Abstract

This dissertation explores the connection between amyotrophic lateral sclerosis pathogenesis and RNA biology via two mechanisms: microRNA regulation of gene expression in the spinal cord and modulation of TDP-43 pathology through synthetic oligonucleotides. Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by selective loss of motor neurons in the brain and spinal cord that leads to progressive weakness. However, despite this shared clinical pattern the known causes of ALS are extremely varied. Only about 10% of ALS cases are familial while the other 90% occur sporadically. Even among that small fraction of familial cases, researchers have identified dozens of genes that are either causative or substantially increase risk. With this level of heterogeneity, there is a need to identify points of convergence so that new therapeutics can benefit the greatest number of patients. Aberrations in RNA regulation serve as one such point of convergence and are the target of the work described here. First, I detail our efforts to profile motor neuron microRNA (miRNA) expression in a mouse model of ALS and our subsequent experiments demonstrating the impact of one differentially regulated transcript, miR-146a, on disease progression. We first observed wide-ranging changes in motor neuron miRNA expression with disease progression, confirming that miR-146a expression is decreased specifically in motor neurons but increased in the whole spinal cord. Subsequently, we crossed miR-146a knockout mice to our ALS model and observed a survival benefit along with a reduction in spinal cord gliosis, though we did not observe motor neuron rescue. In fact, we observed that miR-146a knockout mice developed spontaneous motor neuron loss and paralysis with advanced age, and we demonstrated that miR-146a inhibition reduced motor neuron viability in culture. With this evidence in mind, we hypothesize that miR-146a loss has pleiotropic effects in the spinal cord with opposing influences on ALS progression, limiting damage caused by reactive glia while also sensitizing motor neurons. Next, I describe our progress towards designing a novel oligonucleotide-based approach to counteract the pathological mislocalization and aggregation TDP-43, an RNA-binding protein that shows characteristic pathology in 97% of ALS cases. We first showed that synthetic TDP-43-binding oligonucleotides counteracted aggregation in vitro and that this benefit scaled with oligonucleotide dosage and TDP-43 binding affinity. Moving to a mouse model of TDP-43 proteinopathy, we demonstrated that these TDP-43-binding oligonucleotides were able to distribute widely in the CNS and modestly improved nuclear localization early in disease. However, we did not observe a reduction in TDP-43 aggregation, nor did we observe a behavioral benefit. Taken together, these projects outline two distinct approaches to approaching aberrations in RNA processing during ALS. Better understanding miRNA regulation of the disease provides opportunities to broadly affect gene expression and improve motor neuron resiliency, while TDP-43-binding oligonucleotides offer a means of directly targeting pathology shared by almost all patients with ALS.

Committee Chair

Timothy Miller

Committee Members

Albert A. Davis; Paul H. Taghert; Conrad C. Weihl; Gregory F. Wu

Degree

Doctor of Philosophy (PhD)

Author's Department

Biology & Biomedical Sciences (Neurosciences)

Author's School

Graduate School of Arts and Sciences

Document Type

Dissertation

Date of Award

4-15-2026

Language

English (en)

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