論文 - 詳細
| RRC ID | 42088 |
|---|---|
| 著者 | Eelen G, Verlinden L, van Camp M, van Hummelen P, Marchal K, de Moor B, Mathieu C, Carmeliet G, Bouillon R, Verstuyf A. |
| タイトル | The effects of 1alpha,25-dihydroxyvitamin D3 on the expression of DNA replication genes. |
| ジャーナル | J Bone Miner Res |
| Abstract |
UNLABELLED:To identify key genes in the antiproliferative action of 1,25(OH)2D3, MC3T3-E1 mouse osteoblasts were subjected to cDNA microarray analyses. Eleven E2F-driven DNA replication genes were downregulated by 1,25(OH)2D3. These results were confirmed by quantitative RT-PCR in different cell types, showing the general nature of this action of 1,25(OH)2D3. INTRODUCTION:1Alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3] has a potent antiproliferative action characterized by a blocked transition from the G1- to the S-phase of the cell cycle. This study aims to identify genes whose expression is markedly altered after 1,25(OH)2D3 treatment in parallel with or preceding the observed G1-arrest. MATERIALS AND METHODS:The cDNA microarray technique was used, and the expression of approximately 4600 genes in MC3T3-E1 mouse osteoblasts was studied 6 and 12 h after treatment with 10(-8) M 1,25(OH)2D3. Quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) analyses were performed on MC3T3-E1 cells and on wildtype and vitamin D receptor (VDR) knockout primary murine epidermal keratinocytes (VDRwt MEKs, VDR-/- MEKs) and murine mammary tumor cells (GR) to confirm the microarray data. RESULTS AND CONCLUSIONS:After 12 h of treatment, in parallel with the 1,25(OH)2D3-induced G1 arrest, a particular set of DNA replication genes including a cell division cycle 6 homolog, a DNA polymerase alpha subunit, proliferating cell nuclear antigen, two DNA polymerase delta subunits, and flap-structure specific endonuclease 1, was downregulated at least 2-fold. These genes are known targets of the E2F family of transcription factors, which are probably the central mediators of this action of 1,25(OH)2D3. Indeed, as shown by transfection assays with an E2F reporter construct, 12- and 24-h treatment of MC3T3-E1 cells with 1,25(OH)2D3 reduced E2F activity by 49% and 73%, respectively. Quantitative RT-PCR analyses confirmed the downregulation of these DNA replication genes by 1,25(OH)2D3 in MC3T3-E1, GR, and VDRwt MEKs cells, but not in VDR-/- MEKs cells, showing that this 1,25(OH)2D3-driven antiproliferative action is of a general nature and depends on a functional VDR. |
| 巻・号 | 19(1) |
| ページ | 133-46 |
| 公開日 | 2004-1-1 |
| DOI | 10.1359/JBMR.0301204 |
| PMID | 14753745 |
| MeSH | 3T3 Cells Animals Calcitriol / pharmacology* Calcitriol / physiology Cell Cycle / drug effects Cell Cycle / genetics Cell Cycle Proteins / genetics Cell Line Cyclin-Dependent Kinase Inhibitor p21 Cyclin-Dependent Kinase Inhibitor p27 Cyclins / genetics DNA Replication / drug effects* DNA Replication / genetics DNA-Binding Proteins / genetics DNA-Directed DNA Polymerase / genetics Down-Regulation E2F Transcription Factors Flap Endonucleases / genetics Gene Expression Regulation / drug effects* Gene Silencing Genes, Reporter / genetics Keratinocytes / drug effects Keratinocytes / metabolism Mice Mice, Knockout Models, Biological Oligonucleotide Array Sequence Analysis Osteoblasts / cytology Osteoblasts / drug effects Osteoblasts / metabolism Polymerase Chain Reaction Proliferating Cell Nuclear Antigen / genetics Receptors, Calcitriol / genetics Transcription Factors / genetics Tumor Cells, Cultured Tumor Suppressor Proteins / genetics |
| IF | 5.854 |
| 引用数 | 54 |
| WOS 分野 | ENDOCRINOLOGY & METABOLISM |
| リソース情報 | |
| ヒト・動物細胞 | MC3T3-E1(RCB1126) |