Use of molecular probes to study regulation of aromatase cytochrome P-450.
Aromatase, an enzyme complex localized in the endoplasmic reticulum of estrogen-producing cells, is composed of NADPH-cytochrome P-450 reductase, and aromatase cytochrome P-450 (cytochrome P-450AROM). To define the molecular mechanisms involved in the multifactorial regulation of cytochrome P-450AROM in estrogen-producing cells, we have isolated a cDNA specific for human cytochrome P-450AROM and have used this cDNA to isolate the human cytochrome P-450AROM gene. The cDNA sequence encodes a polypeptide of 503 amino acids and contains--near the carboxy-terminus, a region of high homology with the putative heme-binding regions of other P-450 cytochromes. COS1 cells transfected with an expression plasmid containing the cytochrome P-450AROM cDNA had the capacity to aromatize testosterone, androstenedione and 16 alpha-hydroxyandrostenedione, suggesting that a single polypeptide catalyzes all steps of the aromatization reaction using either of the three major C19-substrates. The human cytochrome P-450AROM gene is greater than 52 kb in size and consists of 10 exons and 9 introns. Hormonally induced changes in aromatase activity of human ovarian granulosa and adipose stromal cells are associated with comparable changes in cytochrome P-450AROM gene expression and synthesis, whereas the reductase component is only modestly affected. Studies are in progress to define the molecular mechanisms involved in the regulation of cytochrome P-450AROM gene expression in estrogen-producing cells.
Mendelson
C R
CR
Department of Biochemistry, Cecil H. and Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, Dallas 75235.
Means
G D
GD
Mahendroo
M S
MS
Corbin
C J
CJ
Steinkampf
M P
MP
Graham-Lorence
S
S
Simpson
E R
ER
D000273:Adipose Tissue / Q000032:analysis / Q000166:cytology / Q000201:enzymology*; D000595:Amino Acid Sequence; D001141:Aromatase / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D002478:Cells, Cultured; D004247:DNA / Q000032:analysis; D015342:DNA Probes; D005260:Female; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D006107:Granulosa Cells / Q000032:analysis / Q000201:enzymology*; D006801:Humans; D008969:Molecular Sequence Data; D012313:RNA / Q000032:analysis; D013379:Substrate Specificity
D015342:DNA Probes; D012313:RNA; D004247:DNA; D001141:Aromatase