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Dentin matrix protein 1 gene cis-regulation: use in osteocytes to characterize local responses to mechanical loading in vitro and in vivo.
- Source :
-
The Journal of biological chemistry [J Biol Chem] 2005 May 27; Vol. 280 (21), pp. 20680-90. Date of Electronic Publication: 2005 Feb 22. - Publication Year :
- 2005
-
Abstract
- Dentin matrix protein 1 (DMP1) is highly expressed in osteocytes and is mechanically responsive. To study osteocyte-specific and mechanically regulated DMP1 gene expression, the transcriptional activity of three cis-regulatory regions was first examined in an osteoblast differentiation model in vitro using a green fluorescent protein (GFP) reporter. Expression of the -9624 to +1996 bp (10 kb) and -7892 to +4439 bp (8 kb) DMP1 cis-regulatory regions dramatically increased in areas of mineralized matrix, in dendritic, osteocyte-like cells. Mineralizing cultures expressing the 8-kb construct show dramatic GFP increases in response to loading in cells with a dendritic morphology. Transgenic mice expressing the 8-kb DMP1-GFP and -2433 to +4439 bp (2.5 kb) DMP1-LacZ were generated. Osteocyte-specific expression was found with the 8 kb but not with the 2.5 kb in postnatal animals. However, the 2.5 kb could support expression in rapidly forming osteoblasts and pre-osteocytes in the embryo. Primary calvarial osteoblast cultures demonstrated that the 2.5 kb supports weak expression in a subset of osteoblasts and pre-osteocytes, but not in mature osteocytes. However, the 8 kb supports robust expression in primary bone marrow cultures. Therefore the region -7892 to -2433 bp, termed a 5.5-kb "Osteocyte Enhancer Module," appears to be required for osteocyte specificity. Ulnae of mice with the 8-kb DMP1-GFP were subjected to mechanical loading where GFP expression increased selectively and locally in osteocytes, distal to the mid-shaft and near the surface of the bone. Thus, the 8-kb region of the DMP1 gene is a target for mechanotransduction in osteocytes, and its cis-regulatory activity may be correlated to local strain in bone.
- Subjects :
- Animals
Biomechanical Phenomena
Bone Marrow Cells
Bone and Bones physiology
Cell Differentiation
Cell Line
Green Fluorescent Proteins genetics
Humans
Mice
Mice, Transgenic
Osteoblasts
Recombinant Fusion Proteins
Regulatory Sequences, Nucleic Acid
Stress, Mechanical
Transfection
Ulna
beta-Galactosidase genetics
Extracellular Matrix Proteins genetics
Gene Expression Regulation
Mechanotransduction, Cellular physiology
Osteocytes metabolism
Phosphoproteins genetics
Subjects
Details
- Language :
- English
- ISSN :
- 0021-9258
- Volume :
- 280
- Issue :
- 21
- Database :
- MEDLINE
- Journal :
- The Journal of biological chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 15728181
- Full Text :
- https://doi.org/10.1074/jbc.M500104200