Pathogenic causes for progressive loss of dopaminergic neurons along with protein aggregates termed Lewy bodies in the substantia nigra in (SN) Parkinson’s disease (PD) are not yet understood. A protein called α-synuclein is the major component of the Lewy bodies. Familial mutations of α-synuclein lead to accelerated protein aggregation and cytotoxicity (1). Interestingly, α-synuclein undergoes a protein post-translational modification termed SUMOylation. It involves attaching the Small Ubiquitin MOdifier-like protein known as SUMO to the protein. Since SUMOylation is found to inhibit α-synuclein-mediated aggregation and cytotoxicity (2), to enhance SUMOylation levels may represent a promising therapeutic strategy against PD pathology. We have recently found that SUMO protease SENP3 negatively regulates SUMOylation levels via a process known as deSUMOylation where it detaches SUMO from SUMOylated proteins (3). This has led us to hypothesize that i) α-synuclein is a deSUMOylation substrate for SENP3, and (ii) SENP3-mediated deSUMOylation exacerbates PD neuropathies due to α-synuclein mutations. The findings from this project would better our understanding of molecular mechanisms underlying PD pathogenesis and identify new target(s) for future PD treatment.
The proposed work will directly test the hypothesis, and it will involve a combination of techniques, including those in molecular biology (e.g., cloning and tagging and site-directed mutagenesis), protein chemistry (e.g., glutathione S-transferase/Histidine pulldowns & co-immunoprecipitations followed by western blotting, protein purification and assays for SUMOylation/deSUMOyation), cell biology (e.g., cultures of clonal cell lines and neuronal cells, and DNA & sh/siRNA transfections/CRISPR-Cas9-mediated knockdown/knockout and replacement), together with In-cell quantitative analysis of the aggregation and the inclusion of α-synuclein under different SUMOylation statuses and biochemical examinations of cell viability (e.g., cytochrome c release, caspase activation, and MTT and LDH assays).
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