The role of BRCA1 in telomere maintenance

Abstract

Telomeres are fundamental structures found at the end of all eukaryotic chromosomes that function to protect the end of chromosomes from end-to-end fusion, erosion and subsequent telomere dysfunction. Telomerase and alternative lengthening of telomere (ALT) mechanisms maintain the telomeres by compensating natural telomeric loss. ALT is found to be present in 15% of human tumours lines and it may be expressed at low levels in the normal mouse tissues. However, the exact mechanism behind ALT depression and/or activation in the mammalian cells is not fully understood. Previous studies have highlighted the role of BRCA1 in telomere dysfunction. Also, it has recently been shown that BRCA1 co-localises at telomeres in the ALT + human cells through BLM and Rad50. However, it is still unclear whether BRCA1 plays a direct role on telomere length maintenance and integrity. The aim of this project was to examine the role of BRCA1 in telomere maintenance associate with ALT in BRCA1 defective mammalian cells. Therefore to achieve this, we have set up series of experiments to look at, (a) hallmarks of ALT activity at the cytological level, (b) measuring of ALT activity using biochemical and immunocytochemistry techniques and (c) understanding the role of BRCA1 in DNA damage response mechanism and telomere dysfunction. Firstly, we found elevated levels of recombination at telomeres in the two human BRCA1 carrier cell lines and mouse embryonic stem cell with deficiency in Brca1-/-. Secondly, our data showed that human and mouse BRCA1 defective cells are significantly more sensitive to ionizing radiation in line with the DNA repair function of BRCA1. Moreover, we found persistent DNA damage at telomeres in the BRCA1 defective environment when after exposure of cells to ionizing radiation. Thirdly, we found evidence of ALT activity in some mouse cell lines, and elevated ALT in mouse cells defective in Brca1. Finally, we examined some other ALT markers using immunofluorescence. Our data indicate differences between human and mouse cells in regulating ALT. Taken together data presented in this thesis revealed that (i) BRCA1 plays a major role in telomere maintenance and defective BRCA1 mammalian cells show evidence of telomere dysfunction and telomere length shortening in line with previous publish data, (ii) BRCA1 defective mouse cells have elevated levels of ALT, (iii) the mouse lymphoblastoid LY-S cells have complete absence of ALT.