-synuclein in parkinson’s disease - cold spring...

a-Synuclein in Parkinson’s Disease

Leonidas Stefanis

Laboratory of Neurodegenerative Diseases, Biomedical Research Foundation of the Academyof Athens, and Second Department of Neurology, University of Athens Medical School,Athens 11527, Greece

Correspondence: [email protected]

a-Synuclein is a presynaptic neuronal protein that is linked genetically and neuropatholog-ically to Parkinson’s disease (PD). a-Synuclein may contribute to PD pathogenesis in anumber of ways, but it is generally thought that its aberrant soluble oligomeric confor-mations, termed protofibrils, are the toxic species that mediate disruption of cellular homeo-stasis and neuronal death, through effects on various intracellular targets, including synapticfunction. Furthermore, secreted a-synuclein may exert deleterious effects on neighboringcells, including seeding of aggregation, thus possibly contributing to disease propagation.Although the extent to which a-synuclein is involved in all cases of PD is not clear, targetingthe toxic functions conferred by this protein when it is dysregulated may lead to novel thera-peutic strategies not only in PD, but also in other neurodegenerative conditions, termed syn-ucleinopathies.

The first link of Parkinson’s disease (PD) to a-synuclein was also the first conclusive demon-

stration of a genetic defect leading to PD, and thushas historical and conceptual value. Althoughfamilial contribution to the disease was discussedby some, it was felt to be minor at best by most.The tangible discovery of a gene defect linked toPD in particular families was a ground-breakingdiscovery,as itopenedthedoortoafloodofstudiesinvestigating the genetic base of the disorder, cul-minating in more recent genome-wide associationstudies (GWAS), that, remarkably, have made afullcircle back to the origins of the molecular geneticera of PD; it turns out that one of the major genesidentified as linked to sporadic PD is none otherthan SNCA, the gene encoding fora-synuclein.

A parallel story that has developed over theyears is the abundant abnormal a-synucleinpathology that characterizes neuropathologicalspecimens derived not only from PD patients,but also patients with other neurodegenerativeconditions, collectively termed “synucleinopa-thies.” The ability to model abnormal a-synu-clein accumulation in various cellular andanimal models has led to the study of the conse-quences of such accumulation, to the decipher-ing of the molecular pathways involved, and tothe identification of potential therapeutic tar-gets. Such targets may prove of general utility,and provide the basis for future therapeuticsin most forms of PD, and, potentially, other syn-ucleinopathies.

Editor: Serge Przedborski

Additional Perspectives on Parkinson’s Disease available at www.perspectivesinmedicine.org

Copyright # 2012 Cold Spring Harbor Laboratory Press; all rights reserved; doi: 10.1101/cshperspect.a009399

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GENETICS OF PD AND OTHERDISORDERS WITH LINKS TOSYNUCLEINOPATHIES

In 1997, Polymeropoulos et al. reported thefirst specific genetic aberration linked to PDin a large Italian kindred, and in some Greekfamilial cases. The mutation corresponded to aG209A substitution in the SNCA gene encodingfor a-synuclein, resulting in an A53T aminoacid change. The pattern of inheritance was au-tosomal-dominant, and the disease was early-onset, usually in the 40s. Although there wassome initial skepticism, based on the fact thatthe threonine was the amino acid in position53 in the rodent SNCA homolog, this was setaside following the identification of two morepoint mutations in SNCA, leading to A30Pand E46K amino acid changes, which were as-sociated with autosomal-dominant PD (Krugeret al. 1998; Zarranz et al. 2004). The next land-mark study that linked SNCA to PD through adifferent mechanism identified triplications ofthe SNCA locus in separate families with an au-tosomal-dominant inheritance pattern (Single-ton et al. 2003). This led to a potential totalof a duplication of the a-synuclein load insuch patients, and this was confirmed laterat the protein level (Miller et al. 2004). Otherfamilies were subsequently described that har-bored duplications of the gene, which wassimilarly associated with autosomal-dominantdisease (Chartier-Harlin et al. 2004). Interest-ingly, a gene dosage effect exists, in that triplica-tion cases are earlieronset and have a more severecourse compared to the duplication cases (Fuchset al. 2007).

When a specific genetic defect is identified inrare familial cases, it is almost a reflex reaction tolook at the same gene in the sporadic disease,with the hope of identifying polymorphismsthat may be linked to the disease in associationstudies. This has not been particularly successfulin other diseases, but in the case of PD and theSNCA gene this approach has borne fruit. Initialstudies focused on the Rep1 polymorphic regionlocated approximately 10 kB upstream of theSNCA transcriptional initiation site. Particularpolymorphisms in this region conferred a rela-

tive risk for sporadic PD in initial associationstudies, which were replicated and then con-firmed in a large metanalysis (Maraganoreet al. 2006). Others studies found associationsof PD with 30 regions of the gene (Mueller et al.2005; Mizuta et al. 2006). Targeted associationstudies are, however, prone to false positives;therefore, it is very important that the link ofSNCA to sporadic PD has been confirmedthrough large unbiased GWAS. SNCA, and, inparticular, the 30 region, has been a consistenthit in such studies, and represents, of the linksidentified, probably the most robust one (Nallset al. 2011).

The conclusion from these genetic studiesis that SNCA is linked to both rare familialand sporadic PD from a genetic standpoint. Itis notable that the association of SNCAvariantsto disease has now also been extended to multi-ple-system atrophy (MSA), another prototypi-cal synucleinopathy, which is invariably spo-radic (Al-Chalabi et al. 2009; Scholz et al. 2009).

THE INVOLVEMENT OF a-SYNUCLEIN INTHE NEUROPATHOLOGICAL PICTURE OFPD AND RELATED DISORDERS

The genetic link of SNCA to PD in 1997 ledinvestigators to rapidly develop appropriate an-tibodies against a-synuclein and use them inhistopathological sections of PD patients. a-Synuclein turned out to be robustly expressedwithin Lewy bodies (LBs), and, in particular,in the “halo” of the inclusions (Spillantini et al.1997, 1998; Baba et al. 1998). In fact,a-synucleinimmunohistochemistry is currently the goldstandard in the neuropathological evaluationof PD. With repeated studies, three major in-sights emerged: (1) a-synuclein pathology inPD is not confined to the cell soma, but is alsoprominent in neuritic processes, (2) it is wide-spread in various brain regions in PD, and (3)it is present in a number of other synucleinopa-thies, such as MSA, dementia with Lewy bodies(DLBs), many cases of Alzheimer’s disease (AD)(the so-called LB variant of AD), neurodegen-eration with brain iron accumulation (NBIA)type I, pure autonomic failure (PAF), and evena subtype of essential tremor (reviewed in Kahle

L. Stefanis

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2008). The distribution of the pathology at thecellular and regional level is different in eachdisease.

