Electronic Thesis and Dissertation Repository

Thesis Format

Monograph

Degree

Master of Science

Program

Physiology and Pharmacology

Supervisor

Betts, Dean H

2nd Supervisor

Watson, Andrew J

Co-Supervisor

Abstract

Reactivation of the multi-subunit ribonucleoprotein telomerase is the primary telomere maintenance mechanism in cancer, but it is rate-limited by the enzymatic component, telomerase reverse transcriptase (TERT). While regulatory in nature, TERT alternative splice variant/isoform regulation and functions are not fully elucidated and are further complicated by their highly diverse expression. In this thesis, I characterized TERT expression across normal and neoplastic tissues using TCGA and GTEx RNA-sequencing data. In doing so, I demonstrated the global overexpression and splicing shift towards full-length TERT in neoplastic tissue. Furthermore, my studies identified tumour subtype expression differences possibly regulated by subtype-specific characteristics, detailed heterogeneity in both isoform function and prognostic potential and determined cancer cell lines with representative tumour specific TERT transcriptomes. Taken together, my work reinforced the need for tissue specific TERT investigations, provided avenues to do so, and brought to light the current technical limitations of bioinformatically analyzing TERT isoform expression.

Summary for Lay Audience

When human cells divide, they replicate their DNA. However, the method used is imperfect and slightly shortens the DNA ends during each replication. To protect important genes, the ends are filled with repeated sequences called telomeres. While telomeres can increase the number of possible healthy replications, they also shorten with each cell division. Most adult cells with critically short telomeres will stop replicating and die off. However, more important cell types that replicate often, like stem cells, avoid cell death by re-lengthening their telomeres using the protein complex telomerase. The motor behind telomerase activity is telomerase reverse transcriptase (TERT). Unfortunately, cancer cells hijack this mechanism to continue dividing indefinitely. One possible way this is done is by changing the alternative splicing of the TERT. Normally, when genes are expressed the transcripts are cut and pasted (alternatively spliced) into different orientations, called isoforms. Some isoforms have the expected gene function, while others have new functions or no function. Adult cells express TERT, but mostly alternatively spliced non-functional isoforms. Therefore, cancer cells may change splicing back towards the functional isoform, amongst increasing overall expression. Complicating matters, the splicing and functions of some isoforms seem to differ between tissue types. In order to investigate this, I compared the expression of TERT isoforms in 33 cancer and 19 normal tissue types. In doing so, I confirmed that cancer cells typically increase overall TERT expression and shift splicing towards the functional isoform, whereas normal cells shift splicing towards non-functional isoforms. In addition, I identified normal tissue types with uncharacteristically increased TERT expression, indicating the possibility of another important function outside of telomere re-lengthening. As well, I identified cancer-specific subtypes with different expression patterns and outlined possible reasons as to why this may occur based on their subtype characteristics. Finally, to potentially guide future investigations, I matched tumours with the most representative cancer cell lines based on TERT isoform expression patterns. Taken together, my work set the groundwork for and reinforced the need for tissue-specific investigations, provided avenues to do so, and brought to light the current limitations of bioinformatically analyzing TERT isoform expression.

Creative Commons License

Creative Commons Attribution 4.0 License
This work is licensed under a Creative Commons Attribution 4.0 License.

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