Biochem. J. (2007) 405
(69–75) (Printed in Great Britain)
Runx3 negatively regulates Osterix expression in dental pulp cells
Li ZHENG*, Koichiro IOHARA*, Masaki ISHIKAWA†, Takeshi INTO*, Teruko TAKANO-YAMAMOTO‡, Kenji MATSUSHITA* and Misako NAKASHIMA*1
*Laboratory of Oral Disease Research, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, Aichi 474-8522, Japan, †Department of Endodontology and Operative Dentistry, Division of Oral Rehabilitation, Faculty of Dental Science, Kyushu University, Fukuoka 812-8582, Japan, and ‡Division of Orthodontics and Dentofacial Orthopedics, Graduate School of Dentistry, Tohoku University, Sendai 980-8575, Japan
Osterix, a zinc-finger-containing transcription factor, is required for osteoblast differentiation and bone formation. Osterix is also expressed in dental mesenchymal cells of the tooth germ. However, transcriptional regulation by Osterix in tooth development is not clear. Genetic studies in osteogenesis place Osterix downstream of Runx2 (Runt-related 2). The expression of Osterix in odontoblasts overlaps with Runx3 during terminal differentiation in vivo. Runx3 down-regulates Osterix expression in mouse DPCs (dental pulp cells). Therefore the regulatory role of Runx3 on Osterix expression in tooth development was investigated. Enforced expression of Runx3 down-regulated the activity of the Osterix promoter in the human embryonic kidney 293 cell line. When the Runx3 responsive element on the Osterix promoter, located at -713 to -707 bp (site 3, AGTGGTT) relative to the cap site, was mutated, this down-regulation was abrogated. Furthermore, electrophoretic mobility-shift assay and chromatin immunoprecipitation assays in mouse DPCs demonstrated direct functional binding of Runx3 to the Osterix promoter. These results demonstrate the transcriptional regulation of Osterix expression by Runx3 during differentiation of dental pulp cells into odontoblasts during tooth development.
Key words: bone morphogenetic protein 2 (BMP2), dental pulp cell (DPC), Osterix, Runx2, Runx3, tooth development.
Abbreviations used: BMP, bone morphogenetic protein; ChIP, chromatin immunoprecipitation; DIG, digoxigenin; DMEM, Dulbecco's modified Eagle's medium; DPC, dental pulp cell; Dspp, dentin sialophosphoprotein; DTT, dithiothreitol; EGFP, enhanced green fluorescent protein; EMSA, electrophoretic mobility-shift assay; FBS, foetal bovine serum; HEK-293, human embryonic kidney 293; KLK4, kallikrein 4; MSCV, murine stem cell virus; P1, postnatal day 1; RT, reverse transcription; Runx, Runt-related.
1To whom correspondence should be addressed (email misako@nils.go.jp).
Received 18 January 2007/19 February 2007; accepted 13 March 2007
Published as BJ Immediate Publication 13 March 2007, doi:10.1042/BJ20070104
© The Authors Journal compilation © 2007 Biochemical Society
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