Braak et al. (2003), in their seminal study ofthe staging of PD pathology, used a-synucleinimmunohistochemistry in a large number ofautopsy cases. Based essentially on a-synucleinneuritic pathology (“Lewy neurites”), they sug-gested a six-stage scheme in which the pathol-ogy begins first at the olfactory bulb and thedorsal vagal nucleus and gradually follows anascending course, culminating in widespreada-synuclein pathology at the later stages, in-volving associative cortical regions. Notably,substantia nigra is only affected in stage 3 ofthis scheme. Although there have been crit-icisms against this hypothesis (Burke et al.2008), and it does not explain the absence ofclinical symptoms in subjects who on autopsyhave widespread a-synuclein pathology, or theclinical picture of DLB, it appears to hold upfor the majority, but not all, of cases examinedin large cohorts of LB cases (Dickson et al.2010). What is important to note here is thatoverall in the PD brain the majority of ab-normally deposited a-synuclein is in neuriticprocesses. The presence of a-synuclein in LBsis likely the tip of the iceberg. In fact, Kramerand Shulz-Schaeffer (2007), using a modifiedprotocol termed protein aggregate filtration,showed in DLB cases a remarkable degree ofaberrant neuritic a-synuclein pathology, in thevirtual absence of LBs.

Overall, it is clear that abnormal a-synu-clein deposition occurs early in PD. Giventhe fact that such pathology may develop sec-ondarily, for example, owing to iron mishan-dling in NBIA type I, we cannot be certainthat it is a primary factor in the disease process.However, the generally quite defined pattern ofa-synuclein pathology in PD, the cellular andanimal models of a-synuclein overexpressionthat will be discussed, and, most importantly,the combination with the genetic data men-tioned above, suggest that this aberrant deposi-tion is the driving force in PD pathogenesis.Such deposition, as mentioned, may be subtleand difficult to detect in the absence of elabo-rate immunohistochemical techniques that, it

is hoped, will become more widely applicableand available.

THE STRUCTURE AND FUNCTION OFa-SYNUCLEIN

The SNCA gene encodes for a 140 amino acidprotein, which in aqueous solutions does nothave a defined structure, hence the term “na-tively unfolded protein.” a-Synuclein, however,does form a-helical structures on binding tonegatively charged lipids, such as phospholipidspresent on cellular membranes, and b-sheet-rich structures on prolonged periods of incu-bation. The protein is composed of three dis-tinct regions: (1) an amino terminus (residues1–60), containing apolipoprotein lipid-bind-ing motifs, which are predicted to form amphi-philic helices conferring the propensity to forma-helical structures on membrane binding, (2)a central hydrophobic region (61–95), so-calledNAC (non-Ab component), which confers theb-sheet potential, and (3) a carboxyl terminusthat is highly negatively charged, and is proneto be unstructured (Fig. 1). There are at leasttwo shorter alternatively spliced variants of theSNCA gene transcript, but their physiologicaland pathological roles have not been well char-acterized (Maroteaux and Scheller 1991; Uedaet al. 1993; Maroteaux et al. 1988).

a-Synuclein is a member of the synucleinfamily of proteins, which also include b- andg-synuclein. What largely sets aparta-synucleinfrom the other members structurally is the NACregion. All three members of the family arepredominantly neuronal proteins that underphysiological conditions localize preferentiallyto presynaptic terminals (George 2002). Thereare some reports of extranigral b- and g-synu-clein neuritic pathology in synucleinopathies,but, if it exists, it does not appear to be wide-spread (Galvin et al. 1999). Interestingly, pointmutations in b-synuclein have been describedin rare cases with DLB (Ohtake et al. 2004),and expression of one of these in a transgenicmouse model-induced neurodegeneration (Fu-jita et al. 2010). On the other hand, wild-type(WT) b-synuclein has been found to be pro-tective in various settings against a-synuclein-


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mediated neurodegeneration (Hashimoto et al.2001; Park and Lansbury, 2003). It would ap-pear therefore that WT b-synuclein may haveneuroprotective properties, but that, under cer-tain situations, it may assume a neurotoxic po-tential.

a-Synuclein is abundantly expressed in thenervous system, comprising 1% of total cy-tosolic protein. On immunohistochemistry innormal brains a-synuclein antibodies confera punctuate neuropil staining pattern, consis-tent with presynaptic terminal staining, whereasneuronal soma staining is less apparent (Kahle2008). In presynaptic terminals a-synuclein ispresent in close proximity, but not within, syn-aptic vesicles. In fact a-synuclein is also veryabundant, for unclear reasons, in erythrocytesand platelets (Barbour et al. 2008). Whether itis expressed at all under physiological condi-tions in glia is somewhat controversial; it wouldappear that, if it exists, such expression is pre-sent at very low levels (Mori et al. 2002a). Theexpression of a-synuclein is induced duringneuronal development, following determina-tion of neuronal phenotype and establishmentof synaptic connections, and lags behind theinduction of proteins involved in synapticstructure (Withers et al. 1997; Kholodilov et al.1999; Murphy et al. 2000; Rideout et al. 2003).Moreover, a-synuclein expression levels are

modulated in conditions that alter plasticity orconfer injury (George et al. 1995; Kholodilovet al. 1999; Vila et al. 2000). Such data have ledto the notion that a-synuclein may be a modu-lator of synaptic transmission. Examinations ofneurophysiological properties of mice null fora-synuclein and of cellular systems in whichWT a-synuclein is overexpressed have been in-strumental in this regard, as they reveal alter-ations in neurotransmitter release, which arenot always consistent across neuronal systems(Abeliovich et al. 2000; Murphy et al. 2000;Cabin et al., 2002; Larsen et al. 2006). Overallthough, it would appear that a-synuclein actsas a subtle break for neurotransmitter releaseunder circumstances of repeated firing. An ele-gant study by Fortin et al. (2005) showed thatfluorescently labeled a-synuclein moves awayfrom the vesicles on neuronal firing, and thengradually returns. These effects may involvetransient binding of a-synuclein to the vesicles,whereas a-synuclein may also be involved in ve-sicular biogenesis, through effects on phospha-tidic acid metabolism, and in compartmenta-lization between resting and readily releasablepools (Murphy et al. 2000). Consistent with arole fora-synuclein at the level of synaptic trans-mission, a-synuclein modulates profoundly thephenotype of mice null for the presynaptic pro-tein CSP-a (cysteine string protein-a), which

9561

Repeat 6Repeat 5

Apolipoprotein motif

Repeat 2 Repeat 4Repeat 3 Acidic tail140

Repeat 1

1N C

A53T

NAC region necessary for aggregation

E46KA30P

Mutations geneticallylinked to familial

autosomal-dominantParkinson’s disease

Figure 1. Schemtic structure of a-synuclein.

