Stevens, Roy H.2020-11-022020-11-022012864885537http://hdl.handle.net/20.500.12613/2405Objective: To examine Dentine matrix protein 1(DMP1) and Dentine sialophosphoprotein (DSPP) expression and subcellular localization in various cell lines to better understand their function. Methods: RT-PCR, immunofluorescent staining and Western-blotting analyses were used to determine the expression and subcellular localization of DMP1 and DSPP in various cell lines, including odontoblast-like (17IIA11), preosteoblast (MC3T3-E1), mesenchymal cells (C3H10T1/2), and human dental pulp stem cells (DPSC). In addition, a haemagglutinin (HA) tagged DMP1 expression construct was generated and examined for its subcellular localization in COS-7 cells. Results: Western-blot analysis showed the presence of DMP1 and DSPP in the cytoplasmic and nuclear extracts of MC3T3-E1, 17IIA11, and C3H10T1/2 cells. DMP1 and DSPP transcripts were consistently detected in all three cell lines by RT-PCR analysis. However, immunofluorescent detection of DMP1 revealed the presence of two distinct subpopulations of cells with either nuclear or cytoplasmic staining; this phenomenon was not noticed with DSPP immunofluorescence. Nuclear and cytoplasmic DMP1 was confirmed in MC3T3-E1 cells by immuofluorescent staining using a rabbit polyclonal antibody; and the staining was inhibited when the antibody was preincubated with the synthetic peptide used to generate the antibody, confirming the specificity of the antibody. Nuclear and cytoplasmic localization was also observed in COS-7 cells transfected with HA-tagged DMP1 expression construct when detected with an antibody against the HA tag. Conclusion: These findings suggest that, apart from their role as a constituent of dentin/bone matrix, both DMP1 and DSPP might play a regulatory or structural role in the nucleus that is not unique to the odontoblast/osteoblast cells.53 pagesengIN COPYRIGHT- This Rights Statement can be used for an Item that is in copyright. Using this statement implies that the organization making this Item available has determined that the Item is in copyright and either is the rights-holder, has obtained permission from the rights-holder(s) to make their Work(s) available, or makes the Item available under an exception or limitation to copyright (including Fair Use) that entitles it to make the Item available.http://rightsstatements.org/vocab/InC/1.0/DentistryBiology, MolecularText