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J. Biol. Chem., Vol. 276, Issue 43, 40001-40007, October 26, 2001

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Smad-interacting Protein 1 Is a Repressor of Liver/Bone/Kidney Alkaline Phosphatase Transcription in Bone Morphogenetic Protein-induced Osteogenic Differentiation of C2C12 Cells^*

Przemko Tylzanowski^§, Kristin Verschueren^¶, Danny Huylebroeck^¶, and Frank P. Luyten

From the  Laboratory of Skeletal Development and Joint Disorders, University of Leuven, Herestraat 49, 3000 Leuven, Belgium and the ^¶ Department of Cell Growth, Differentiation and Development (VIB-07), the Flanders Interuniversity Institute for Biotechnology (VIB) and the Laboratory of Molecular Biology, University of Leuven, Herestraat 49, B-3000 Leuven, Belgium

Up-regulation of liver/bone/kidney alkaline phosphatase (LBK-ALP) has been associated with the onset of osteogenesis in vitro. Its transcription can be up-regulated by bone morphogenetic proteins (BMPs), constitutively active forms of their cognate receptors, or appropriate Smads. The promoter of LBK-ALP has been characterized partially, but not much is known about its transcriptional modulation by BMPs. A few Smad-interacting transcriptional factors have been isolated to date. One of them, Smad-interacting protein 1 (SIP1), belongs to the family of two-handed zinc finger proteins binding to E2-box sequences present, among others, in the promoter of mouse LBK-ALP. In the present study we investigated whether SIP1 could be a candidate regulator of LBK-ALP transcription in C2C12 cells. We demonstrate that SIP1 can repress LBK-ALP promoter activity induced by constitutively active Alk2-Smad1/Smad5 and that this repression depends on the binding of SIP1 to the CACCT/CACCTG cluster present in this promoter. Interestingly, SIP1 and alkaline phosphatase expression domains in developing mouse limb are mutually exclusive, suggesting the possibility that SIP1 could also be involved in the transcriptional regulation of LBK-ALP in vivo. Taken together, these results offer an intriguing possibility that ALP up-regulation at the onset of BMP-induced osteogenesis could involve Smad/SIP1 interactions, resulting in the derepression of that gene.

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