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show synaptic degeneration (Chandra et al.2005). It is believed that a-synuclein, througha chaperone-like function, may act in synergywith CSP-a in the assembly of the SNARE com-plex (Chandra et al. 2005), and this effect mayinvolve, in particular, binding to synaptotag-min-2 (Burree et al. 2010).

Therefore, the main function ofa-synucleinwould appear to be the control of neurotrans-mitter release, through effects on the SNAREcomplex. Recent evidence, to be discussed be-low, suggests that this physiological functionmay provide insight into the aberrant functionunleashed in disease states.

SYNUCLEINOPATHY MODELS

Based on the genetic and pathological data, at-tempts have been made to create cellular andanimal models in which various forms of a-synuclein are overexpressed. We have electedto not present such models with any great detailhere, but rather highlight the inferences gainedfrom such models regarding the potentialpathological (and sometimes physiological) ef-fects of a-synuclein. Suffice it to say here, thatthere are a number of transgenic mice available,in which expression of various forms of a-syn-uclein is achieved through different promoters,allowing in some cases regional and temporalcontrol of expression. In such mice, there is gen-erally some degree of neurodegneration and LB-like pathology, but it does not involve primarilythe dopaminergic system. Some mice developbehavioral phenotypes that could be akin topresymptomatic PD (reviewed by Chesselet2008). An alternative approach has been theviral-mediated transduction of dopaminergicneurons of the substantia nigra of rodentswith a-synuclein, and this has been more suc-cessful in modeling the dopaminergic cell deathaspect of the disease (Kirik et al. 2002). A num-ber of invertebrate models, including fly (Feanyand Bender 2000) and Caenorhabditis elegans(Lasko et al. 2003), have also been created, andhave been very useful in testing theories regard-ing the toxic pathways elicited by aberrant a-synuclein.

THE AGGREGATION POTENTIAL OFa-SYNUCLEIN

The ability of a-synuclein to generate b-sheetstructures under certain circumstances gener-ated a lot of enthusiasm, as it provided parallelsto the b-sheet structure of b-amyloid, and con-sequently, a unified pathogenetic basis for thetwo most common neurodegenerative diseases.Indeed, WTas well as disease-related mutants ofa-synuclein form amyloid-like fibrils on pro-longed incubation in solution (Conway et al.2000). It is widely agreed on that such fibrilsform the basis of the mature LBs and Lewy neu-rites (LNs) present in synucleinopathies. In theprocess of fibril formation various intermediateforms ofa-synuclein develop. These are initiallysoluble oligomeric forms ofa-synuclein that as-sume spherical, ring-, and stringlike characteris-tics when seen under the electron microscope.Such structures, collectively termed protofibrils,gradually become insoluble and coalesce into fi-brils. It is difficult to capture these intermediatestructures in cells or in vivo, but their biochemi-cal equivalent isthought to besolubleoligomericforms, some, but not all, of which are SDS-stableand can be detected on SDS/polyacrylamidegel electrophoresis (PAGE) gels. Other forms,however, can only be captured on native gelsor by size-exclusion chromatography (Emma-nouilidou et al. 2010a). This cascade of events,starting from the natively unfolded proteinand culminating in the mature fibril formationis collectively termed a-synuclein aggregation.

The general consensus in the field is thataggregation is a main pathogenic feature of a-synuclein. This is buttressed by the lack of tox-icity conferred by WT b-synuclein or by a-syn-uclein variants that do not contain the NAC do-main and do not form aggregates (Periquet et al.2007), by the beneficial effects of overxepressingHSPs, which assist in refolding of aggregation-prone proteins, in synucleinopathy models(Auluck and Bonini 2002; Auluck et al. 2002;Klucken et al. 2004), and by the protection con-ferred in cellular models of a-synuclein overex-pression by reagents that inhibit or neutralizeaggregated species (Vekrellis et al. 2009; Bieschkeet al. 2010). However, the protective effects of



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HSP overexpression are not always apparent(Shimshek et al. 2010). There is less agreementas to which particular species are neurotoxic.A predominant idea is that some of the earlysoluble oligomeric species have such a potential.This is based on the following pieces of data:

(1) A53T and A30P a-synuclein form moreprotofibrils than the WT protein, whereasonly A53T forms more readily mature fi-brils; however, E46K a-synuclein formsless protofibrils than the WT, thereforethere is no simple correlation that can ex-plain everything using this scheme, whichis based on in vitro cell-free systems (Con-way et al. 2000; Fredenburg et al. 2007).

(2) Dopamine and its metabolites act as inhib-itors of the conversion of protofibrils tomature fibrils in vitro and in vivo, presum-ably through the formation of dopamine/a-synuclein adducts (Conway et al. 2001;Mazzulli et al. 2006; Tsika et al. 2010), sug-gesting that the preferential vulnerability ofdopaminergic neurons to the pathogenicprocess of PD may be attributable to theenhanced presence of soluble oligomers(although, surprisingly, mouse nigral neu-rons, which contain more oligomers, ap-pear relatively resistant to transgenic a-synuclein-mediated neurodegeneration).

