neurodegenerative diseases, NDD, proteinaceous deposits, Alzheimer’s disease, protofibrils, oxidative stress, aging, pathogenesis, expression vectors, yeast Transformation, Fluorescence Microscopy, Protein Expression, Serial Spotting, Rate of Aggregation, Mutagenesis, non-toxic protofibril model, dopaminergic degeneration
Parkinson's disease (PD) is characterized by the progressive death of dopaminergic neurons in the human brain. The misfolding and aggregation of alpha-synuclein, as well as the presence of reactive oxygen species (ROS), are throught to contribute to the cytoxicity. The mechanism of interaction between these two pathways is unknown. Mitochondrial dysfunction, specifically, incomplete respiratory metabolism and loss of antioxidants, has long been implicated as the culprit of ROS accumulation. Our lab has previously developed budding and fission yeast models to study genetic regulation of alpha-synuclein misfolding and toxicity. My thesis is composed of two studies. For my first goal, I tested the hypothesis that a mitochondrial dysfunction would enhance alpha- synuclein misfolding, aggregation, and toxicity. I used two budding yeast strains, one knocked-out for alpha-ketoglutarate dehydrogenase (the respiratory mutant) and mitochondrial glyoxalase II (the anti-oxidant mutant) to study the combinatorial effects of oxidative stress and expression of human alpha-synuclein. We found no cellular toxicity and decreased alpha-synuclein expression in both strains, yet intracellular aggregates only in the respiratory mutant. For my second foal, I tested the hypothesis that changing key amino acids within alpha-synuclein would affect its folding and toxic properties. Specifically, I investigated a mathematically modeled mutation of alpha-synuclein, Ala76a-Glu, which is thought to change its shape in a way that decreases its rate of aggregation. In fission yeast, A76E-alpha-synuclein formed aggregates more slowly than the normal form and did not change toxicity or expression. In budding yeast, A76E-alpha-synuclein reached the membrane more slowly, initially had decreased expression, and exhibited no toxicity. Results from these two studies assert that mitochondrial dysfunction and alpha-synuclein shapes play a role in the manifestation of Parkinson's disease.
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