TISSUE-SPECIFIC PROMOTERS REGULATE AROMATASE CYTOCHROME-P450 GENE-EXPRESSION IN HUMAN OVARY AND FETAL TISSUES

The formation of estrogens from C19 steroids is catalyzed by a specific form of cytochrome P450, aromatase cytochrome P450 (P450AROM; the product of the CYP19 gene). Previous studies have demonstrated that aromatase activity in human adipose and ovarian granulosa cells is subject to complex multifactorial regulation and that changes in activity are correlated with changes in the levels of mRNA encoding P450AROM. We have previously isolated the human CYP19 gene. Two unique untranslated first exons (exons I.1 and I.2) have been identified in mRNA specific for P450AROM in human placenta. Although the proportion of transcripts encoding exon I.2 is very small genomic clones encoding the sequences of both exons I.1 and I.2 have recently been isolated. The corpus luteum of human ovary differs in that promoters I.1 and I.2 are completely inactive. Sequence analysis of the DNA immediately 5' of exon II (which contains the start site of translation) demonstrates the presence of a TATAA sequence beginning 149 basepairs 5' of the ATG initiation codon identified in placental exon II. Using a combination of primer extension and S1 nuclease protection analysis, it appears that the initiation site of ovarian P450AROM transcripts aligns 26 basepairs down-stream of the sequence TATAA. It appears, therefore, that the expression of P450AROM-specific mRNA in corpus luteum is regulated by an additional promoter (promoter II), which is located just 5' of exon II. Consistent with these observations, Northern analysis of poly(A)+ RNA isolated from placenta and corpus luteum demonstrates that the major promoter of placental P450AROM is promoter I.1, while the major promoter in the corpus luteum is promoter II. Analysis of the tissue-specific utilization of these promoters was accomplished by means of the polymerase chain reaction to amplify specific 5'-termini from mRNA templates. In addition to placenta, JEG-3 cells, hydatid moles, and fetal liver appear to use promoter I.1 and, to a limited extent, promoter I.2. These results suggest that the tissue-specific regulation of the human P450AROM gene is in part the consequence of the utilization of tissue-specific promoters.