Environmental pesticide exposure is connected to the occurrence of Parkinson's disease (PD), the most common neurodegenerative movement disorder in America. Epidemiological investigations indicate exposure to the pesticides paraquat (PQ) and maneb (MB) increase the risk of PD, but the underlying mechanisms are unknown. Mitochondrial oxidative stress appears to be central to environmental causes of PD, which is supported by research on the antioxidant glutathione (GSH). However, recent data show that peroxiredoxin 3 (Prx3), a peroxidase supported by the antioxidant thioredoxin-2 (Trx2), has a predominant role in mitochondrial peroxide metabolism. Oxidation of Trx2 activates cell death mechanisms through the mitochondrial permeability transition and activation of apoptosis signal regulating kinase-1 (ASK1). The proposed research is designed to investigate the hypothesis that the Trx2/Prx3 system is an important mitochondrial antioxidant system in the substatntia nigra that is disrupted by MB-potentiated PQ toxicity. To accomplish this, two specific aims consisting of in vitro and in vivo models will be used. In Aim 1, dopaminergic SH-SY5Y cells will be treated with PQ, MB or in combination to evaluate stress responses, potentiation of Trx2/Prx3 oxidation and activation of cell death pathways. This research will involve specific mechanistic targets of mitochondria and mass spectrometry-based proteomic analyses. To examine MB-mediated effects, aldehyde dehydrogenase activity and intracellular copper accumulation will be measured. In Aim 2, a genetic mouse model (Prx3 over- expressing mouse) will be used to study specific mitochondrial redox effects of PQ + MB in vivo. Abundance and compartmental redox effects will be measured by mass spectrometry-based proteomics and discovery- based metabolomic analyses. This research will provide a foundation for future translation of results into human studies, where archival samples from exposed farm workers are available. Lastly, these studies will advance the understanding of the mechanistic role of the Trx2/Prx3 system in pesticide-mediated neurodegeneration.