University of Pittsburgh
Grant: $378,750 - National Institutes of Health - Jul. 17, 2009
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Award Description: Abstract: The goal of this proposal is to extend and confirm our in vitro findings in vivo using mouse models to measure the impact of a-Syn on PP2A and TH with regard to DA synthesis. To achieve this, we will use multiple transgenic lines of a-Syn mice and assays utilizing recombinant proteins. In vivo we will modulate a-Syn levels by lentiviral transduction and then measure the effects of soluble and aggregated a-Syn on TH and PP2A by immunoblots, enzyme assays, immunohistochemistry, and confocal microscopy. In vitro we will use recombinant proteins to measure the impact of aggregated a-Syn on TH and PP2A activity as measured using well characterized enzymatic assays as we test the following hypotheses: 1) that a-Syn inhibits DA synthesis by effects on TH and PP2A in vivo, and 2) that soluble a-Syn stimulates PP2A and inhibits TH activity, while aggregated a-Syn is unable to do so, as measured in vitro and in vivo. We are uniquely positioned to elucidate the biological and translational relevance of a-Syn as a regulator of DA homeostasis, with the potential to identify novel preventative or restorative PD therapies.
Project Description: Aim 1 We have collected tissues from the various transgenic mice and are measuring: - TH activity by enzymatic assay - TH phosphorylation by immunohistochemistry and immunoblot *Preliminary findings suggest that, much like in vitro, a-Syn inhibits TH in vivo - PP2A activity by enzymatic assay *Preliminary findings suggest that, much like in vitro, a-Syn activates PP2A in vivo Aim 2 We have begun assaying the impact of aggregated a-Syn on PP2A activity: - Aggregation of a-Syn on Coomassie stained SDS-PAGE gels and on immunoblots - We saw less PP2A activation by aggregated a-Syn in enzymatic assays *Preliminary data indicate that aggregated a-Syn is less capable of stimulating PP2A activity.
Jobs Summary: Prime Recipient was able to retain a Research Assistant Professor, Assistant Professor, Research Associate and Visiting Scholar. (Total jobs reported: 2)
Project Status: Less Than 50% Completed
This award's data was last updated on Jul. 17, 2009. Help expand these official descriptions using the wiki below.
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