Exon-specific northern analysis and rapid amplification of cDNA ends (RACE) reveal that the proximal promoter II (PII) is responsible for aromatase cytochrome P450 (CYP19) expression in human ovary.
Estrogens are synthesized from C19 steroids by a unique form of cytochrome P450, aromatase cytochrome P-450 (P-450AROM; the product of the CYP19 gene). We have shown that tissue-specific expression of human P-450AROM is determined, in part, by the use of alternative promoters. Previous methods of analysis for determining the specific 5'-termini of the different transcripts included S1 nuclease protection, primer extension, and Northern analysis. In the present study we have used the RACE procedure (rapid amplification of cDNA ends) to amplify and clone the 5' termini of P-450AROM transcripts expressed in human corpus luteum (CL). Sequencing of the resulting clones supports the results of the previously performed studies. Specifically, the proximal promoter, PII, is the predominant promoter utilized in CL, such that the start of transcription occurs 26 bp downstream of the putative TATA sequence. A minority of the clones possess an alternative 5'-end, namely I.3. Exon-specific Northern analysis confirms that the majority of the P-450AROM transcripts in CL tissue contain sequence specific for promoter II. Similarly, exon-specific Northern analysis indicates that transcripts in human follicles, as well as granulosa cells in culture, contain primarily sequence specific for promoter II.
Jenkins
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Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, Dallas 75235-9051.
Michael
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Mahendroo
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Simpson
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D001141:Aromatase / Q000096:biosynthesis* / Q000235:genetics; D001483:Base Sequence; D015152:Blotting, Northern; D003338:Corpus Luteum / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics; D004790:Enzyme Induction; D005091:Exons; D005260:Female; D006107:Granulosa Cells / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D009928:Organ Specificity; D006080:Ovarian Follicle / Q000378:metabolism; D010053:Ovary / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D012326:RNA Splicing; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics
D018076:DNA, Complementary; D012333:RNA, Messenger; D001141:Aromatase