The Mitochondrial Chaperone Protein TRAP1 Mitigates a- Synuclein Toxicity Erin K. Butler1,2,3., Aaron Voigt1., A. Kathrin Lutz4, Jane P. Toegel1, Ellen Gerhardt2, Peter Karsten 1 , Bjo¨ rn Falkenburger1, Andrea Reinartz5, Konstanze F. Winklhofer4,6, Jo¨ rg B. Schulz1,7* 1 Department of Neurology, University Medical Center, RWTH Aachen, Aachen, Germany, 2 Department of Neurodegeneration and Restorative Research, Center Molecular Physiology of the Brain (CMPB), Georg-August University Go¨ ttingen, Go¨ ttingen, Germany, 3 Go¨ ttingen Graduate School for Neurosciences and Molecular Biology (GGNB), Go¨ ttingen, Germany, 4 Neurobiochemistry, Adolf-Butenandt-Institute, Ludwig Maximilians University, Munich, Germany, 5 Department of Pathology, University Medical Center, RWTH Aachen, Aachen, Germany, 6 German Center for Neurodegenerative Diseases (DZNE), Munich, Germany, 7 J u¨lich-Aachen Research Alliance (JARA) Brain, Ju¨ lich/Aachen, Germany Abstract Overexpression or mutation of a-Synuclein is associated with protein aggregation and interferes with a number of cellular processes, including mitochondrial integrity and function. We used a whole-genome screen in the fruit fly Drosophila melanogaster to search for novel genetic modifiers of human [A53T]a-Synuclein–induced neurotoxicity. Decreased expression of the mitochondrial chaperone protein tumor necrosis factor receptor associated protein-1 (TRAP1) was found to enhance age-dependent loss of fly head dopamine (DA) and DA neuron number resulting from [A53T]a-Synuclein expression. In addition, decreased TRAP1 expression in [A53T]a-Synuclein–expressing flies resulted in enhanced loss of climbing ability and sensitivity to oxidative stress. Overexpression of human TRAP1 was able to rescue these phenotypes. Similarly, human TRAP1 overexpression in rat primary cortical neurons rescued [A53T]a-Synuclein–induced sensitivity to rotenone treatment. In human (non)neuronal cell lines, small interfering RNA directed against TRAP1 enhanced [A53T]a- Synuclein–induced sensitivity to oxidative stress treatment. [A53T]a-Synuclein directly interfered with mitochondrial function, as its expression reduced Complex I activity in HEK293 cells. These effects were blocked by TRAP1 overexpression. Moreover, TRAP1 was able to prevent alteration in mitochondrial morphology caused by [A53T]a-Synuclein overexpression in human SH-SY5Y cells. These results indicate that [A53T]a-Synuclein toxicity is intimately connected to mitochondrial dysfunction and that toxicity reduction in fly and rat primary neurons and human cell lines can be achieved using overexpression of the mitochondrial chaperone TRAP1. Interestingly, TRAP1 has previously been shown to be phosphorylated by the serine/threonine kinase PINK1, thus providing a potential link of PINK1 via TRAP1 to a-Synuclein. Citation: Butler EK, Voigt A, Lutz AK, Toegel JP, Gerhardt E, et al. (2012) The Mitochondrial Chaperone Protein TRAP1 Mitigates a-Synuclein Toxicity. PLoS Genet 8(2): e1002488. doi:10.1371/journal.pgen.1002488 Editor: Harry T. Orr, University of Minnesota, United States of America Received April 7, 2011; Accepted December 2, 2011; Published February 2, 2012 Copyright: 2012 Butler et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: EKB was supported by a European Union Marie Curie Action doctoral stipend, Neuroscience Early Stage Research Training (NEUREST). This work is funded by the BMBF (NGFN+: 01GS08137 to JBS and AV and 01GS08139 to KFW; KNDD: 01GI0703 to JBS) and DFG grants to KFW (SFB 596, A14) as well as grants from the Helmholtz Alliance ‘‘Mental Health in an Aging Society.’’ The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: jschulz@ukaachen.de . These authors contributed equally to this work. Introduction Parkinson’s disease (PD) is the second most prevalent neurode- generative disease behind Alzheimer’s disease (AD), with an incidence rate of approximately 110–300 per 100,000 persons above the age of 50 [1]. The movement disorder is characterized by the selective death of dopaminergic neurons in the substantia nigra pars compacta (SNc) [2]. Death of SNc neurons results in a reduction of dopamine (DA) levels within their key efferent target, the striatum [3]. Mitochondrial Complex I activity deficit and evidence of enhanced oxidative stress within affected brain regions are also observed in PD [4–6]. Age and pesticide/herbicide exposure are the most important disease risk factors [7–9]. Importantly, there is no clinical therapy available that has been shown to slow or reverse PD. While the majority of PD is diagnosed as idiopathic, 5–10% of cases are attributable to familial forms of PD [10]. Although genetic PD represents only a small percentage of patients, mutations in these genes point to underlying biochemical pathways that could also be relevant to sporadic PD patients. Three missense mutations in the small pre-synaptic protein a- Synuclein (SNCA/PARK1/4; GenBank ID 6622) have been shown to result in autosomal-dominant PD. A critical effect of protein dose on pathology is implicated by disease-causing gene duplication and triplication [11–14]. a-Synuclein is also a major protein component of the Lewy Bodies (LB), the key histologic feature of dopaminergic and non-dopaminergic neurons found in PD patients [15]. Thus, a-Synuclein is strongly suggested to be a causal factor in PD pathogenesis. Human a-Synuclein mutation or overexpression results in cytotoxicity, with [A53T]a-Synuclein being the most toxic variant known. Direct cell loss can be induced in both in vitro and in vivo models of yeast, C. elegans, Drosophila, rat, mouse, and non-human primate [16–23]. The formation of a-Synuclein oligomers from PLoS Genetics | www.plosgenetics.org 1 February 2012 | Volume 8 | Issue 2 | e1002488