Biochemical fractionation of induced pluripotent stem cell derived motor neurons from an ALS patient with the Glycine-298-Serine TDP-43 mutation
Goldszer, Isaac M.
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Transactive response DNA-binding protein (TDP-43), and fused in sarcoma/translocated in liposarcoma (FUS/TLS) form protein aggregates in amyotrophic lateral sclerosis (ALS) and fronto-temporal lobar degeneration (FTLD). The sequencing of the TDP-43 gene TARDBP in a large patient population has shown more than 40 missense mutations that are now known to cause disease. The effect of genetic mutation on protein aggregation, and the pathogenesis of ALS is the focus of this study. In order to determine the effect of the TARDBP Glycine-298- Serine (G298S) missense mutation on protein aggregation in disease, induced pluripotent stem cells (iPSCs) were reprogrammed from control and G298S mutant fibroblasts, and differentiated into motor neurons using defined factors, and fractio- nated to determine the soluble and insoluble TDP-43 burden. There was an increase in insoluble TDP-43 in the ALS-patient-derived motor neuron lysates over a normal control, but the significance could not be assessed because of the small sample size. A toxicity assay using fluorescence activated cell sorting showed an unexpected trend towards healthier control neurons. Future studies should include quantified immunohistochemical analysis of motor neurons and use novel pharmaceuticals to attempt to correct aberrant TDP-43-mediated RNA processing.
Thesis (M.A.)--Boston University