Limits...
A novel promoter controls Cyp19a1 gene expression in mouse adipose tissue.

Zhao H, Innes J, Brooks DC, Reierstad S, Yilmaz MB, Lin Z, Bulun SE - Reprod. Biol. Endocrinol. (2009)

Bottom Line: Subcutaneous and brown adipose tissue did not contain detectable Cyp19a1 mRNA.Dexamethasone significantly induced activity of this adipose-specific promoter region.These results expand the known 5'-regulatory region of the murine Cyp19a1 gene to 75 kb upstream of the translation start site.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Obstetrics and Gynecology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois 60611, USA. h-zhao@northwestern.edu

ABSTRACT

Background: Aromatase, the key enzyme in estrogen biosynthesis, is encoded by the Cyp19a1 gene. Thus far, 3 unique untranslated first exons associated with distinct promoters in the mouse Cyp19a1 gene have been described (brain, ovary, and testis-specific). It remains unknown whether aromatase is expressed in other mouse tissues via novel and tissue-specific promoters.

Methods: Real-time PCR was used to examine the aromatase expression levels in various C57BL/6 mouse tissues. 5'-rapid amplification of cDNA ends (5'-RACE) was used to determine the transcriptional start sites of Cyp19a1 transcripts. Promoter activity was measured using serial deletion mutants of DNA fused to the luciferase reporter gene. Primary mouse adipose fibroblasts were isolated and cultured from 16-week-old mouse gonadal fat pads.

Results: We systematically analyzed Cyp19a1 expression in a large number of mouse tissues, and demonstrated for the first time that aromatase was expressed in the male but not female gonadal fat pad. Subcutaneous and brown adipose tissue did not contain detectable Cyp19a1 mRNA. We used 5'-RACE to clone a novel gonadal fat-specific untranslated first exon, which is spliced onto a common junction 15 bp upstream of the translation start site. This adipose-specific first exon was mapped to approximately 75 kb upstream of the translation start site. Transfection of luciferase reporter gene plasmids containing the promoter region upstream of the adipose-specific first exon into murine 3T3-L1 adipose fibroblasts demonstrated significant basal promoter activity conferred primarily by the sequence located at -343/-1 bp. Dexamethasone significantly induced activity of this adipose-specific promoter region. Adipose-specific Cyp19a1 mRNA was expressed in primary mouse adipose fibroblasts and significantly induced by dexamethasone alone or serum plus dexamethasone.

Conclusion: Taken together, this research identified a novel, adipose-specific first exon of Cyp19a1 and its hormonally regulated promoter region in male murine gonadal fat. These results expand the known 5'-regulatory region of the murine Cyp19a1 gene to 75 kb upstream of the translation start site. Cyp19a1 expression in mouse adipose tissue may play an important role in reproductive biology and lipid metabolism.

Show MeSH

Related in: MedlinePlus

Cyp19a1 gene expression in 10- and 16-week-old male mouse gonadal fat. (A) Gonadal fat pad weight as percentage of body weight. (B) Cyp19a1 mRNA levels in male mouse gonadal fat were measured by real-time RT-PCR. Data are expressed as mean ± SE with n = 5–8 mice in each group. *p < 0.05 for 16-week-old mice vs. 10-week-old mice.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2684739&req=5

Figure 2: Cyp19a1 gene expression in 10- and 16-week-old male mouse gonadal fat. (A) Gonadal fat pad weight as percentage of body weight. (B) Cyp19a1 mRNA levels in male mouse gonadal fat were measured by real-time RT-PCR. Data are expressed as mean ± SE with n = 5–8 mice in each group. *p < 0.05 for 16-week-old mice vs. 10-week-old mice.

Mentions: We next determined whether Cyp19a1 expression in male gonadal fat varied with age or amount of tissue. While gonadal fat pad weight increased by 43% in 16-week-old mice compared with 10-week-old mice (Figure 2A), Cyp19a1 mRNA levels were not significantly different (Figure 2B).


