Dentinogenesis imperfecta (DGI) is a major genetic disorder affecting dentin with an estimated incidence of 1 in 6000-8000. DGI is clinically characterized by an opalescent dentin resulting in discoloration of the teeth. The dentin is poorly formed due to the irregular arrangement of dentinal tubules with abnormally low mineralization. In this disorder, teeth usually wear down rapidly, leaving short and brown stumps. Mutation and/or aberrant expression of dentin sialophosphoprotein (DSPP), a noncollagenous protein, has been implicated in DGI-II disorder. In order to delineate precise function of DSPP, we have cloned and characterized the murine dentin sialophosphoprotein gene (dspp) to establish the structure, regulation and functions associated with the gene products. The DSPP is translated initially as a single 940aa peptide from about 4.4kb mRNA and, later by unknown mechanisms, cleaved into N terminal as dentin sialoprotein (DSP) and the C terminal as dentinphosphoprotein (DPP). We have also characterized the basal promoter, enhancer and suppressor elements within 5' upstream sequences using a series of deletions in the established mouse odontoblast cell line MO6-G3. Transgenic animal model was developed with a reporter gene (?-galactosidase) under the control of 5.7 kb 5? flanking sequences to establish the presence of all the necessary elements within the promoter that would mimic the temporal and spatial expression pattern of the endogenous gene. Two independent transgenic lines harboring the transgenes were analyzed for the DSPP-LacZ expression profiles. Developmental expression patterns of the transgene was found to be very similar to the endogenous DSPP gene. We have further harnessed this promoter to generate odontoblast specific Cre transgenic mouse lines to create tooth specific knockout mouse models. To characterize the molecular mechanisms underlying dentinogenesis mediated by dspp, we have generated dspp -/- mice. The mutant mice showed dentin hypomineralization, enlarged pulp chambers, increased predentin and pulp exposures similar to human DGI-III. We found increased distribution of biglycan and decorin in the inter-calcospherite spaces, suggesting that these proteins may likely to interfere progression of calcospheritee coalescence in forming relatively uniform mineralization front. Biglycan and decorin belong to a group small leucine repeat proteins, that are known to interact with collagen fibrils in the process of dentin mineralization. In addition to the mineral nucleation, we have identified a novel negative regulatory function to Dspp protein in controlling the levels of biglycan and decorin, which is essential in maintaining the normal predentin. Amelogenins are mainly expressed by ameloblasts and secreted to form a major component of the enamel matrix. These proteins play an important role in enamel formation, defects in which are implicated in amelogenesis imperfecta. Amelogenin null mice generated in our laboratory displayed characteristic discoloration and attrition of teeth associated with defective enamel formation. Presence of amelogenins in cementum, as shown by immunostaining using anti-amelogenin antibodies, suggests a potential role for these proteins in cementum. The objective of the present study was to analyze the defects in the cementum in the amelogenin null mice in order to delineate the function of amelogenin during cementogenesis. Teeth of amelogenin null mice and their littermate controls at different ages were analyzed using light microscopy, electron microscopy and micro-radiography. EM analysis of the null teeth revealed cobbled roughness on their root surface whereas wild-type mice have a relatively smooth surface. Light microscopic analysis of tooth sections of the null mice displayed significant increase in the number of cementicles (~6 fold increase in the null mice). A number of lacunaes were noticed in the cementum of the null mice indicating resorption of cementum and dentin. Periodental-ligament fibers were frequently seen deeply embedded into these lacunaes. These findings suggest that amelogenin plays an important role not only in enamel formation but may also participate in cementogenesis. We have earlier reported that targeted over-expression of transforming growth factor-?1 (TGF-1) in the teeth of the transgenic mice (dTGF-?1) results into a novel tooth phenotype phenomimicking most prevalent tooth disorders in humans. This phenotype was associated with discoloration and attrition of teeth due to defective mineralization. We have now identified a novel expression of crystallin-family members in developing mouse teeth and its regulation by TGF-?1 in these transgenic mice. Crystallins are believed to be stress-related proteins, their expression in teeth implicates them in similar role since teeth are constantly subjected to physical friction and temperature fluctuations.
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