloning, Molecular; D016384:Consensus Sequence; D004247:DNA / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D005091:Exons; D005809:Genes, Regulator; D015698:Genomic Library; D006801:Humans; D007438:Introns; D008262:Macrophage Activation / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012689:Sequence Homology, Nucleic Acid
C111792:CCL18 protein, human; D019742:Chemokines, CC; D017931:DNA Primers; D004247:DNA