Elucidating the role of RNA oxidation on amyotrophic lateral sclerosis onset and progression

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PROJECT SUMMARY A growing body of evidence implicates oxidative stress in Amyotrophic Lateral Sclerosis (ALS). Nevertheless, it is currently unknow if it is a cause, a by-product or a consequence of disease. The effects of oxidative stress on cellular damage caused by reactive oxygen species is usually attributed to modifying proteins and DNA. However, RNA oxidation occurs ten times more often than DNA oxidation. Importantly, high levels of oxidized RNA are detected in relevant neuronal tissues of patients with ALS while, mouse models of ALS show increased RNA oxidation in motor neurons of the spinal cord at an early pre-symptomatic stage. What remains lacking, however, is an understanding of the functional relationship between RNA oxidation and ALS onset and progression. Thus, there is a critical need to identify which motor neuron transcripts are oxidized in early, pre- symptomatic stages of ALS and how this dysregulation contributes to neuronal death and other molecular hallmarks of ALS. I believe that there are many, yet to be discovered, RNA Binding Proteins (RBPs) that are crucial for controlling the fate of oxidized RNAs. I hypothesize that RNA oxidation drives motor neuron degeneration in ALS by dysregulating proper RNA processing by RBPs. I will test this hypothesis by (1) elucidating, in iPSC-derived motor neurons, the RNA targets and the consequences of depletion of known RBPs that interact with oxidized RNAs and identify novel ones; (2) identifying and comparing oxidized RNAs in iPSC- derived motor neuron models; and (3) investigating, at a single cell resolution, the effects of RNA oxidation on transcription, translation and RBP-RNA interactions in iPSC-derived spinal organoid ALS models. If successful, this project will generate the foundational methods and insights to enable early, pre-symptomatic-stage diagnostic approaches and interventions to reduce RNA oxidation levels in high-risk individuals. My background in DNA damage and repair and single cell transcriptomics together with the Yeo lab’s expertise on RNA processing and neurodegeneration make me an ideal candidate to accomplish the research proposed above. These three aims will serve as a basis for my independent academic position generating the foundational methods and insights to study the effects of RNA damage. The Yeo lab at UCSD is an ideal environment to perform this research and complete my training towards pursuit of an independent academic faculty position, as it has consistently been a leader in developing both experimental and computational methods to characterize RNA processing and RBP regulation. Additionally, the location of the Yeo lab proximal to outstanding researchers at UCSD, the Salk Institute, and other research institutes and biotechnology companies in La Jolla will provide ample opportunities for mentored training in performing research and developing an independent research program.