Date of Award
Doctor of Philosophy (PhD)
Edward B. Lee
Aging-related neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS) and frontotemporal degeneration (FTD) are two fatal progressive neurodegenerative diseases that carry genetic and pathologic overlap: a hexanucleotide G4C2 repeat expansion in C9orf72 and loss of a nuclear RNA binding protein, TAR DNA binding protein-43 (TDP-43), into cytoplasmic aggregates. The C9orf72 expansion is the most common genetic cause of ALS/FTD and is associated with reduced C9orf72 expression and accumulation of toxic RNA and protein aggregates. In Chapter 2 of my thesis, using molecular analyses from human post-mortem ALS/FTD brain, I show that the C9orf72 promoter is hypermethylated within a subset of expansion carriers. C9orf72 promoter hypermethylation is associated with reduced C9orf72 pathology and may be protective in these patients. Another key feature in ALS/FTD is the characteristic pathology of nuclear TDP-43 loss in degenerating neurons. TDP-43 is a ubiquitous nuclear RNA binding protein and is heavily involved in RNA processing. TDP-43 has been shown to bind genic elements and repetitive transposable elements such as long interspersed nuclear elements (LINE). Considering that TDP-43 is a ubiquitous RNA binding protein, I hypothesize that nuclear TDP-43 loss can lead to large transcriptomic changes and may contribute to alterations in LINE elements. For Chapters 3 and 4 of my thesis, I use a novel method of subcellular fractionation and fluorescent activated cell sorting (FACS) from post-mortem ALS/FTD human brain to perform high-throughput sequencing analyses to study neuronal molecular changes. In Chapter 3 of my thesis, I use FACS coupled with RNA-seq on neuronal nuclei with and without TDP-43 to show that loss of nuclear TDP-43 is associated with large transcriptome changes and increased LINE accessibility. Furthermore, loss of nuclear TDP-43 leads to increased retrotransposition. I also extend this subcellular fractionation-FACS method to study the effects of the C9orf72 expansion in neuronal nuclei. In Chapter 4, I demonstrate that the C9orf72 expansion is linked to mild gene expression changes that reflect C9orf72 protein loss and not gain of toxic C9orf72 RNA. Through my work, I have shown disease mechanisms linked to repetitive DNA elements, in that I propose (1) the C9orf72 repeat expansion may contribute to disease primarily via a gain of toxic C9orf72 pathology (2) loss of neuronal nuclear TDP-43 may be associated with increase retrotransposon activity which may contribute to disease. Overall, my work has broadened the field of neurodegeneration in my implementation of cell-type specific molecular analyses on post-mortem brain of ALS/FTD patients to identify disease mechanisms with the intent of discovering new therapeutics and biomarkers that can be extended into the clinic.
Liu, Elaine, "Molecular Analysis Of Amyotrophic Lateral Sclerosis And Frontotemporal Degeneration Brain Tissue Identifies Disease Mechanisms Associated With Repetitive Dna Elements" (2017). Publicly Accessible Penn Dissertations. 2697.