Date of Award

2015

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Neuroscience

First Advisor

Claire H. Mitchell

Abstract

Alzheimer’s disease (AD) is the most common form of dementia, leading to memory loss progressive cognitive decline over the course of what can be many years. Mutations in the catalytically active component of the γ-secretase complex, presenilin 1 (PS1), are the most common cause of familial Alzheimer’s disease (fAD), a less-prevalent but earlier-onset form of AD. PS1 mutation is associated with more severe lysosomal and autophagic pathologies than are found in sporadic AD; these pathologies may be a result of lysosomal pH dysregulation. The goal of this dissertation was to confirm a role for elevated lysosomal pH in cells from PS1-fAD patients, to investigate the repercussions of this lysosomal dysfunction, and to identify a therapeutic approach by which to restore both pH and lysosomal pathology to normal. Using human skin fibroblasts containing the PS1-fAD mutation A246E, the present work identified a small but significant increase in lysosomal pH in PS1-fAD mutant cells when compared to control fibroblasts. The pH data were supported by a reduction in mature cathepsin D (Cat D) and Cat D active site availability in PS1-fAD cells, as well as by substantial accumulation of autophagic substrates and up-regulation of components of the lysosomal and autophagic degradative systems, both at the mRNA and protein levels. Treatment with cAMP proved restorative, bringing lysosomal pH in PS1-fAD cells back to baseline while having minimal effect on control cells. cAMP increased Cat D active site availability, reduced autophagic backlog, and led both to mTOR phosphorylation and to down-regulation of genes involved in lysosomal function. Interestingly, cAMP-induced pH restoration, as well as Cat D increase and mTOR phosphorylation, was found to be PKA-dependent, suggesting a signaling pathway that may serve as a useful target for future treatment, and implicating PKA in the immediate, upstream response to cAMP treatment, and mTOR and gene expression modulation in the downstream response. This treatment proved effective in compromised lysosomes from primary neuronal cultures, as well, supporting the general utility of a cAMP-based approach. Together, these results identify lysosomal pH elevation as an important factor in AD pathology, and suggest several possible targets for future therapeutic investigation.

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