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Transcriptional repression of the Dspp gene leads to dentinogenesis imperfecta phenotype in Col1a1-Trps1 transgenic mice.

Napierala D, Sun Y, Maciejewska I, Bertin TK, Dawson B, D'Souza R, Qin C, Lee B - J. Bone Miner. Res. (2012)

Bottom Line: On the molecular level, we demonstrated that sustained high levels of Trps1 in odontoblasts lead to dramatic decrease of Dspp expression as a result of direct inhibition of the Dspp promoter by Trps1.During tooth development Trps1 is highly expressed in preodontoblasts, but in mature odontoblasts secreting matrix its expression significantly decreases, which suggests a Trps1 role in odontoblast development.Thus, we provide novel insights into mechanisms of transcriptional dysregulation that leads to DGI.

View Article: PubMed Central - PubMed

Affiliation: Institute of Oral Health Research, Department of Oral and Maxillofacial Surgery, School of Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294-0007, USA. dobrawan@uab.edu

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Decreased levels of DSP and DPP proteins, and Dspp gene expression in odontoblast of Col1a1-Trps1 mice. (A) Detection of SIBLING proteins isolated from dentin of 8-week-old WT and transgenic mice. Proteins were extracted from dentin and separated into 120 fractions using ion-exchange chromatography. Selected fractions containing SIBLING proteins were analyzed on an SDS-PAGE gel. SIBLING proteins were visualized by Stains-All. Numbers above the gel image indicate the chromatographic protein fractions analyzed on the gel; these fractions contained DPP and DMP1 fragments. The dashed-line box marks DPP protein, the solid-line box marks the N-terminal fragment of DMP1 protein. In transgenic mice a total level of DPP is decreased in comparison with WT mice, whereas levels of DMP1 show no difference. (B) Western blot with DSP antibodies shows decreased levels of DSP in transgenic dentin. (C) RNA in situ hybridization with probes specific for Dspp and Dmp1 (antisense molars, P2; sense molars P4). The boxed areas are magnified on the images in the right low corner of each respective image. In WT animals Dspp is expressed in both odontoblasts and ameloblasts, while in Col1a1-Trps1 mice Dspp expression in odontoblasts is significantly reduced. There is no difference in expression of Dmp1 between WT and transgenic odontoblasts. Od = odontoblasts; Am = ameloblasts. Scale bars = 100 µm.
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fig03: Decreased levels of DSP and DPP proteins, and Dspp gene expression in odontoblast of Col1a1-Trps1 mice. (A) Detection of SIBLING proteins isolated from dentin of 8-week-old WT and transgenic mice. Proteins were extracted from dentin and separated into 120 fractions using ion-exchange chromatography. Selected fractions containing SIBLING proteins were analyzed on an SDS-PAGE gel. SIBLING proteins were visualized by Stains-All. Numbers above the gel image indicate the chromatographic protein fractions analyzed on the gel; these fractions contained DPP and DMP1 fragments. The dashed-line box marks DPP protein, the solid-line box marks the N-terminal fragment of DMP1 protein. In transgenic mice a total level of DPP is decreased in comparison with WT mice, whereas levels of DMP1 show no difference. (B) Western blot with DSP antibodies shows decreased levels of DSP in transgenic dentin. (C) RNA in situ hybridization with probes specific for Dspp and Dmp1 (antisense molars, P2; sense molars P4). The boxed areas are magnified on the images in the right low corner of each respective image. In WT animals Dspp is expressed in both odontoblasts and ameloblasts, while in Col1a1-Trps1 mice Dspp expression in odontoblasts is significantly reduced. There is no difference in expression of Dmp1 between WT and transgenic odontoblasts. Od = odontoblasts; Am = ameloblasts. Scale bars = 100 µm.

Mentions: In contrast to DGI-I, in which both bone and dentin are affected, presentation of DGI-II and DGI-III is restricted to dental abnormalities. These forms of DGI are caused by mutations in the DSPP gene. The DSPP gene encodes a protein that is proteolytically processed to form DSP and DPP, which are the most abundant noncollagenous proteins in the dentin matrix. To understand the molecular basis of abnormal dentin formation in Col1a1-Trps1 mice, we compared the relative abundance of DSP, DPP, and other SIBLING proteins in the dentin matrices between WT and transgenic animals.30–32 These analyses showed dramatically reduced levels of DPP in transgenic animals (Fig. 3A). At the same time, levels of the N-terminal fragment of DMP1 showed no apparent differences as evaluated by Stains-All staining. Since DSP is less sensitive to Stains-All staining, we compared levels of DSP between WT and transgenic mice by Western blot. These analyses also showed dramatically decreased levels of DSP (Fig. 3B).


Transcriptional repression of the Dspp gene leads to dentinogenesis imperfecta phenotype in Col1a1-Trps1 transgenic mice.

