UC Santa Cruz

Traditionally regarded as integral players in mRNA translation, recent studies suggest that transfer RNA (tRNA) expression, processing, and modification are dynamically regulated in response to changing cellular conditions. These mechanisms facilitate the production of a wide variety of unique tRNAs and tDRs (tRNA-derived small RNAs) with the capacity to affect numerous cellular pathways. Increasing evidence implicates these transcripts as critical players in disease progression; however, due to technical limitations associated with studying these highly-modified RNAs in a high-throughput manner, their accurate quantification in relevant disease contexts and steady-state levels has proven difficult.Initially, I sought to determine the basal tissue variance of tRNA and tDR transcripts by systematically profiling the abundance and potential sites of base modification of tRNAs, tDRs, and other small RNAs using a newly developed small RNA sequencing method, Ordered Two-Template Relay sequencing (OTTR-seq), across a panel of 20 mouse tissues. In the first part of his dissertation, I identify novel tissue-biased abundance patterns of both tRNAs and tDRs, on a tissue-by-tissue basis. Using these mouse sequencing data and a variety of other eukaryotic sequencing data I generated, I developed a framework for identifying and semi-quantitating tRNA modifications leveraging the modification-induced misincorporations left by the reverse transcriptase. In this manner, these data provide one of the most precise maps of select tRNA modifications in various model organisms. With a focus on disease, I apply these methodologies to a mouse model of colorectal cancer model to identify tRNA transcripts, and tRNA-related proteins, potentially playing critical roles in disease progression. Additionally, as part of a collaborative effort in the final part of this dissertation, I worked to identify distinct stress-related profiles of intra- and extracellular tDRs. To determine the clinical implications for these, we identified some of these stress-related tDRs in patients' blood plasma following pulmonary bypass surgery. Furthermore, we determined the enzymes responsible for generating these tDR products through various perturbations under cellular stressors. In this manner, this dissertation provides the most detailed look at the changes in tRNA and tDR transcript profiles across cell types and aberrations in disease.