Endocrine Abstracts

CYP11B1 and CYP11B2 are responsible for the final steps in cortisol and aldosterone synthesis, respectively, in human. These two genes share 95% identity in coding regions and 90% identity in introns, but have very dissimilar promoter regions. To investigate whether there is a regulatory link between human CYP11B1 and CYP11B2 genes, we analyzed their upstream sequences using a pattern-search program termed multiple index sequence alignment. Three common sequence segments, cre, Ad5 and SF-1 binding site, were located in the proximal upstream regions. The Ad5 site is essential for basal expression of both genes. Mutation of the Ad5 site of CYP11B1 reduced promoter activity to a minimum level in the human adrenocortical cells. Overexpression of COUP-TFI increased CYP11B1 and CYP11B2 promoter activities. However, COUP-TFI probably exerted its stimulatory effect through cre, not Ad5 as previously reported. The presence of the SF-1 binding site also exhibited a positive effect on both promoters, but to a lesser extent compared to Ad5. Additionally, we discovered two clusters of common sequence segments in the distal upstream regions. Each cluster represents a short Alu repeat. Alu modulated CYP11B1 and CYP11B2 promoter activities as an enhancer. Copy number but not orientation affected its enhancement. Three truncated long L1 repeats were found in the CYP11B1 promoter, while one was located in the CYP11B2 promoter. The one situated 343-bp upstream of CYP11B1 diminished promoter activation of the gene. Removal of this L1 repeat regained promoter activity. The inhibition was colocalized with a putative open reading frame ORF2 encoded within L1. Although this ORF2 is nonfunctional owing to a premature termination, inhibition may arise from transcriptional expression of ORF2. Our results denied this possibility because no sign of expression was detected when ORF2 was replaced by a fluorescent protein. L1 apparently contains negative regulatory sequences or structures.