Multiple System Atrophy and Parkinson ’ s disease

The synucleinopathies are a diverse group of neurodegenerative disorders that share a common pathologic intracellular lesion, composed primarily of aggregates of insoluble α-Synuclein (α-Syn) protein in selectively vulnerable populations of neurons and glia. The term, “synucleinopathy,” was introduced in 1998 after it was recognized that filamentous α-Syn deposits might represent a link between Multiple system atrophy (MSA), Parkinson’s disease (PD), and Dementia with Lewy bodies (DLB). In this thesis I have focused on two of the synucleinopathies, PD and MSA. -Syn is an abundant neuronal protein enriched in presynaptic nerve terminals, and it has been critically implicated in the genetics and neuropathology of synucleinopathies. α-Syn is a soluble protein, yet, under certain conditions it gradually oligomerizes and deposits in intra-cellular insoluble inclusions. These inclusions, when in neuronal cells, are called Lewy bodies. When in oligodendrocytes, the inclusions are called glial cytoplasmic inclusions (GCIs), and are characteristics of MSA histopatholgy. Specific point mutations in the α-Syn gene, as well as duplications and triplications at the locus for wild-type -Syn, increase its propensity to self-oligomerize, in addition to increasing its pathogenicity. In recent years, extracellular -Syn became the focus of many studies due to its potential role in disease initiation and progression While secretion of -Syn may occur normally from healthy neurons, growing evidence indicates that enhanced secretion is associated with -Syn pathogenesis. The mechanisms involved in -Syn secretion are not yet fully understood. The general aims of the thesis are: 1. To elucidate the mechanisms underlying the pathogenic accumulation of -Syn, a neuronal protein in oligodendroglia, as observed in Multiple System Atrophy. 2. To elucidate the mechanisms by which -Syn expression affects catalase, the antioxidative stress enzyme. In the first part of the study, I tested the hypothesis that oligodendrocytes can take up neuronal-secreted -Syn as part of the pathogenic mechanisms leading to MSA. I reported that increases in the degree of soluble α-Syn oligomers and intracellular Syn levels, enhance its secretion from cultured MN9D dopaminergic cells, stably expressing the protein. In accord, I showed that the oligodendroglial cell line, OliNeu, can take-up neuronal-secreted or exogenously-added -Syn from their conditioning medium. This uptake is concentration-, time-, and clathrin-dependent. Utilizing the demonstrated effect of polyunsaturated fatty acids (PUFAs) to enhance α-Syn neuropathology, I showed in vivo that increases in neuronal -Syn levels enhance its localization to oligodendrocytes in the brains of a mouse model for synucleinopathies, transgenic for the human A53T -Syn mutant form. Thus, the pathogenic mechanisms leading to elevated levels of α-Syn in neurons underlie neuronal secretion, and the subsequent uptake of -Syn by oligodendrocytes in MSA. In the second part of the study I aimed at investigating the mechanisms by which Syn expression affects catalase, the anti-oxidative stress enzyme. Previous studies in our lab showed evidence suggesting that -Syn specifically affects catalase expression levels and enzymatic activity. Searching for a mechanism through which -Syn expression affects catalase, I found that peroxisome proliferator-activated receptor (PPAR which is known to control catalase transcription, has inhibited transcription activity when -Syn is expressed. It is important to note that activating PPARα, PPARγ and RXR, but not PPARβ/δ, restored and enhanced catalase activity. Based on these results, I have suggested that through its involvement in fatty