Uncovering the Function of TDP43 Splice Variants in Physiology and Neurodegeneration

Abstract

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are devastating neurodegenerative conditions lacking comprehensive treatment options. ALS is a disease of the upper and lower motor neurons, causing muscle atrophy that eventually leads to paralysis and respiratory failure. FTD, on the other hand, is characterized by the loss of neurons in the frontal and temporal lobes of the brain, which drives personality changes, language impairments, and cognitive decline. Although these conditions are seemingly unrelated, there is a growing body of evidence to suggest significant clinical, pathological, and genetic overlap between ALS and FTD. Consistent with this, ALS and FTD share a pathological hallmark characterized by the mislocalization and aggregation of the RNA-binding protein TDP43 (Transactive response DNA/RNA-binding protein, 43 kDa), which is observed in more than 95% of ALS patients and approximately half of FTD patients. Moreover, mutations in several other ALS/FTD-associated genes result in TDP43 pathology, highlighting a fundamental connection between TDP43 and these conditions. TDP43 is a ubiquitously expressed nuclear protein that serves an essential role in RNA processing. As such, small alterations in TDP43 localization and abundance are sufficient to disrupt critical biological processes. To compensate for this, TDP43 regulates its own levels through a negative feedback loop wherein TDP43 binds its own 3’ untranslated region (3’UTR), triggering alternative splicing of TARDBP, the gene that encodes TDP43, and the generation of multiple alternative TDP43 isoforms that are targeted for degradation. We previously found that two of these splice variants, termed “short” (s)TDP43 due to a C-terminal truncation, accumulate in the cytosol and are enriched in vulnerable motor neurons. Moreover, through its intact N-terminus, sTDP43 can sequester full-length (fl)TDP43 within cytoplasmic inclusions reminiscent of those in disease, indicating that sTDP43 production may contribute to the susceptibility of motor neurons in ALS/FTD. This dissertation focuses on uncovering the regulation of sTDP43 variants, as well as their potential function and contribution to ALS/FTD pathogenesis. Chapter 1 begins with a review of the past and present literature surrounding sTDP43 isoforms, focusing specifically on sTDP43 regulation by nonsense-mediated decay, neuronal hyperactivity, as well as multiple post-translational pathways dictating sTDP43 clearance. I also review sTDP43 localization, as well as phase separation and RNA binding abilities to predict the cellular function of these variants. In Chapter 2, I conduct an in-depth analysis of the transcriptional and post-translational pathways responsible for sTDP43 regulation. Moreover, I examine the specific domains responsible for sTDP43-mediated toxicity and identify splicing abnormalities in disease-associated transcripts upon sTDP43 accumulation. In Chapter 3, I explore the abundance and distribution of sTDP43 in secretory tissue and provide the first evidence for endogenous sTDP43 and flTDP43 secretion in saliva. Chapter 4 summarizes key findings from this research and discusses open questions surrounding sTDP43 stability, localization, and function. Appendix A provides an overview of strategies for selective elimination of sTDP43, and Appendix B contains a scientific commentary describing the membraneless organelles in which TDP43 has been detected. Taken together, this dissertation characterizes the regulation, potential function, and peripheral expression of sTDP43 isoforms, as well as provides insights into a probable role for sTDP43 in neurodegenerative disease.