Year: 2010
Type: dissertation
Abstract: The nuclear matrix (NM) is proposed to be a permanent network of core filaments underlying thicker fibres, present regardless of transcriptional activity.It is found to be both RNA and protein rich; indeed, numerous important nuclear proteins are components of the structure.In addition to mediating the organisation of entire chromosomes, the NM has also been demonstrated to tether telomeres via their TTAGGG repeats.In order to examine telomeric interactions with the NM, a technique known as the DNA halo preparation has been employed.Regions of DNA that are tightly attached to the structure are found within a so-called residual nucleus, while those sequences forming lesser associations produce a halo of DNA.Coupled with various FISH methodologies, this technique allowed the anchorage of genomic regions by the NM, to be analysed.In normal fibroblasts, the majority of chromosomes and telomeres were extensively anchored to the NM.Such interactions did not vary significantly in proliferating and senescent nuclei.However, a decrease in NM-associated telomeres was detected in quiescence.Since lamin A is an integral component of the NM, it seemed pertinent to examine chromosome and telomere NM-anchorage in Hutchinson-Gilford Progeria Syndrome (HGPS) fibroblasts, which contain mutant forms of lamin A. Indeed, genome tethering by the NM was perturbed in HGPS.In immortalised HGPS fibroblasts, this disrupted anchorage appeared to be rescued; the implications of this finding will be discussed.This study also suggested that telomere-NM interactions are aberrant in X-linked Emery-Dreifuss Muscular Dystrophy (X-EDMD), which is caused by mutant forms of emerin, another NM-associated protein.The positioning of selected genes in control and X-EDMD cell lines was examined in unextracted nuclei using 2D and 3D FISH.Subtle shifts in the organisation of these genes were detected in diseased cells; however, their expression levels remained unaltered.Furthermore, in order to examine the architectural integrity of the nuclear lamina in lamin A and emerin mutant cell lines, scanning electron microscopy (SEM) was funded my PhD research.
Author: Lauren S. Godwin
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W173176439