acid metabolism, -Syn expression affects the pool of fatty acid metabolites, which normally act as activating ligands for PPAR. The affected PPAR activity inhibits catalase expression and results in the accumulation of oxidative damage that characterizes PD. I concluded that α-Syn down-regulates catalase expression and activity by interfering with the complex and overlapping network of nuclear receptor dimerization and transcription activation. In summary, my results may contribute to the understanding of the pathogenic mechanisms involved in -Syn toxicity in the synucleinopathies. A role for brain lipids, and specifically for PUFAs, has been demonstrated. List of abbreviation: 1-methyl-4-phenylpyridinium ion (MPP + ). 2,4-dinitrophenylhydrazine (DNPH) 3′-Cyclic Nucleotide 3′-Phosphodiesterase (CNPase) 3-nitropropionic acid (3-NP) 4-hydroxynonenal (4-HNE) 6-hydroxydopamine (6-OHDA) carbonic anhydrase II (CAII) cerebrospinal fluid (CSF) clathrin-mediated endocytosis (CME) conditioned medium (CM) deep brain stimulation (DBS) Dementia with Lewy bodies (DLB) docosahexaenoic acid (DHA, 22:6) docosohexaenoic acid (DHA) eicosapentaenoic acid (EPA, 20:5) genome-wide association (GWA) glial cytoplasmic inclusions (GCI) Glial fibrillary acidic protein (GFAP) glial nuclear inclusions (GNIs) glutathione (GSH) hydrogen peroxide (H2O2) hydroxyeicosatetraenoic acid (HETE) Immunocytochemistry (ICC) Immunohistochemistry (IHC levodopa (L-dopa) Lewy bodies (LB), Lewy neurites (LN) linolenic acid (ALA, 18:3) Multiple System Atrophy (MSA) Muno unsaturated fatty acid (MUFA) myelin basic protein (MBP) neuronal cytoplasmic inclusions (NCIs) neuronal nuclear inclusions (NNIs) Neuronal Nuclei (NeuN) nuclear receptor related 1 (NURR1) Olivopontocerebellar Atrophy (OPCA) paraformaldehyde (PFA) Parkinson’s disease (PD) peroxisome proliferatoractivated receptor  coactivator  (PGC-1-) peroxisome proliferator-activated receptor gamma(PPAR) polyunsaturated fatty acids (PUFAs) prostaglandin J2 (PGJ2) proteolipid protein (PLP) quinolinic acid (QA) reactive oxygen species (ROS) Retinoid X Receptor (RXR) Reverse transcription polymerase chain reaction (RT-PCR) saturated fatty acids (SFAs) Shy-Drager Syndrome (SDS) single nucleotide polymorphisms (SNPs) small hairpin RNA (shRNA) Striatonigral Degeneration (SND) substania nigra pars compacta (SNpc) succinate dehydrogenase inhibitor superoxide dismutase (SOD) superoxide dismutase (SOD1) transgenic (tg) tubulin polymerization promoting protein (TPPP/P25) wild type (wt) α-Synuclein (α-Syn) List of publication: 1. The clathrin-dependent localization of dopamine transporter to surface membranes is affected by α-Synuclein" Haya Kisos, Tziona Ben-Gedalya and Ronit Sharon. Submitted. 2. -Synuclein expression inhibits catalase activity in brains of mice modeling Parkinson's disease. Eugenia Yakunin, Haya Kisos, Willem Kulik,; Ronald J. A Wanders,and Ronit Sharon. Submitted. 3. Enhanced neuronal -Synuclein pathology associates with its accumulation in oligodendrocytes in mice modeling Synucleinopathies. Haya Kisos, Katharina Pukaß, Tamir Ben-Hur, Christiane Richter-Landsberg and Ronit Sharon. PLoS One,2012 4. Docosahexaenoic acid controls α-synuclein neuropathology in a mouse model. Eugenia Yakunin ,Virginie Loeb ,Haya Kisos ,Yoav Biala ,Shlomo Yehuda, Yoel Yaari, Dennis J. Selkoe and Ronit Sharon. Brain Pathol. 2012 5. Alpha-synuclein expression selectively affects tumorigenesis in mice modeling Parkinson's disease. αSynuclein Expression Selectively Affects Tumorigenesis in Mice Modeling Parkinson's Disease Eitan Israeli,Eugenia Yakunin,Yonaton Zarbiv,Amir HacohenSolovich,Haya Kisos,Virginie Loeb,Michal Lichtenstein,Tziona BenGedalya,Ofra Sabag,Eli Pikarsky,Haya Lorberboum-Galski,Ronit Sharon PLoS One. 201