HUMAN 4-HYDROXYPHENYLPYRUVATE DIOXYGENASE GENE (HPD)

Overlapping DNA fragments spanning approximately 21 kb of genomic DNA and encompassing the human 4-hydroxyphenglpyruvate dioxygenase gene (H PD) have been cloned by screening a human leukocyte genomic library an d by PCR amplification of human fibroblastic DNA. A continuous gene se quence of 20,890 nucleotides was established, including 1957 bp of the 5'-flanking region. The 4-hydroxyphenylpyruvate dioxygenase gene is c omposed of 14 exons interrupted. by 13 introns, all exhibiting convent ional vertebrate splicing, Computer analysis of the DNA sequence revea led 12 complete repetitive Alu elements, 1 in the 5'-flanking region a nd 11 in the intervening segments of the gene, The transcriptional ini tiation site was mapped to a position 35 nt upstream of the translatio nal start point. The computer analysis also identified several potenti al transcription regulatory elements, including one CRE site, two AP-2 sites, and two Sp1 sites, in the sequence upstream of the transcripti on initiation site. Functional analysis of promoter activity by transi ent transfection of chloramphenicolacetyl transferase reporter plasmid s revealed a possible involvement of cyclic adenosine monophosphate in the regulation of transcription. The highest level of expression of 4 -hydroxyphenylpyruvate dioxygenase was found in human Liver tissue as demonstrated by Northern blot analysis. (C) 1997 Academic Press.