Molecular cloning of wild-type and mutant Osterix and its localization in MC3T3-E1 cells
Date
2013
DOI
Authors
Morcos, Joseph Mounir
Version
OA Version
Citation
Abstract
Bones are crucial organs in the human body - deviations from proper formation results in harmful and sometimes lethal effects. Thus, it is imperative that the regulatory factors of bone formation be investigated. One such factor is Osterix/SP7 (OSX), a Zinc-finger-containing transcription factor required for bone formation first determined in Osx null mice. In patients with mutant Osterix (MT-OSX), bone was malformed. Hence it is clear that wild-type (WT)-OSX and MT-OSX have different effects on the biological mechanisms involved in bone formation. This study aims to identify these differences at the cellular level by investigating the localization of WT-OSX and MT-OSX by confocal immunofluorescence microscopy. WT and MT forms of OSX were first cloned into pcDNA3-FLAG tagged vectors. Over expression of WT-OSX and MT-OSX were observed and indicated that both WT-OSX and MT-OSX (frameshift mutation at c.1052) experienced post translational modifications. In order to visualize the localization of WT-OSX and MT-OSX in the cell, pcDNA3-FLAG-WT-OSX and pcDNA3-FLAG-MT-OSX were transfected into MC3T3-E1 cells. Cells were stained based on immunofluorescence methods and it was concluded that MT-OSX was unable to enter into some parts of the nucleus. Although the exact locations of WT-OSX and MT-OSX were not determined, there is a clear difference in the localization of WT-OSX and MT-OSX in the cells; this ultimately affects the transcriptional properties of MT-OSX which inhibits proper bone formation.
Description
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Thesis (MSD) --Boston University, Henry M. Goldman School of Dental Medicine, 2013 (Department of Periodontology and Oral Biology).
x, 47 leaves : ill.
Includes bibliographic references: leaves 39-44.
Thesis (MSD) --Boston University, Henry M. Goldman School of Dental Medicine, 2013 (Department of Periodontology and Oral Biology).
x, 47 leaves : ill.
Includes bibliographic references: leaves 39-44.
License
This work is protected by copyright. Downloading is restricted to the BU community. If you are the author of this work and would like to make it publicly available, please contact open-help@bu.edu.