Expressing Alpha-Synuclein in RCSN-3 Cells to Improve the Study of Pathways in Parkinson’s Disease
Alpha-synuclein (αS) is a dopamine-regulating protein in the brain that can form aggregates, clusters of misfolded proteins, and cause neurodegeneration in the form of Parkinson’s Disease (PD). This ongoing research project explores the furthered development of the RCSN-3 cell line where it can express fluorescently tagged human αS. The development of this in vitro model will necessitate transfection of αS DNA, fluorescence microscopy, and differentiation of the cells into neurons, with the ultimate goal of having a better model to both screen small-molecule drugs that could inhibit αS aggregation and study other related pathways, such as the polyamine pathway. With an efficacious working model of dopaminergic neurons, we can now work to replicate the results of cell viability and aggregation assays accomplished by our lab in past years to show the applicability of the findings to the pathogenesis of PD.
We went about furthering the RCSN-3 cell line using DNA plasmid transfection protocols, seeding of preformed αS fibrils into confluent cell culture, and fluorescence microscopy to establish the viability of the cell line as a model. Our next steps involve differentiating the cells into neurons and replicating assays with previously established results to expand on data implicating the polyamine pathway in PD pathogenesis, as well as obtaining samples to send for Proteomics and Metabolomics analysis.