A novel promoter controls Cyp19a1 gene expression in mouse adipose tissue.

Zhao H, Innes J, Brooks DC, Reierstad S, Yilmaz MB, Lin Z, Bulun SE - Reprod. Biol. Endocrinol. (2009)

Cyp19a1 gene expression in 10- and 16-week-old male mouse gonadal fat. (A) Gonadal fat pad weight as percentage of body weight. (B) Cyp19a1 mRNA levels in male mouse gonadal fat were measured by real-time RT-PCR. Data are expressed as mean ± SE with n = 5–8 mice in each group. *p < 0.05 for 16-week-old mice vs. 10-week-old mice.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2684739&req=5

Figure 2: Cyp19a1 gene expression in 10- and 16-week-old male mouse gonadal fat. (A) Gonadal fat pad weight as percentage of body weight. (B) Cyp19a1 mRNA levels in male mouse gonadal fat were measured by real-time RT-PCR. Data are expressed as mean ± SE with n = 5–8 mice in each group. *p < 0.05 for 16-week-old mice vs. 10-week-old mice.
Mentions: We next determined whether Cyp19a1 expression in male gonadal fat varied with age or amount of tissue. While gonadal fat pad weight increased by 43% in 16-week-old mice compared with 10-week-old mice (Figure 2A), Cyp19a1 mRNA levels were not significantly different (Figure 2B).

Bottom Line: Subcutaneous and brown adipose tissue did not contain detectable Cyp19a1 mRNA.Dexamethasone significantly induced activity of this adipose-specific promoter region.These results expand the known 5'-regulatory region of the murine Cyp19a1 gene to 75 kb upstream of the translation start site.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Obstetrics and Gynecology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois 60611, USA. h-zhao@northwestern.edu

ABSTRACT

Background: Aromatase, the key enzyme in estrogen biosynthesis, is encoded by the Cyp19a1 gene. Thus far, 3 unique untranslated first exons associated with distinct promoters in the mouse Cyp19a1 gene have been described (brain, ovary, and testis-specific). It remains unknown whether aromatase is expressed in other mouse tissues via novel and tissue-specific promoters.

Methods: Real-time PCR was used to examine the aromatase expression levels in various C57BL/6 mouse tissues. 5'-rapid amplification of cDNA ends (5'-RACE) was used to determine the transcriptional start sites of Cyp19a1 transcripts. Promoter activity was measured using serial deletion mutants of DNA fused to the luciferase reporter gene. Primary mouse adipose fibroblasts were isolated and cultured from 16-week-old mouse gonadal fat pads.

Results: We systematically analyzed Cyp19a1 expression in a large number of mouse tissues, and demonstrated for the first time that aromatase was expressed in the male but not female gonadal fat pad. Subcutaneous and brown adipose tissue did not contain detectable Cyp19a1 mRNA. We used 5'-RACE to clone a novel gonadal fat-specific untranslated first exon, which is spliced onto a common junction 15 bp upstream of the translation start site. This adipose-specific first exon was mapped to approximately 75 kb upstream of the translation start site. Transfection of luciferase reporter gene plasmids containing the promoter region upstream of the adipose-specific first exon into murine 3T3-L1 adipose fibroblasts demonstrated significant basal promoter activity conferred primarily by the sequence located at -343/-1 bp. Dexamethasone significantly induced activity of this adipose-specific promoter region. Adipose-specific Cyp19a1 mRNA was expressed in primary mouse adipose fibroblasts and significantly induced by dexamethasone alone or serum plus dexamethasone.

Conclusion: Taken together, this research identified a novel, adipose-specific first exon of Cyp19a1 and its hormonally regulated promoter region in male murine gonadal fat. These results expand the known 5'-regulatory region of the murine Cyp19a1 gene to 75 kb upstream of the translation start site. Cyp19a1 expression in mouse adipose tissue may play an important role in reproductive biology and lipid metabolism.

Show MeSH
Related in: MedlinePlus