| RRC ID |
39465
|
| 著者 |
Kimura T, Mizumoto Y, Ivell R.
|
| タイトル |
Differential protein-DNA binding analysis identifies a novel enhancer element, US-1, involved in the upregulation of the oxytocin receptor gene in human myometrium at term.
|
| ジャーナル |
Mol Cell Endocrinol
|
| Abstract |
In order to investigate the regulatory mechanisms involved in the transcription of the human oxytocin receptor (OTR) gene in the human myometrium at term of pregnancy, we subjected the 5' flanking region of the gene to a differential EMSA (electrophoretic mobility shift assay) procedure. Comparing nuclear proteins from term myometrium, in which OTR gene transcription is massively up-regulated, with those from the non-pregnant myometrium, indicated a prominent DNA-protein complex using the former extract. The sequence of the protein binding site was determined within 20 bp (TCTGCCTTCATCCAGCC) and designated as uterine stimulator motif-1 (US-1). The concatemerized US-1 sequence exhibited enhancer activity using a minimal thymidine kinase promoter (tk-81) in transfected SKN cells. We partially purified US-1 binding protein from SKN cells using a resin bead affinity procedure. Binding activity could be concentrated, although the protein eluate still comprised more than 20 component polypeptides. The molecular weight of the principal protein-DNA complex was determined following UV crosslinking to be 70 kDa. In circumstances where a cell-line mimicking the pregnant uterus at term is not available, the differential EMSA strategy, comparing OTR DNA protein binding in up- and down-regulated tissues, provides a powerful tool to investigate OTR regulation in the uterus. However, the precise characterization and identity of the specific DNA-binding protein(s) and consequent experimental verification of regulatory mechanisms still require elucidation.
|
| 巻・号 |
148(1-2)
|
| ページ |
137-49
|
| 公開日 |
1999-2-25
|
| DOI |
10.1016/s0303-7207(98)00225-1
|
| PII |
S0303-7207(98)00225-1
|
| PMID |
10221779
|
| MeSH |
Base Sequence
Cell Line
Cell Nucleus / metabolism
DNA Footprinting
Enhancer Elements, Genetic*
Female
Gene Expression Regulation*
Humans
Leiomyosarcoma
Molecular Sequence Data
Myometrium / metabolism*
Pregnancy / metabolism*
Pregnancy Trimester, Third
Promoter Regions, Genetic
Receptors, Oxytocin / biosynthesis
Receptors, Oxytocin / genetics*
Thymidine Kinase / genetics
Transfection
Tumor Cells, Cultured
Up-Regulation
|
| IF |
3.871
|
| 引用数 |
9
|
|
WOS 分野
|
ENDOCRINOLOGY & METABOLISM
CELL BIOLOGY
|
| リソース情報 |
| ヒト・動物細胞 |
|
