A shared promoter element regulates the expression of three steroidogenic enzymes

Abstract

The adrenal cortex of the mouse coordinately expresses three cytochrome P450 enzymes that are required for the biosynthesis of corticosteroids: cholesterol side-chain cleavage enzyme (SCC), steroid 21-hydroxylase (21-OHase), and steroid 11 beta-hydroxylase (11 beta-OHase). Within their 5'-flanking regions, we previously identified six elements containing variations of an AGGTC motif that regulated expression in mouse Y1 adrenocortical cells: 21-OHase elements at -210, -140, and -65; SCC elements at -70 and -40; and an 11 beta-OHase element at -310. We demonstrate here that all six elements interact with the same, or closely related, DNA-binding protein(s). First, these elements all formed complexes of similar mobility in gel shift assays, suggesting that they interacted with protein(s) of similar size. Additional larger complexes were seen with those probes containing exact AGGTCA sequences. Second, competition experiments confirmed that the factor(s) interacting with different elements had closely related or identical recognition specificities. Finally, indistinguishable profiles of shift activities were seen upon fractionation of nuclear proteins over sequential chromatographic columns. Collectively, these results suggest that related elements interact with a shared protein to regulate three essential steroidogenic enzymes. An AGGTCA sequence motif comprises the response element for several members of the nuclear hormone receptor family. Oligonucleotide competitions and specific effects of antisera in gel shift assays implicated chicken ovalbumin upstream promoter-transcription factor in the formation of the larger complexes seen with the elements containing exact AGGTCA sequences. Therefore, this member of the nuclear hormone receptor family also may regulate the expression of the adrenal steroidogenic enzymes.