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J. Biol. Chem., Vol. 279, Issue 40, 42182-42191, October 1, 2004

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Regulation of the Cell Type-specific Dentin Sialophosphoprotein Gene Expression in Mouse Odontoblasts by a Novel Transcription Repressor and an Activator CCAAT-binding Factor^*

Shuo Chen, Aaron Unterbrink, Sheela Kadapakkam, Juan Dong, Ting Ting Gu, Julie Dickson, Hui-Hsiu Chuang, and Mary MacDougall¶

From the Department of Pediatric Dentistry, Dental School and the Department of Pharmacology, University of Texas Health Science Center, San Antonio, Texas 78229-3900

Dentin sialophosphoprotein (DSPP) is an extracellular matrix protein that is cleaved into dentin sialoprotein (DSP) and dentin phosphoprotein (DPP) with a highly restricted expression pattern in tooth and bone. Mutations of the DSPP gene are associated with dentin genetic diseases. Regulation of tissue-specific DSPP expression has not been described. To define the molecular basis of this cell-specific expression, we characterized the promoter responsible for the cell-specific expression of the DSPP gene in odontoblasts. Within this region, DNase I footprinting and electrophoretic mobility shift assays delineated one element that contains an inverted CCAAT-binding factor site and a protein-DNA binding site using nuclear extracts from odontoblasts. A series of competitive electrophoretic mobility shift assay analyses showed that the protein-DNA binding core sequence, ACCCCCA, is a novel site sufficient for protein binding. These two protein-DNA binding sequences are conserved at the same proximal position in the mouse, rat, and human DSPP gene promoters and are ubiquitously present in the promoters of other tooth/bone genes. Mutations of the CCAAT-binding factor binding site resulted in a 5-fold decrease in promoter activity, whereas abolishment of the novel protein-DNA binding site increased promoter activity by about 4.6-fold. In contrast to DSPP, expression levels of the novel protein were significantly reduced during odontoblastic differentiation and dentin mineralization. The novel protein was shown to have a molecular mass of 72 kDa. This study shows that expression of the cell type-specific DSPP gene is mediated by the combination of inhibitory and activating mechanisms.

Received for publication, March 4, 2004 , and in revised form, July 14, 2004.

^* This work was supported by NIDCR, National Institutes of Health, Grants PO1 DE113221 (to M. M.) and R03 DE014484 (to S. C.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

^¶ To whom correspondence should be addressed: Dept. of Pediatric Dentistry, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Dr., San Antonio, TX 78229-3900. Tel.: 210-567-3798; Fax: 210-567-6603; E-mail: MacDougall{at}uthscsa.edu.

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