(3) In cellular systems and in vivo, the presenceof soluble oligomers, and not frank inclu-sions, generally correlates best with neu-rotoxicity (Gosavi et al. 2002; Lo Biancoet al. 2002). Two major studies providedsupport to this idea, as they showed en-hanced neurotoxicity both in cell cultureand in vivo with artificial mutants of a-synuclein that favored the formation of sol-uble oligomers over mature fibrils (Karpinaret al. 2009; Winner et al. 2011). It shouldbe noted, however, that the issue is notsettled; the protofibrils are heterogeneous,“off pathway” oligomers that do not leadto toxicity may exist, and frank inclusionshave the potential of being neurotoxic them-selvesover time,especially ifpresent inaxons,where they can block neuronal trafficking

or “suck in” essential neuronal components.On the other hand, if the protofibril theoryis correct, enhancing inclusions could beviewed as a method to remove the toxic inter-mediate oligomeric species, and this has beenattempted with some success in cellularmodels (Bodner et al. 2006).

a-SYNUCLEIN POSTTRANSLATIONALMODIFICATIONS AND INTERACTIONS

a-Synuclein may be modified by phosphoryla-tion, oxidation, nitrosylation, glycation, or gly-cosylation. Of all such modifications, the beststudied is phosphorylation. Antibodies specificto S129-phosphorylated a-synuclein promi-nently decorate LBs in synucleinopathies (Fuji-wara et al. 2002). Although the initial idea,which appeared to be buttressed by experimentsin transgenic flies, was that such phosphoryla-tion enhanced a-synuclein-soluble oligomers(but actually decreased a-synuclein inclusions)and neurotoxicity (Chen et al. 2005, 2009);subsequent studies have cast doubt on this no-tion, providing contradictory results (Gorba-tyuk et al. 2008; Paleologou et al. 2008; Azeredoda Silveira et al. 2009). In fact, it may be thatS129 phosphorylation, which occurs minimallyunder physiological circumstances, occurs afterthe fact in already formed LBs (Paleologou et al.2008). Enhancing a-synuclein phosphatase ac-tivity was, however, protective against a-synu-clein-mediated neurotoxicity, suggesting a causaldetrimental role of a-synuclein phosphorylationin the disease process (Lee et al. 2011). Polo-likekinases appear to be the physiological kinasesfor a-synuclein S129 phosphorylation (Ingliset al. 2009; Mbefo et al. 2010). S87 is anotherphosphorylation site for a-synuclein, and S87-Pa-synuclein is also apparent within LBs, but itspathological significance is unclear (Paleologouet al. 2010). Phosphorylation on a-synuclein ty-rosine residues has also been reported and mayact in opposing fashion to S129 phosphorylation,at least in Drosophila (Chen et al. 2009).

A number of studies in cell-free systemsand in cell culture have shown that a-synucleincan be oxidized, and this seems to drive its oli-gomerization (Lee et al. 2002; Betarbet et al.

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2006; Mirzaei et al. 2006; Qin et al. 2007). Ironand dopamine appear to exert a similar effect,although whether this is a direct interaction ormediated by oxidation is less clear (Ostrerova-Golts et al. 2000; Conway et al. 2001; Kostkaet al. 2008; Leong et al. 2009). The potent oxi-dant 4-hydroxy-2-nonenal (HNE) promotedstable soluble oligomers, inhibiting their con-version to fibrils; such stable soluble oligomerswere highly toxic, favoring again the solubleoligomer toxicity theory (Qin et al. 2007). In asimilar vein, nitration of a-synuclein has beenreported; in fact, an antibody specifically gener-ated against nitrosylated a-synucelin decoratesLB-like pathology (Giasson et al. 2000). Mono-meric and dimeric nitrated a-synuclein pro-moted fibril formation (Hodara et al. 2004),whereas oligomeric nitrated a-synuclein actu-ally inhibited it, stabilizing as soluble oligomers(Uversky et al. 2005), suggesting complex effectsof nitrosylation on a-synuclein aggregation. Aswith S129 phosphorylation, it is possible thatthe intense nitrosylation of a-synuclein withinLB-like structures occurs after the fact.

Given the fact that oxidative/nitrative stressinduced by environmental factors, includingmitochondrial toxins, has been heavily impli-cated in PD, oxidation- or nitration-induceda-synuclein oligomerization has attracted at-tention, as a point of potential convergence be-tween environmental and genetic factors lead-ing to the disease. It should be noted, however,that a-synuclein aggregation could be dissoci-ated from mitochondrial toxicity-induced death(Liu et al. 2008), arguing against simplistic lin-ear pathways to neurodegeneration, and raisingthe issue whether in certain instances the ab-undant a-synuclein-related pathology may bean epiphenomenon, and not causally related toneuronal dysfunction and death. Similar infer-ences can be taken from a study in proteasomalinhibitor-treated neurons, where the b-sheet-like pathology of a-synuclein was dissociatedfrom death (Rideout et al. 2004).

Another modification that may be impor-tant is carboxy-terminal truncation of a-synu-clein, leading to fragments lacking part or thewhole of the acidic tail (Li et al. 2005). Thesefragments appear to be more prone to fibrilliza-

tion (Kim et al. 2002; Murrayet al. 2003). Calpain,one of the enzymes thought to perform suchprocessing, is especially interesting given its cal-cium dependence and localization at presynapticterminals (Mishizen-Eberz et al. 2003, 2005).

a-Synuclein is quite promiscuous in itsbinding properties, and may bind various pro-teins in neuronal cells, including componentsof dopamine metabolism, such as tyrosine hy-droxylase (TH) and the dopamine transporter(DAT), modulating their function (Ostrerovaet al. 1999; Perez et al. 2002; Wersinger andSidhu 2003; Wersinger et al. 2003; Peng et al.2005). There are quite a few reports suggestingthat through such binding to various proteins,a-synuclein may exert pro- or antiapoptoticeffects (Ostrerova et al. 1999; da Costa et al.2000; Jin et al. 2011), although no effect on pro-grammed neuronal cell death has been shown inmice null for a-synuclein (Stefanis et al. 2004).Interestingly, however, these mice are resistantto the well-known mitochondrial neurotoxinMPTP, through mechanisms that are still notdeciphered, but may involve steps both up-stream of and downstream from mitochon-drial dysfunction (Dauer et al. 2002). Such miceare also resistant against neuroinflammation-induced nigral degeneration conferred by lipo-polysaccharide (LPS) (Gao et al. 2008), suggest-ing that endogenous a-synuclein is somehowinvolved in such death pathways.

A number of interactions of a-synuclein withother proteins have been implicated in its aggrega-tion potential; most notable amongst these is syn-philin-1, a protein identified through a yeast-two-hybrid screen as an AS-interacting protein,which promotes a-synuclein aggregation andinclusion formation (Engelenderet al. 1999). In-terestingly, transgenic expression of synphilin-1on the background of A53T a-synuclein trans-genic mice attenuates neurodegeneration, as as-sessed by indices of cell death and gliosis, whileenhancing inclusion formation, suggesting thatpromoting a-synuclein inclusion formation isneuroprotective (Smith et al. 2010).

