Abstract
Melanocortin receptor 3 (MC3R) is expressed in the hypothalamus and pituitary in humans and rodents, and is involved in the control of feeding, energy metabolism, and pituitary function. In the mouse pituitary, MC3R is detected in mammotrophs. This study aimed to clarify the regulatory mechanism for Mc3r expression in the mouse pituitary. The promoter activities of reporter constructs for the MC3R gene 5'-flanking region up to - 4000. bp (transcription initiation site designated as +. 1) were analyzed. The promoter activity significantly increased in the - 86/+. 109 construct, but decreased in the - 38/+. 109 construct, indicating that the minimal promoter required for basal expression of Mc3r is located in the - 86/+. 109 region. Putative binding sites for transcription factors AP-1 and ATF4 were found in the 5'-flanking region of Mc3r. Site-directed mutation or deletion of these sites affected the promoter activities. In gel-shift assays with a nuclear extract of mouse anterior pituitary cells, band-shifts were detected for both sites after the addition of the nuclear extract, and were decreased in the presence of excess unlabeled probe competitors. These results indicated that both sites were involved in the regulation of Mc3r expression in anterior pituitary cells. Estradiol-17β treatment increased the Mc3r promoter activity, indicating that the gene is regulated by estradiol-17β. In conclusion, we have demonstrated the minimum promoter region required for Mc3r expression, and identified two binding sites for AP-1 and ATF4 and in the 5' upstream-flanking region of Mc3r that are essential for Mc3r expression.
| Original language | English |
|---|---|
| Pages (from-to) | 62-69 |
| Number of pages | 8 |
| Journal | Gene |
| Volume | 562 |
| Issue number | 1 |
| DOIs | |
| Publication status | Published - May 10 2015 |
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Keywords
- Gene expression
- Pituitary
- Promoter
- Proopiomelanocortin
- α-MSH
ASJC Scopus subject areas
- Genetics
- Medicine(all)
Cite this
Functional characterization of the mouse melanocortin 3 receptor gene promoter. / Okutsu, Keisuke; Ojima, Fumiya; Shinohara, Naoto; Taniuchi, Shusuke; Mizote, Yasusyo; Aoki, Kenji; Kudo, Toshiyuki; Ogoshi, Maho; Takeuchi, Sakae; Takahashi, Sumio.
In: Gene, Vol. 562, No. 1, 10.05.2015, p. 62-69.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Functional characterization of the mouse melanocortin 3 receptor gene promoter
AU - Okutsu, Keisuke
AU - Ojima, Fumiya
AU - Shinohara, Naoto
AU - Taniuchi, Shusuke
AU - Mizote, Yasusyo
AU - Aoki, Kenji
AU - Kudo, Toshiyuki
AU - Ogoshi, Maho
AU - Takeuchi, Sakae
AU - Takahashi, Sumio
PY - 2015/5/10
Y1 - 2015/5/10
N2 - Melanocortin receptor 3 (MC3R) is expressed in the hypothalamus and pituitary in humans and rodents, and is involved in the control of feeding, energy metabolism, and pituitary function. In the mouse pituitary, MC3R is detected in mammotrophs. This study aimed to clarify the regulatory mechanism for Mc3r expression in the mouse pituitary. The promoter activities of reporter constructs for the MC3R gene 5'-flanking region up to - 4000. bp (transcription initiation site designated as +. 1) were analyzed. The promoter activity significantly increased in the - 86/+. 109 construct, but decreased in the - 38/+. 109 construct, indicating that the minimal promoter required for basal expression of Mc3r is located in the - 86/+. 109 region. Putative binding sites for transcription factors AP-1 and ATF4 were found in the 5'-flanking region of Mc3r. Site-directed mutation or deletion of these sites affected the promoter activities. In gel-shift assays with a nuclear extract of mouse anterior pituitary cells, band-shifts were detected for both sites after the addition of the nuclear extract, and were decreased in the presence of excess unlabeled probe competitors. These results indicated that both sites were involved in the regulation of Mc3r expression in anterior pituitary cells. Estradiol-17β treatment increased the Mc3r promoter activity, indicating that the gene is regulated by estradiol-17β. In conclusion, we have demonstrated the minimum promoter region required for Mc3r expression, and identified two binding sites for AP-1 and ATF4 and in the 5' upstream-flanking region of Mc3r that are essential for Mc3r expression.
AB - Melanocortin receptor 3 (MC3R) is expressed in the hypothalamus and pituitary in humans and rodents, and is involved in the control of feeding, energy metabolism, and pituitary function. In the mouse pituitary, MC3R is detected in mammotrophs. This study aimed to clarify the regulatory mechanism for Mc3r expression in the mouse pituitary. The promoter activities of reporter constructs for the MC3R gene 5'-flanking region up to - 4000. bp (transcription initiation site designated as +. 1) were analyzed. The promoter activity significantly increased in the - 86/+. 109 construct, but decreased in the - 38/+. 109 construct, indicating that the minimal promoter required for basal expression of Mc3r is located in the - 86/+. 109 region. Putative binding sites for transcription factors AP-1 and ATF4 were found in the 5'-flanking region of Mc3r. Site-directed mutation or deletion of these sites affected the promoter activities. In gel-shift assays with a nuclear extract of mouse anterior pituitary cells, band-shifts were detected for both sites after the addition of the nuclear extract, and were decreased in the presence of excess unlabeled probe competitors. These results indicated that both sites were involved in the regulation of Mc3r expression in anterior pituitary cells. Estradiol-17β treatment increased the Mc3r promoter activity, indicating that the gene is regulated by estradiol-17β. In conclusion, we have demonstrated the minimum promoter region required for Mc3r expression, and identified two binding sites for AP-1 and ATF4 and in the 5' upstream-flanking region of Mc3r that are essential for Mc3r expression.
KW - Gene expression
KW - Pituitary
KW - Promoter
KW - Proopiomelanocortin
KW - α-MSH
UR - http://www.scopus.com/inward/record.url?scp=84925004500&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84925004500&partnerID=8YFLogxK
U2 - 10.1016/j.gene.2015.02.043
DO - 10.1016/j.gene.2015.02.043
M3 - Article
C2 - 25701401
AN - SCOPUS:84925004500
VL - 562
SP - 62
EP - 69
JO - Gene
JF - Gene
SN - 0378-1119
IS - 1
ER -