title: Functional characterization of the mouse melanocortin 3 receptor gene promoter.
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.
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan.
authors.forename: Keisuke
authors.initials: K
authors.lastname: Okutsu
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan.
authors.forename: Fumiya
authors.initials: F
authors.lastname: Ojima
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan.
authors.forename: Naoto
authors.initials: N
authors.lastname: Shinohara
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan.
authors.forename: Shusuke
authors.initials: S
authors.lastname: Taniuchi
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan.
authors.forename: Yasusyo
authors.initials: Y
authors.lastname: Mizote
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan.
authors.forename: Kenji
authors.initials: K
authors.lastname: Aoki
authors.affiliation: Department of Biological Pharmacy, School of Pharmacy, Shujitsu University, Kita-ku, Okayama 703-8516, Japan.
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Kudo
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan.
authors.forename: Maho
authors.initials: M
authors.lastname: Ogoshi
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan.
authors.forename: Sakae
authors.initials: S
authors.lastname: Takeuchi
authors.affiliation: Department of Biology, Graduate School of Natural Science and Technology, Okayama University, Tsushima, Kita-ku, Okayama 700-8530, Japan. Electronic address: stakaha@cc.okayama-u.ac.jp.
authors.forename: Sumio
authors.initials: S
authors.lastname: Takahashi
mesh_terms: D024506:5' Flanking Region; D051701:Activating Transcription Factor 4 / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D004958:Estradiol / Q000494:pharmacology; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D057809:HEK293 Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008813:Mice, Inbred ICR; D010903:Pituitary Gland, Anterior / Q000187:drug effects / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D044104:Receptor, Melanocortin, Type 3 / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D018808:Transcription Factor AP-1 / Q000235:genetics* / Q000378:metabolism
chemical_list: C081452:Atf4 protein, mouse; C470053:Mc3r protein, mouse; D044104:Receptor, Melanocortin, Type 3; D018808:Transcription Factor AP-1; D051701:Activating Transcription Factor 4; D004958:Estradiol; D008156:Luciferases
keywords: Gene expression; Pituitary; Promoter; Proopiomelanocortin; α-MSH