Napierala D, Sun Y, Maciejewska I, Bertin TK, Dawson B, D'Souza R, Qin C, Lee B - J. Bone Miner. Res. (2012)

Decreased levels of DSP and DPP proteins, and Dspp gene expression in odontoblast of Col1a1-Trps1 mice. (A) Detection of SIBLING proteins isolated from dentin of 8-week-old WT and transgenic mice. Proteins were extracted from dentin and separated into 120 fractions using ion-exchange chromatography. Selected fractions containing SIBLING proteins were analyzed on an SDS-PAGE gel. SIBLING proteins were visualized by Stains-All. Numbers above the gel image indicate the chromatographic protein fractions analyzed on the gel; these fractions contained DPP and DMP1 fragments. The dashed-line box marks DPP protein, the solid-line box marks the N-terminal fragment of DMP1 protein. In transgenic mice a total level of DPP is decreased in comparison with WT mice, whereas levels of DMP1 show no difference. (B) Western blot with DSP antibodies shows decreased levels of DSP in transgenic dentin. (C) RNA in situ hybridization with probes specific for Dspp and Dmp1 (antisense molars, P2; sense molars P4). The boxed areas are magnified on the images in the right low corner of each respective image. In WT animals Dspp is expressed in both odontoblasts and ameloblasts, while in Col1a1-Trps1 mice Dspp expression in odontoblasts is significantly reduced. There is no difference in expression of Dmp1 between WT and transgenic odontoblasts. Od = odontoblasts; Am = ameloblasts. Scale bars = 100 µm.
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Related In: Results  -  Collection

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fig03: Decreased levels of DSP and DPP proteins, and Dspp gene expression in odontoblast of Col1a1-Trps1 mice. (A) Detection of SIBLING proteins isolated from dentin of 8-week-old WT and transgenic mice. Proteins were extracted from dentin and separated into 120 fractions using ion-exchange chromatography. Selected fractions containing SIBLING proteins were analyzed on an SDS-PAGE gel. SIBLING proteins were visualized by Stains-All. Numbers above the gel image indicate the chromatographic protein fractions analyzed on the gel; these fractions contained DPP and DMP1 fragments. The dashed-line box marks DPP protein, the solid-line box marks the N-terminal fragment of DMP1 protein. In transgenic mice a total level of DPP is decreased in comparison with WT mice, whereas levels of DMP1 show no difference. (B) Western blot with DSP antibodies shows decreased levels of DSP in transgenic dentin. (C) RNA in situ hybridization with probes specific for Dspp and Dmp1 (antisense molars, P2; sense molars P4). The boxed areas are magnified on the images in the right low corner of each respective image. In WT animals Dspp is expressed in both odontoblasts and ameloblasts, while in Col1a1-Trps1 mice Dspp expression in odontoblasts is significantly reduced. There is no difference in expression of Dmp1 between WT and transgenic odontoblasts. Od = odontoblasts; Am = ameloblasts. Scale bars = 100 µm.
Mentions: In contrast to DGI-I, in which both bone and dentin are affected, presentation of DGI-II and DGI-III is restricted to dental abnormalities. These forms of DGI are caused by mutations in the DSPP gene. The DSPP gene encodes a protein that is proteolytically processed to form DSP and DPP, which are the most abundant noncollagenous proteins in the dentin matrix. To understand the molecular basis of abnormal dentin formation in Col1a1-Trps1 mice, we compared the relative abundance of DSP, DPP, and other SIBLING proteins in the dentin matrices between WT and transgenic animals.30–32 These analyses showed dramatically reduced levels of DPP in transgenic animals (Fig. 3A). At the same time, levels of the N-terminal fragment of DMP1 showed no apparent differences as evaluated by Stains-All staining. Since DSP is less sensitive to Stains-All staining, we compared levels of DSP between WT and transgenic mice by Western blot. These analyses also showed dramatically decreased levels of DSP (Fig. 3B).

Bottom Line: On the molecular level, we demonstrated that sustained high levels of Trps1 in odontoblasts lead to dramatic decrease of Dspp expression as a result of direct inhibition of the Dspp promoter by Trps1.During tooth development Trps1 is highly expressed in preodontoblasts, but in mature odontoblasts secreting matrix its expression significantly decreases, which suggests a Trps1 role in odontoblast development.Thus, we provide novel insights into mechanisms of transcriptional dysregulation that leads to DGI.

View Article: PubMed Central - PubMed

Affiliation: Institute of Oral Health Research, Department of Oral and Maxillofacial Surgery, School of Dentistry, University of Alabama at Birmingham, Birmingham, AL 35294-0007, USA. dobrawan@uab.edu

Show MeSH
Related in: MedlinePlus