title: Functional analysis of the I.3, I.6, pII and I.4 promoters of CYP19 (aromatase) gene in human osteoblasts and their role in vitamin D and dexamethasone stimulation.
abstract: OBJECTIVE
  Current evidence suggests that extragonadal estrogens play an important role in bone metabolism. Estrogen biosynthesis is catalyzed by P450aromatase, encoded by the CYP19 gene. The aims of this paper were to study CYP19 gene expression in human osteoblasts under several hormone and cytokine treatments and to define promoter regions involved in this regulation.
  
  
  METHODS
  CYP19 transcript levels were measured from primary human osteoblasts and MG-63 cells by real-time PCR in basal conditions, and in response to seven different hormones and cytokines. Four promoters of CYP19 gene were cloned upstream of the luciferase gene and transfected into MG-63 cells. The effect of vitamin D and dexamethasone in these promoter activities was evaluated.
  
  
  RESULTS
  Vitamin D and dexamethasone were potent stimulators of CYP19 transcription, while testosterone and 17beta-estradiol stimulated moderately. Promoter pII proved the most potent in driving transient luciferase expression. Promoter I.4 displayed moderate activity, while promoters I.3 and I.6 were weak. A region upstream of exon I.3, including exon I.6, was identified as containing repressor elements of promoter pII. Promoter I.3 activity was modulated by repressors located within exon I.3, while an enhancer of promoter I.4 was detected within exon I.4. In the absence of fetal calf serum, dexamethasone stimulation was observed on promoters I.3 and I.4, while vitamin D stimulation acted only on promoter I.3.
  
  
  CONCLUSIONS
  Four regulatory regions of promoters pII, I.3 and I.4 are relevant to CYP19 expression in human osteoblasts. Vitamin D and dexamethasone modulate transcription through these regions.
authors.affiliation: URFOA-IMIM, Hospital del Mar, Universitat Autònoma de Barcelona, E-08003 Barcelona, Spain. aenjuanes@clinic.ub.es
authors.forename: Anna
authors.initials: A
authors.lastname: Enjuanes
authors.forename: Natalia
authors.initials: N
authors.lastname: Garcia-Giralt
authors.forename: August
authors.initials: A
authors.lastname: Supervía
authors.forename: Xavier
authors.initials: X
authors.lastname: Nogués
authors.forename: Silvia
authors.initials: S
authors.lastname: Ruiz-Gaspà
authors.forename: Mariona
authors.initials: M
authors.lastname: Bustamante
authors.forename: Leonardo
authors.initials: L
authors.lastname: Mellibovsky
authors.forename: Daniel
authors.initials: D
authors.lastname: Grinberg
authors.forename: Susana
authors.initials: S
authors.lastname: Balcells
authors.forename: Adolfo
authors.initials: A
authors.lastname: Díez-Pérez
mesh_terms: D000368:Aged; D001141:Aromatase / Q000096:biosynthesis / Q000235:genetics*; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D003907:Dexamethasone / Q000494:pharmacology*; D004958:Estradiol / Q000494:pharmacology; D005260:Female; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D020738:Leptin / Q000494:pharmacology; D008297:Male; D008875:Middle Aged; D010006:Osteoblasts / Q000201:enzymology*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D013739:Testosterone / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D014807:Vitamin D / Q000494:pharmacology*
chemical_list: D007375:Interleukin-1; D020738:Leptin; D014409:Tumor Necrosis Factor-alpha; D014807:Vitamin D; D013739:Testosterone; D004958:Estradiol; D003907:Dexamethasone; D001141:Aromatase