A Specific Cis-acting Element Regulates in Vitro Transcription of Sex- dependent Mouse Steroid 16wHydroxylase (C-P4501s,) Gene*

The mouse steroid 16n-hydroxylase (C-P450& family consists of structurally similar genes with very divergent expressions (Wong, G., Itakura, T., Kawajiri, K., Skow, L., and Negishi, M. (1989) J. Biol. Chem. 264,2920-292’7). Of five family members, the C-P4501e, gene exhibits a male-specific expression whereas the P450cb gene is equally expressed in both males and females although the level of expression is much lower than that of the C-P4501e, gene. By DNase I footprinting and in vitro transcription of the C-P4501s, gene, we determined two cis-acting transcription elements. SD1 (sex difference information) in the -841-102 region from the transcription start site is one of the two cis-acting transcription elements, which is specific to the sex-specific C-P4501e, gene and confers a high transcription level of this gene. We substituted the nucleotide(s) in the SD1 sequence to the corresponding base(s) in the P450cb gene promoter and tested each substituted gene promoter by in vitro transcription. The results indicate that the function of SD1 as the transcription element depends most critically on the nucleotides at positions -100 to -98. Another cis-acting transcription element, CTE (common transcription element), is located between -44 and -68 in both C-P4501e, and P450cb genes. A column chromatography of nuclear extract indicates that the proteins bound to SD1 and CTE are different. We speculate therefore that SD1 is a specific and necessary element involved in the male-specific C-P45016, gene transcription. The sequence comparisons of the corresponding regions of SD1 and CTE among the mouse and the closely related rat P450 genes suggest that SD1 is a newly evolved cis-acting element in the sex-specific C-P4501e, gene which arose through gene duplication and selective nucleotide substitution.