Telomere Length Studies In Human Cancer Cells
Buseman, Christen Marie
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Telomeres consist of repetitive DNA sequences and their associated binding proteins, and serve to protect linear chromosome ends from being recognized as double stranded breaks in need of repair. The telomeres of most normal diploid cells shorten with every cell division until they reach a critically short length, at which time the cells undergo senescence or apoptosis. Cancer cells which have the ability to divide indefinitely must prevent their telomeres from becoming critically short, and the majority of cancer cells achieve this by upregulating telomerase. Maintaining telomere length involves regulating the dynamic between telomere shortening and telomere elongation. However, there are still many aspects of this dynamic regulatory process that are unknown. Many methods of telomere length assessment have been developed that utilize a variety of molecular techniques, but a major shortcoming of these methods is that they lack the ability to detect single short telomeres that are thought to trigger replicative senescence. Thus, the objective of this work was to develop an assay, named Universal STELA, which can generate an accurate distribution of telomere lengths on all chromosomes and allow for the study of the shortest telomeres in experimental settings. Universal STELA was first used to determine if cancer stem cells are susceptible to telomerase inhibition therapy because they have a larger fraction of shorter telomeres than non cancer stem cells. Cancer stem cells are thought to contribute to cancer metastasis and recurrence, and therapies like telomerase inhibition that target cancer stem cells may lead to more durable treatment outcomes. Universal STELA was next used to investigate regulation of telomerase action. C- and G-STELA were used to determine that telomerase activity is coupled to telomere replication, while C-strand fill-in is delayed until S/G2. Universal STELA was used to compare the rate of elongation of short, average and long telomeres when telomeres are shorter than their maintenance length.