Creation and establishment of transgenic mouse models for for Mecp2 gene, causing Rett syndrome
Year: 2007
Type: dissertation
Abstract: The MeCP2 protein is a transcriptional repressor binding to methylated cytosines and modifying the chromatin structure. Mecp2 gene was eventually found to be the cause of 95% of Rett Syndrome cases. In spite of its ubiquitous expression, it is believed to have some unique functions in the neurons. But till now, very less is known about the role of MeCP2 in neuronal cells and the mechanism by which it brings about the deteriorating disorder. The aim of the project was to produce two different transgenic mouse models expressing Mecp2 tagged with EGFP as a reporter to further characterize Mecp2 and obtain an appropriate functional analysis. The transgenic constructs have been generated by engineering a BAC clone (B22804) containing the entire Mecp2 gene of the mouse and flanking sequences, by GET recombination. First mouse model overexpresses the wild type Mecp2 labeled at C-terminus with EGFP protein. The second transgenic model overexpresses an early truncated (human mutation) Mecp2 protein, lacking the nuclear localization signal, also labeled at the C-terminus with the EGFP protein. The investigation of the transgenic mouse lines shall allow the spatio-temporal analysis of the Mecp2 protein by tracing the EGFP protein. Moreover, the transgenic mouse model which ectopically expresses an early truncated Mecp2 protein will help to understand the role of the mutated protein in the pathogenesis of the Rett Syndrome. Although Mecp2 is more predominantly expressed in the brain, it doesn"t seem to express in all the cells including glial cells. So, the Mecp2+/EGFP+ specific brain cells were isolated by Fluorescence Activated Cell Sorting (FACS) using our unique transgenic mouse models and the identity of them could be characterized using different markers by immunostaining. A transcriptomics approach using the Mecp2+/EGFP+ cell sub-population between the two transgenic models would validate the regulation of already found target genes or determine new target genes specifically. Further analysis will be done in the following directions: Live cell imaging of Mecp2 at the synapses, analysis of Mecp2+/EGFP+ cells in the respiratory network through the mouse development, and determining the size of the neurons expressing the transgenic proteins.
DOI: 10.53846/goediss-205
Author: Arunachalam, Jayamuruga Pandian
Language: English
Subfield: Genetics
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W2991506