Interactions of a-synuclein with lipids maybe important not only for their physiologicfunctions, but also for their aberrant effects. Re-garding the former, it is interesting to note that



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sequestration of the fatty acid arachidonic acidaway from the SNARE complex by a-synucleinhas been suggested to underlie its inhibitoryeffect on neuronal transmission (Darios et al.2010), and that alterations in levels of key en-zymes in lipid signaling, such as PLD2 andPLA2, may underlie the phenotype in SNCAnull microglia, which are overreactive but lackphagocytic potential (Austin et al. 2006, 2011).Regarding the latter, polyunsaturated fatty acids(PUFAs) have been shown to enhance oligome-rization and neurotoxicity of a-synuclein (Per-rin et al. 2001; Sharon et al. 2003; Assayag et al.2007).

GAIN-OF-FUNCTION ANDACCUMULATION OF a-SYNUCLEIN

The notion that enhanceda-synuclein levels arecausative in PD pathogenesis is derived fromthe familial SNCA multiplication cases show-ing a dose-dependent correlation ofa-synucleinload to the PD phenotype, the autosomal-dom-inant pattern of inheritance for the point muta-tions, the accumulation of a-synuclein in synu-cleinopathy brains, and the demonstration thatfeatures of the disease can be mimicked by WTa-synuclein overexpression in various models.Consistent with this idea, a-synuclein proteinlevels are increased with aging in the substantianigra, and correlate with decreased TH immu-nostaining (Chu and Kordower 2007). How-ever, it should be noted that there is insufficientevidence that there is a generalized excess ofa-synuclein in PD brains. In fact, mRNA stud-ies have been quite contradictory in this regard,with some even showing a decrease of SNCA ex-pression in PD nigra (Dachsel et al. 2007). Pro-tein levels are not obviously increased overall inPD brains, although clearly there is induction ofinsoluble components, including monomericand oligomeric species. A comprehensive studyof multiple brain areas showed that membrane-associated monomeric a-synuclein levels wereonly modestly increased in the substantia nigra,and not in other brain regions, of PD patientscompared to controls; even so, the levels insome PD cases were within the spectrum of con-trol values. In contrast, in MSA, the membrane-

associated a-synuclein increase was robust invulnerable brain regions (Tong et al. 2010). Itmay of course be that neurons with the highestexpression levels ofa-synuclein are most vulner-able and succumb early in the disease process,giving their place to glia, thus confounding theresults. To partly circumvent this issue, Grunde-mann et al. (2008) performed laser-capture mi-crodissection studies, and reported a consider-able increase of SNCA expression in survivingnigral neurons derived from PD brains com-pared with controls. However, this increase didnot appear to be specific for SNCA (Grunde-mann et al. 2008). Therefore, the issue is stillopen.

An indirect way of approaching this is byassessing whether polymorphic SNCA variantsthat confer disease risk in association studiesalter a-synuclein levels in model systems. Ininitial studies, the Rep1 polymorphism associ-ated with the disease led to increased SNCAtranscriptional activity in neuronal cell lines(Chiba-Falek and Nussbaum 2001); this findinghas been confirmed in mice (Cronin et al.2009), and even in the human brain (Linnertzet al. 2009), although in another study an asso-ciation between Rep1 polymorphisms and a-synuclein mRNA and protein levels in the brainwas not apparent (Fuchs et al. 2008). Polymor-phisms within the 30 region may also correlatewith AS levels (Fuchs et al. 2008; Mata et al.2010). Intriguingly, it was reported that risk-conferring alleles at the 30 untranslated region(UTR) were associated with increased expres-sion of an alternatively spliced form of SNCA(McCarthy et al. 2011), suggesting another me-chanism through which gene variations may belinked to the disease.

It is generally assumed that a-synuclein leadsto disease through a toxic gain-of-function thatis likely inherent in the normal protein when it ex-ceeds a certain level. In apparent agreement withthis, a-synuclein null mice, in contrast to trans-genic overexpressors, display no overt neuropa-thological or behavioral phenotype (Abeliovichet al. 2000; Cabin et al. 2002). However, whenstudied more closely, such mice do have somedeficits in the dopaminergic system, in par-ticular, a slightly reduced number of postnatal

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dopaminergic neurons (Robertson et al. 2004),and a drop of dopaminergic terminals with ag-ing (Al-Wandi et al. 2010). Thus, the theory thatpart of the PD phenotype might be attributableto the sequestering of physiological a-synucleinin inclusions, which could cause neuronal, and,in particular (in view of the normal function ofthe protein), synaptic damage, cannot be dis-carded.

Despite the fact that transcriptional regu-lation of a-synuclein is so important for PDpathogenesis, little is known about the mecha-nisms involved. Transcription factors GATA-1(Scherzer et al. 2008) and ZSCAN21 (Cloughand Stefanis 2007; Clough et al. 2009) withinIntron 1 have been proposed to play a role asinducers of transcription. In the latter case, asignaling pathway involving ERK/PI3 kinasesignaling mediated ZSCAN-induced SNCAtranscriptional activation (Clough and Stefanis2007; Clough et al. 2009). A similar ERK-de-pendent signaling pathway regulating SNCAproduction was apparent in enteric neurons(Paillusson et al. 2010). Methylation may beimportant in SNCA expression, as methylationwithin Intron 1 suppressed SNCA transcrip-tional activity, and brains of patients with LBdiseases showed decreased methylation withinthis region, suggesting epigenetic regulation ofSNCA expression (Jowaed et al. 2010; Matsu-moto et al. 2010). In one study, it was proposedthat sequestration of the methylation factorDnmt1 away from the nucleus by a-synucleinmay lead to reduced methylation and enhancedSNCA exression, thus creating a feed-forwardloop of SNCA overexpression (Desplats et al.2011). Posttranscriptional regulation of a-syn-uclein can also occur through endogenousmicro RNAs, which bind to the 30 end of thegene (Junn et al. 2009; Doxakis 2010). A partic-ularly interesting twist to the effects of familialpoint mutations in SNCA has been providedby a series of studies that, somewhat counterin-tuitively, suggest that the expression of the mu-tant alleles is suppressed, especially in cases withprolonged disease (Markopoulou et al. 1999;Kobayashi et al. 2003), through a mechanismthat may involve histone modifications (Vout-sinas et al. 2010). Remarkably, Voutsinas et al.

