Studies of dentin matrix protein 1 (DMP1) regulation and function in vivo

Dentin matrix protein 1 (DMP1) is a highly phosphorylated extracellular matrix protein, predominantly expressed in osteocytes in bone. Full-length DMP1 is enzymatically processed into a 37 kDa N- and a 57 kDa C-terminal fragment in vivo. The goal of this study was to identify the cis-regulatory regions that control osteoctye-specific expression of the DMP1 gene, to determine the in vivo function of the 57 kDa C-terminal fragment and to determine the role of DMP1 in control of phosphate homeostasis.; In vitro transient transfection assays showed that a 2.4-kb cis-regulatory region of the DMP1 gene contains basal promoter activity and that a 9.6-kb cis-regulatory region has the highest promoter activity. Transgenic mice bearing a lacZ reporter driven by these two different cis-regulatory regions separately were then generated and characterized. The region between -9.6 and -2.4-kb of the DMP1 gene has been identified as an osteocyte-specific module, which confers the osteocyte-specific expression of the lacZ reporter gene in vivo.; DMP1-null mice display a rachitic bone phenotype, elevated circulating FGF23 levels and hypophosphatemia. Transient transfection of the full-length DMP1 into various cell lines showed that DMP1 is processed similarly in vitro as in vivo into 37 kDa and 57 kDa fragments. Targeted re-expression of the full-length DMP1 or the 57 kDa fragment driven by a 3.6-kb Colla1 promoter in DMP1-null mice rescues the skeletal abnormalities, suggesting that the 57 kDa fragment is the essential functional domain of DMP1.; FGF23, a potent phosphaturic hormone, was sharply increased in DMP1-null osteocytes and serum and re-expression of DMP1 restored FGF23 to normal. Furthermore, a high phosphate diet rescued the rachitic phenotype in the growth plate and dramatically improved mineralization abnormalities in DMP1-null mice. However, co-expression of PHEX, a membrane protease whose mutations lead to a similar phenotype as that of DMP1-null mice, and DMP1 showed that PHEX had no effect on DMP1 cleavage in vitro, suggesting that both genes function independently in control of phosphate homeostasis.; In conclusion, the 57 kDa C-terminal fragment is the essential functional domain of DMP1 which plays a critical role in control of phosphate homeostasis through regulation of FGF23.