論文 - 詳細
| RRC ID | 61279 |
|---|---|
| 著者 | Tamiya H, Ikeda T, Jeong JH, Saito T, Yano F, Jung YK, Ohba S, Kawaguchi H, Chung UI, Choi JY. |
| タイトル | Analysis of the Runx2 promoter in osseous and non-osseous cells and identification of HIF2A as a potent transcription activator. |
| ジャーナル | Gene |
| Abstract |
Little is known about the upstream regulator of Runx2, a master regulator of osteoblast differentiation in bone tissues. To elucidate the molecular mechanism of Runx2 gene expression, we analyzed Runx2 promoter activity in osseous (MC3T3-E1, KS483, Kusa) and non-osseous (NIH3T3, C3H10T1/2, mouse embryonic fibroblasts) cells and also identified Runx2 upstream regulator using a Runx2 promoter-derived luciferase reporter system. After cloning 15 serial deletion constructs from -6832 bp/+390 bp to -37 bp/+390 bp of the Runx2-P1 promoter, we performed a transient transfection assay in osseous and non-osseous cells. A reduction in Runx2 promoter activity was observed in two regions; one was between -3 kb and -1 kb, and the other was between -155 bp and -75 bp. The step-down pattern in promoter activity between -3 kb and -1 kb was observed only in osseous cells. Interestingly, the step-down pattern between -155 bp and -75 bp was revealed in both cell types. Consistently, beta-galactosidase staining in axial skeleton of -3 kb-Runx2-P1-LacZ transgenic mice was positive, but that of all skeletal tissues of -1 kb-Runx2-P1-LacZ transgenic mice was negative. To identify upstream regulators of the Runx2-P1 promoter, we screened 100 transcription factors using Runx2-P1-luciferase reporter constructs in NIH3T3 fibroblasts and HeLa cells. Among them, HIF2A was identified as the strongest activator of Runx2-P1 promoter activity. A HIF2A-responsive site on the Runx2 promoter was identified between -106 bp and -104 bp by mutation analysis. An electrophoretic mobility shift assay and chromatin immunoprecipitation assay confirmed the binding of HIF2A to the Runx2-P1 promoter in vitro and in vivo, respectively. Knock-down using siRNA against HIF2A confirmed that HIF2A is an important regulator of Runx2 gene expression. Collectively, these results suggest that the region between -3 kb and -1 kb is required for the minimal skeletal tissue-specific expression of Runx2, and that the region between -155 bp and -75 bp is important for its basal transcription, which may be in part mediated by HIF2A in bone tissues. |
| 巻・号 | 416(1-2) |
| ページ | 53-60 |
| 公開日 | 2008-6-15 |
| DOI | 10.1016/j.gene.2008.03.003 |
| PII | S0378-1119(08)00112-1 |
| PMID | 18442887 |
| MeSH | Animals Base Sequence Bone and Bones / metabolism* Cell Line Cloning, Molecular Core Binding Factor Alpha 1 Subunit / genetics* DNA Mutational Analysis Humans Hypoxia-Inducible Factor 1, alpha Subunit / physiology* Mice Mice, Transgenic Promoter Regions, Genetic* Transcriptional Activation* Transfection |
| IF | 2.984 |
| オルトメトリクス指標 |
オルトメトリクス指標項目
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| 最多言及媒体 | X(Twitter) |
| 各媒体での言及数の合計 | 1 |
| 過去6か月間でのオルトメトリクス指標の変動値 | 0.0 |
| リソース情報 | |
| ヒト・動物細胞 | MC3T3-E1(RCB1126) |