(2010) found that the WT allele is induced in acompensatory fashion, and that the total levelof SNCA messenger RNA (mRNA) exceeds thatof controls, suggesting that total SNCA levels,rather than the presence of the A53T mutant,are responsible for the disease in these cases.Although this study was based on a single pa-tient, it does bring up again the issue of totalSNCA levels as the major determinant of toxicity.

In view of the above, a reasonable avenue toattack synucleinopathies may be the loweringof a-synuclein levels. A note of caution is in or-der here, given the uncertainty about potentialneurotoxicity associated with severe reductionof a-synuclein levels, as mentioned above. Infact, Gorbatyuk et al. (2010) found that siRNA-mediated down-regulation of a-synuclein ledto quite considerable neurotoxicity within therat nigral dopaminergic system within a periodof 4 wk. In contrast, no toxicity was observedwhen small interfering RNAs (siRNAs) againsta-synuclein were injected directly into the mon-key nigra, achieving 50%–60% down-regula-tion of expression (McCormack et al. 2010).

Control of a-synuclein levels can also beachieved at the level of its degradation. a-Synu-clein degradation has been controversial, but itappears that the bulk of degradation of at leastmonomeric WT a-synuclein in neuronal cellsystems occurs through the lysosomal pathwaysof chaperone-mediated autophagy (CMA) andmacroautophagy (Paxinou et al. 2001; Webbet al. 2003; Cuervo et al. 2004; Lee et al. 2004;Vogiatzi et al. 2008; Mak et al. 2010). It hasbeen proposed that dysfunction of these degra-dation pathways may be a contributing factorto PD pathogenesis (Xilouri et al. 2008). a-Syn-uclein may also be turned over by the protea-some in other experimental settings (Tofariset al. 2001; Webb et al. 2003). A factor thatmay play a role is the exact species ofa-synucleinstudied. In any case, strategies directed towardpromoting such endogenous degradation sys-tems to enhance clearance of excess a-synucleinhave the advantage that they could also alleviatethe aberrant effects ofa-synuclein on their func-tion (see below). Indeed, stimulation of macro-autophagy through pharmacological or molecu-lar means leads to increased WT a-synuclein



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clearance and neuroprotection (Webb et al.2003; Spencer et al. 2009). Another attractivemethod of enhancing clearance of a-synucleinthrough the lysosomal pathway may be throughimmunization (Masliah et al. 2005, 2011).

POTENTIAL PATHOGENIC EFFECTS OFa-SYNUCLEIN: a-SYNUCLEIN AT THESYNAPSE

Given the physiological role of a-synuclein atthe synapse, it is not surprising that studieshave identified synaptic effects as significant de-terminants of a-synuclein-induced neurotoxic-ity in cell culture and in vivo models based onoverexpression of the protein; in some of thesecases levels of overexpression are modest, ex-pected to be present in at least the brains ofthe SNCA multiplication cases. The effectsseen include loss of presynaptic proteins, de-crease of neurotransmitter release, redistribu-tion of SNARE proteins, enlargement of synap-tic vesicles, and inhibition of synaptic vesiclerecycling (Chung et al. 2009; Garcia-Reitbocket al. 2010; Nemani et al. 2010; Scott et al.2010). It is likely that such deficits precede overtneuropathology and are causal in synaptic andneuritic degeneration, although the sequenceof synaptic events and the identification of theexact point in the cascade in which aberranta-synuclein assumes its neurotoxic potentialare still elusive.

Given the role of calcium is presynaptic sig-naling events, it is tempting to speculate thatperturbations in calcium homeostasis mayplay a role in the neuritic degeneration inducedby a-synuclein. Indeed, there is some evidencefor this, albeit still somewhat fragmentary,implicating primarily a-synuclein oligomers,which could form pores on cellular membranesor alter the properties of voltage-gated recep-tors, leading to excess calcium influx (Vollesand Lansbury 2002; Danzer et al. 2007; Hettiar-achchi et al. 2009). Through a similar porelikemechanism, oligomeric AS may also attack syn-aptic vesicles, leading to leakage of neurotrans-mitter in the cytosol; in the case of dopaminevesicles, this would mean excess cytosolic dop-amine, which could lead to oxidative stress-

induced death (Mosharov et al. 2006). Excessintracellular calcium enhanced cytosolic dopa-mine, and AS was required for death inducedby excess dopamine, creating a possible scenarioin which intracellular calcium, cystolic dopa-mine, and a-synuclein interact with each otherin a self-feeding cascade leading to neurodegen-eration (Mosharov et al. 2009). Another factorthat could be important in this setting is cal-cium-mediated calpain activation that couldlead, as mentioned, to carboxy-terminal a-syn-uclein truncation, consequent oligomerization,and further calcium influx, creating a viciouscycle.

a-SYNUCLEIN AT THE CYTOSKELETON

a-Synuclein may impact cytoskeletal dynamics.Most of the evidence suggests an association ofa-synuclein with microtubules. Various investi-gators have detected an interaction of a-synu-clein with tubulin (Alim et al. 2002; Zhouet al. 2010), but the effects reported on tubulinpolymerization are conflicting, with others re-porting enhanced (Alim et al. 2004; Lin et al.2009), and others reduced (Lee et al. 2006;Zhou et al. 2010) tubulin polymerization as aresult of a-synuclein overexpression. Further-more, tubulin depolymerization may lead toimpaired maturation of a-synuclein protofi-brils into mature fibrils, which may lead toenhanced neurotoxicity (Lee and Lee 2002).Another factor that may be important is the re-lationship of a-synuclein to tau, which is likelyimportant in the pathogenesis of sporadic dis-ease, as in GWAS the two corresponding genesare the ones showing the highest associationto PD (Nalls et al. 2011). Although tauopathyis not a prominent feature in PD, it is appreci-ated in certain cases (Kotzbauer et al. 2004).In such cases, a-synuclein and tau pathologymay occur in the same cell, but the formed ag-gregates are separate. In fact, each one of thesetwo proteins has the tendency to seed the aggre-gation of the other (Giasson et al. 2003). a-Syn-uclein appears to enhance tau phosphorylation,and this may lead to secondary effects on themicrotubular network (Jensen et al. 1999; Hag-gerty et al. 2011; Qureshi and Paudel 2011).

