Date of Award
PhD (Doctor of Philosophy)
Dr Leonie Young
Telomere maintenance is essential for the continued proliferation of tumour cells and is performed through the mechanism of telomerase activation or by a recombination-based process referred to as alternative lengthening of telomeres (ALT). The most important components of active telomerase are the reverse transcriptase enzyme (hTERT) and the non-coding RNA subunit (hTR). hTERT is the rate-limiting component and is highly regulated. The detection of genes that regulate either telomere maintenance mechanism is therefore of potential therapeutic significance. To address this, the study used stable isotope labelling of/with amino acids in cell culture (SILAC) to compare the proteomes of 2 cell lines, 6G and 1-3C that had been previously derived from a single SV4O-immortalised jejunal fibroblast cell. Both cell lines are therefore genetically similar except for the fact that the 6G and 1-3C lines use different TMMs (telomerase and ALT respectively) to counteract telomere attrition. An ATPase-dependent DNA helicase protein called RuvBI2 was found to be relatively overexpressed in the 6G cell line. RuvBI2 is known to be involved in the control of gene expression and a putative role in hTERT regulation was investigated. siRNA knockdown of RuvBI2 reduced hTERT protein levels in the 6G cell line. RuvB12 was able to interact with the hTERT promoter and was also shown to upregulate hTERT gene transcription. Collectively the data in this study suggest that RuvBI2 is a positive regulator of hTERT gene expression and this observation supports the existing body of data that RuvBI2 has a multitiude of functions that contribute to the tumorigenesis.
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Flavin P. RuvB12 as a regulator of hTERT [PhD Thesis]. Dublin: Royal College of Surgeons in Ireland; 2010