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Mechanisms of Neurodegeneration in ALS and FTD

dc.contributor.authorFlores, Brittany
dc.date.accessioned2019-02-07T17:54:35Z
dc.date.availableNO_RESTRICTION
dc.date.available2019-02-07T17:54:35Z
dc.date.issued2018
dc.date.submitted2018
dc.identifier.urihttps://hdl.handle.net/2027.42/147577
dc.description.abstractAmyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are devastating neurodegenerative conditions that share key clinical, pathologic, and genetic characteristics. Neuronal inclusions rich in the RNA binding protein TDP43 are found in the majority of ALS and FTD. Moreover, the most common cause of familial ALS and FTD is a hexanucleotide (G4C2) repeat expansion mutation within the first intron of chromosome 9 open reading frame 72, or C9orf72. Mutant C9orf72 transcripts undergo repeat associated non-AUG (RAN) translation, generating five unique dipeptide repeat proteins (DPRs) that accumulate in degenerating neurons in C9orf72-associated ALS/FTD, but their significance in disease pathogenesis remains unclear. My dissertation investigates C9orf72 RAN peptides, TDP43 deposition, and their respective contributions to neurodegeneration. My central hypothesis is that C9orf72 RAN peptides disrupt TDP43 metabolism, leading to neurodegeneration via TDP43-dependent RNA misprocessing. My thesis addresses the molecular pathways responsible for neurodegeneration in ALS and FTD. Chapter 1 reviews central features of ALS and FTD, including an overview of the proposed mechanisms of C9orf72-related neurodegeneration and aspects of TDP43 deposition. I first determined whether C9orf72 RAN peptides, and more specifically which RAN peptides, are toxic to neurons. In collaboration with Dr. Magdalena Ivanova, we synthesized short polymers corresponding to the three sense-strand C9orf72 RAN products, analyzed their structures by electron microscopy and assessed their relative toxicity when applied to rodent primary cortical neurons. In doing so, we observed unique structural features for each dipeptide that correlated with their cellular internalization and relative toxicity. This work is described in further detail in Chapter 2. I next began investigating the intrinsic properties of TDP43 that are critical for downstream neuronal toxicity in disease models. TDP43 binds thousands of transcripts, particularly UG-rich sequences, and TDP43-dependent toxicity is tightly tied to its ability to recognize RNA. Intramolecular interactions between TDP43’s RNA binding domains, mediated by a salt bridge, are necessary for maintaining specificity for UG sequences. How sequence specificity of TDP43 binding to RNA affects TDP43 accumulation and survival remain unclear. Here, I show that genetically engineered mutations disrupting the TDP43 salt bridge reduce the affinity of nucleic acid binding and eliminate recognition of its native RNA targets. These same mutations dramatically destabilize TDP43, alter nuclear localization and abrogate toxicity upon overexpression in primary neurons. High-throughput RNA sequencing and splicing analyses indicated that TDP43 accumulation predominantly affects transcripts encoding components of the ribosome and oxidative phosphorylation pathways. These studies are illustrated in Chapter 3. Chapter 4 describes relevant preliminary work with implications of a connection between the mutant C9orf72 repeat expansion and TDP43 deposition. Briefly, I demonstrate that G4C2 oligonucleotides are recognized by TDP43 variants containing salt bridge-disrupting mutations, and co-expression of G4C2 and TDP43 enhance cytoplasmic mislocalization and neuronal toxicity. Chapter 5 concludes the dissertation outlining the next steps moving forward with this work. Taken together, this dissertation uncovers novel disease pathways that can be targeted for therapy development.
dc.language.isoen_US
dc.subjectamyotrophic lateral sclerosis
dc.subjectneurodegeneration
dc.titleMechanisms of Neurodegeneration in ALS and FTD
dc.typeThesisen_US
dc.description.thesisdegreenamePhDen_US
dc.description.thesisdegreedisciplineCellular & Molecular Biology
dc.description.thesisdegreegrantorUniversity of Michigan, Horace H. Rackham School of Graduate Studies
dc.contributor.committeememberBarmada, Sami J
dc.contributor.committeememberFuller, Robert S
dc.contributor.committeememberPaulson, Henry L
dc.contributor.committeememberTodd, Peter
dc.subject.hlbsecondlevelMolecular, Cellular and Developmental Biology
dc.subject.hlbtoplevelScience
dc.description.bitstreamurlhttps://deepblue.lib.umich.edu/bitstream/2027.42/147577/1/bnflores_1.pdf
dc.identifier.orcid0000-0002-5614-6901
dc.identifier.name-orcidFlores, Brittany; 0000-0002-5614-6901en_US
dc.owningcollnameDissertations and Theses (Ph.D. and Master's)


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