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Furthermore, a-synuclein may bind and in-fluence polymerization of actin, and thus per-haps indirectly affect cellular trafficking andsynaptic function; disparate effects on actinwere observed between WT and A30P mutanta-synuclein (Sousa et al. 2009). Another aspectthat appears to be affected by aberrant a-synu-clein, and may be related to the effects on thecytoskeleton, is axonal transport. In an adeno-associated virus (AAV) model of nigral synu-cleinopathy, early changes before cell deathincluded a diminution at the level of the stria-tum of motor proteins involved in anterogradetransport, and an increase of those involved inretrograde transport (Chung et al. 2009).

a-SYNUCLEIN AND PROTEINDEGRADATION SYSTEMS

An issue that has attracted a lot of attention is thepossibility that aberrant a-synuclein may im-pact protein degradation systems. This notionis attractive, as a number of neurodegenerativeconditions are associated with impairments ofprotein clearance, and such impairments havethe potential to create a vicious loop involvingaberrant a-synuclein accumulation. Aberranta-synuclein, mutant, or under certain circum-stances, WT, can induce proteasomal dysfunc-tion in a number of cell-free, cellular, and invivo settings (Stefanis et al. 2001; Tanaka et al.2001; Petrucelli et al. 2002; Snyder et al. 2003),although this is not universally observed (Mar-tin-Clemente et al. 2004). It is thought that oli-gomeric a-synuclein, and, in particular, smallsoluble oligomeric forms that are themselves de-graded by the proteasome, may exert this effect(Lindersson et al. 2004; Emmanouilidou et al.2010a). This would create the potential for thevicious cycle mentioned above.

Mutant a-synuclein may also aberrantlyimpact the lysosomes, causing impairment ofCMA (Cuervo et al. 2004; Xilouri et al. 2009;Alvarez-Erviti et al. 2010). WTAS, in an adductwith dopamine, can cause a similar effect (Mar-tinez-Vicente et al. 2008). Interestingly, the mainproteins involved in CMA, Lamp2a and Hsc70,are diminished in PD brains (Alvarez-Ervitiet al. 2010). Such CMA impairment may cause

compensatory macroautophagy induction thatmay be detrimental to neuronal survival (Xi-louri et al. 2009). The issue is, however, contro-versial, as others find that WT a-synuclein ac-tually diminishes macroautophagy, through aninhibitory action at the level of phagophore for-mation (Winslow et al. 2010).

An interesting issue that relates to a-synu-clein degradation and its effects on protein deg-radation systems is the relationship of PD toGaucher disease and glucocerebrosidase (GBA).Heterozygote and homozygote carriers of GBAmutations are at an increased risk of develop-ing PD. It is controversial whether this repre-sents a loss-of-function of GBA or a gain-of-function conferred by the mutants. Accordingto some, loss-of-function of GBA is sufficientto induce a-synuclein accumulation and aggre-gation (Manning-Bog et al. 2010; Mazzulli et al.2011; Xu et al. 2011), which may occur second-ary to lysosomal dysfunction (Mazzulli et al.2011); others find that a-synuclein accumulatesonly on overexpression of GBA mutants, favor-ing a gain of toxic function mechanism (Cullenet al. 2011). The accumulation of substratessuch as glycosylceramide owing to GBA loss-of-function may also confer increased tendencyof a-synuclein to oligomerize (Mazzulli et al.2011; Xu et al. 2011); on the other hand, oligo-meric a-synuclein may impact on GBA, leadingto its dysfunction, creating a vicious cycle of ly-sosomal dysfunction and aberrant a-synucleinaccumulation (Mazzulli et al. 2011).

a-SYNUCLEIN AT THE MITOCHONDRIAAND RELATIONSHIP TO OXIDATIVESTRESS

Many studies now have confirmed that a pro-portion of a-synuclein, endogenous or overex-pressed, resides within mitochondria, and thatit causes down-regulation of complex I activity,thus linking a-synuclein with the effects of mi-tochondrial toxins, and potentially, sporadicPD (Li et al. 2007; Devi et al. 2008; Nakamuraet al. 2008; Liu et al. 2009; Loeb et al. 2010).Furthermore, brain mitochondria morphologyis disrupted in a-synuclein transgenic mice(Martin et al. 2006). a-Synuclein, potentially



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in the form of small oligomers, appears toinduce mitochondrial fragmentation, whichmay be responsible for subsequent mitochon-drial dysfunction and death (Kamp et al. 2010,Nakamura et al. 2011). a-Synuclein, and espe-cially the mutant form, perhaps through thesame mechanism of fragmentation, also in-duces inappropriate excessive mitophagy, whichleads to mitochondrial removal and neuronaldeath (Choubey et al. 2011). Interestingly, en-hancing the expression of PGC-1a, the mastertranscriptional regulator of nuclearly encodedmitochondrial subunits, which is down-regu-lated in PD brains, leads to protection againsta-synuclein-induced neurodegeneration in cel-lular models (Zheng et al. 2010).

Through such effects on mitochondria, andespecially the complex I inhibition, a-synucleincould potentially induce reactive oxygen species(ROS) production, oxidative stress, and resultantneuronal death. There is in fact some evidencethat a-synuclein overexpression can induce ROS,and that suppressing these through molec-ular or pharmacological means may be neuro-protective in mammalian (Hsu et al. 2000; Clarket al. 2010; Liu et al. 2011) or insect systems(Wassef et al. 2007; Botella et al. 2008).

a-SYNUCLEIN AT THE ER/GOLGI

In a neuronal cell model based on adenoviralexpression of a-synuclein, Golgi fragmentationwas an early feature and correlated with the ap-pearance of soluble oligomeric species (Gosaviet al. 2002). Smith et al. (2005) observed endo-plasmic reticulum (ER) stress as an early eventin an inducible model of A53T a-synuclein-mediated neurotoxicity, and were able to inhibitdeath using a pharmacological inhibitor of ERstress, also suggesting that a primary target ofa-synuclein may be the ER/Golgi. In a blindedscreen in a yeast model, the major class of modu-lators ofa-synuclein toxicity was genes involvedin vesicular trafficking (Willingham et al. 2003).This was taken one step further by Cooper et al.(2006), who identified ER-to-Golgi traffickingas the major cellular perturbation in this yeastmodel, and Rab1, a protein involved in this tran-sition, as a critical determinant of a-synuclein-

mediated toxicity in a variety of cellular and invivo models. Thayanidhi et al. (2010) suggestedthat in mammalian systems the ER/Golgi transi-tion was delayed owing to an antagonistic effectof a-synuclein on ER/Golgi SNAREs. Over-expression of other Rabs, more closely linkedto synaptic vesicular function, such as Rab3A,also prevented dopaminergic toxicity in the a-synuclein C. elegans model, suggesting that thesephenomena may more broadly reflect the inter-action and effects of a-synuclein on vesiculardynamics (Gitler et al. 2008).

