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High-Throughput Approaches To Understanding RNA Processing


Nascent RNAs are immediately coated in proteins, termed RNA binding proteins (RBPs) that regulate RNA transcription, splicing, localization, and turnover. Because of their ubiquitous nature, RBPs both directly and indirectly influence every aspect of cellular function, and therefore it is unsurprising that their misregulation leads to disease. Understanding the delicate and complex interplay between RBPs and their targets is not only critical to elucidating fundamental biological processes, but also essential to identifying mechanisms of pathology and possible therapeutic targets. This dissertation explores the application of high-throughput sequencing technologies and the use and development of downstream computational approaches to understanding RBP-mediated RNA processing. Furthermore, because the brain has the greatest abundance and diversity of RBPs, neurons and neural tissue are of particular interest when considering systems in which to study RBP:RNA interactions. This dissertation will examine known relationships between RNA processing and neurodegeneration as well as applications of high-throughput techniques to the characterization of RNA processing. I will review several cutting-edge, high-throughput sequencing technologies used in the dissection of RBP-RNA interactions. I will also discuss how these technologies have been or could be applied to understanding how the misregulation of RBPs leads to aberrant RNA processing and subsequently causes neurodegenerative disease. In trying to understand RBP-mediate RNA processing, I have developed a novel high-throughput computational approach to leveraging enhanced crosslinking and immuno-precipitation (eCLIP) datasets to identify regulators of microRNA (miRNA) biogenesis. In doing so, I have identified several regulators of miRNA biogenesis.

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