Cells exposed to DNA damaging agents activate a network of mechanisms called DNA damage response, including telomere length regulation. Telomeres are specialized structures that protect chromosome ends from degrading and being fused together. Mouse-knockout experiments revealed that cell lines deficient of DNA-PKcs or Ku70/80 resulted in high amount of telomere end-to-end fusion. Numerous other studies have shown a functional interplay between DNA damage response and telomere maintenance. The aim of this project is to examine this interplay further by investigating mechanisms of DNA damage response, using cell lines from X-linked homozygous recessive form of Dyskeratosis Congenita (DC) patients, which have dysfunctional telomere maintenance. DC is a multi-system disorder characterised by abnormalities of the bone marrow, immune deficiency and a predisposition to cancer. In this work we have shown that cells with defective DKC1 (the gene implicated in the X- linked homozygous recessive form of DC) exhibit a defective DNA damage response by examining two types of cells: fibroblast and lymphoblastoid cell lines. By using various biomarkers (H2AX, TIF assay etc) we analysed the DNA damage response by exposing DC cell lines to ionizing radiation. Our results demonstrated that DC cell lines have an abnormal DNA damage response and as a result show radiosensitivity. We have also knocked down the DKC1 gene in normal cell lines using siRNA oligonucleotides and demonstrated that this knock-down causes radiosensitivity. Therefore our results conclusively show an abnormal DNA damage response in cells derived from DC patients. Finally we used TA-65, a novel telomerase activator derived from the plant Astragalus membranaceus and showed radioprotective effects of this compound in normal lymphoblastoid cell lines. Taken together our results potentiate further the link between telomere maintenance and DNA damage response.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:687649 |
| Date | January 2016 |
| Creators | Sharma, Chetana Devi |
| Contributors | Slijepcevic, P. ; Parris, C. |
| Publisher | Brunel University |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://bura.brunel.ac.uk/handle/2438/12729 |
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