Generation and Characterization of DSPP-Cerulean/DMP1-Cherry reporter mice

The non-collagenous proteins (NCPs) in dentin include two dentin-specific proteins, Dentin Sialoprotein (DSP) and Dentin Phosphoprotein (DPP) and Dentin Matrix Protein-1 (Dmp1), which is also found in bone.  DSP and DPP are generated from a larger precursor, the dentin sialophosphoprotein (DSPP) protein. High levels of expression of DSPP and DSP are the hallmark of odontoblast differentiation and are routinely used to distinguish differentiated odontoblasts from undifferentiated progenitors and osteoblasts.

Objective: To generate a new fluorescent protein reporter transgenic mice that allows identification and isolation of odontoblasts at later stages of differentiation.  

Method: DSPP-Cerulean/DMP1-Cherry transgenic mouse were  generated using a bacterial recombination strategy with the mouse BAC clone RP24-258g7. The expression of both fluorescent reporters in the mandibular molars and incisor was examined in frozen sections at various stages of tooth development. Primary pulp cultures from these animals were used to examine the stage-specific activation of both fluorescent reporters during in vitro mineralization. Reporter gene expression was compared to the expression of endogenous Dspp and Dmp1.

Result: DSPP-Cerulean and DMP1-Cherry were not expressed in pre-odonblasts, ameloblasts or dental pulp. The expression of both reporter genes was first detected in functional odontoblasts associated with pre-dentin and increased in newly differentiated odontoblasts. Fully differentiated odontoblasts exhibited high levels of DSPP-Cerulean and reduced levels of DMP1-Cherry expression. Unlike DSPP-Cerulean, DMP1-Cherry was also expressed at high levels in the osteoblasts and osteocytes within the alveolar bone. In primary pulp cultures DSPP-Cerulean expressing cells appeared within the mineralized regions. There was close correlation between the expression DSPP-Cerulean and endogenous Dspp.

Conclusion: DSPP-Cerulean/DMP1-Cherry transgenic mice provide a new experimental model to study the molecular mechanisms that regulate the progression of progenitors into odontoblasts.  Supported by a fellowship from AADR and Grant R01-DE016689.