a-SYNUCLEIN IN THE NUCLEUS

Synuclein bears its name from its apparent lo-calization in the nucleus and presynatic nerveterminals (Maroteaux et al. 1988). However,nuclear localization of a-synuclein has onlybeen observed inconsistently (Mori et al.2002b; Yu et al. 2007); it actually appears that,at least in transgenic mice, a-synuclein phos-phorylated at S129 may be preferentially local-ized in the nucleus (Schell et al. 2009). a-Synu-clein has been found to interact with histoneson nuclear translocation in nigral neurons in re-sponse to the herbicide paraquat (Goers et al.2003), and to reduce histone acetylation (Kon-topoulos et al. 2006). Inhibitors of histone de-acetylase (HDAC) rescued a-synuclein-medi-ated neurotoxicity in cell culture and in vivoDrosophila models (Kontopoulos et al. 2006).Furthermore, inhibition of Sirtuin 2, a mamma-lian HDAC, which led to increased acetylation,was associated with neuroprotection in variousmodels of synucleinopathy, although whetherpotential acetylation effects on histone were in-volved was not ascertained (Outeiro et al. 2007).

a-SYNUCLEIN OUTSIDE THE CELL:THE THEORY OF PROPAGATION

Although initially a-synuclein was thought tobe a purely intracellular protein, as it lacks a sig-naling sequence that would direct it to the secre-tory pathway, it gradually became clear that itcould be detected in the conditioned mediumof cells and in extracellular fluids, such asplasma and CSF (El-Agnaf et al. 2003; Lee

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et al. 2005). The mechanism of a-synuclein se-cretion is not well understood, but it appears tooccur through a nonclassical secretory pathway(Lee et al. 2005), possibly within exosomes, en-docytic vesicles that are contained within mul-tivesicular bodies and are released on calciuminflux (Emmanouilidou et al. 2010b). The exis-tence of extracellular a-synuclein in rodent andhuman brain interstitial fluid has been con-firmed by microdialysis (Emmanouilidou et al.2011). Importantly, secreted a-synuclein canimpact neuronal homeostasis and lead to neu-ronal death, even at concentrations close tothose identified in bodily fluids, and the respon-sible species, at least in part, appear to be solubleoligomeric (Emmanouilidou et al. 2010b; Dan-zer et al. 2011). Furthermore, various reportshave shown that extracellular a-synuclein caninduce inflammatory reactions in glial cells,thus further potentiating neurodegeneration(Zhang et al. 2005; Klegeris et al. 2008; Leeet al. 2010). It should be noted, however, thatsuch studies are often based on high concentra-tions of recombinant a-synuclein that may notbe physiologic. This issue is also touched on byGendelman and colleagues (Mosley et al. inpress).

A related aspect of a-synuclein pathobiol-ogy that has attracted a lot of attention is its ap-parent ability to be uptaken by cells, although,at least in cell culture, this is highly dependenton the cell type and the particular species ofa-synuclein and is not easily achieved in intactneuronal cells (Emmanouilidou et al. 2010b).Oligomeric-aggregated a-synuclein species ap-pear to be especially prone to uptake and havethe potential to “seed” fibrillization of endoge-nous a-synuclein (Danzer et al. 2007, 2009,2011; Luk et al. 2009; Nonaka et al. 2010; Wax-man and Giasson 2010; Hansen et al. 2011).This observation has created a lot of excitement,as it could potentially explain the propagationof pathology observed in PD, according to theBraak hypothesis of staging of the disease. Twokey in vivo observations support this “prionlikepropagation” theory. On the one hand, progen-itor cells implanted in the hippocampus of re-cipient mice transgenic for a-synuclein incor-porated host a-synuclein in their cell soma

(Desplats et al. 2009), and fetal dopaminergicneurons also showed uptake of hosta-synuclein(Hansen et al. 2011); on the other, fetal dopami-nergic grafts in humans showed LB pathologymany years after implantation, although wheth-er the a-synuclein within the grafted neuronswas host-derived could not be determined withcertainty (Kordower et al. 2008). An intriguingstudy has brought together the mitochondrialdysfunction with the a-synuclein propagationtheories. Intragastric administration of the com-plex I inhibitor rotenone led to enteric a-synu-clein pathology, which gradually spread to thecentral nervous system (CNS), including dopa-minergic neurons, and caused delayed neuro-toxicity, suggesting that, once initiated by mito-chondrial dysfunction, a-synuclein aberrantconformations assume an independent propa-gating seeding and neurotoxic potential (Pan-Montojo et al. 2010). This would be consistentwith the findings of early enteric a-synucleinpathology in PD (Braak et al. 2006) and in a-synuclein transgenic mice (Kuo et al. 2010). Anote of caution is indicated in the general as-sumption that seeding is equated with neuro-toxicity. In fact, seeding and toxicity of extracel-lular a-synuclein can be dissociated (Danzeret al. 2007).

a-SYNUCLEIN BIOMARKERS

In view of the importance of a-synuclein inPD pathogenesis, a number of studies have at-tempted to use measurements of a-synucleinas biomarkers for the disease and for othersynucleinopathies. In the cerebrospinal fluid(CSF), most studies appear to show a reductionof total a-synuclein levels in PD patients com-pared with controls (Tokuda et al. 2006; Mol-lenhauer et al. 2008, 2011; Hong et al. 2010; Ka-suga et al. 2010); in fact, a comprehensive studysuggested thata-synuclein levels, measured by asensitive ELISA technique, may help to differen-tiate synucleinopathies, where a-synuclein isreduced compared with other neurodegenera-tive conditions, such as tauopathies or amy-loidoses (Mollenhauer et al. 2011). In contrast,oligomeric a-synuclein, measured by a specificELISA method, was higher in PD compared



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with control CSF (Tokuda et al. 2010), extend-ing previous findings of identification of solubleoligomeric a-synuclein in DLB brain tissue(Paleologou et al. 2009), but this intriguing re-sult awaits replication in a larger cohort. Datafrom a-synuclein measurements in peripheraltissues/fluids have been inconsistent, and over-all peripherala-synuclein levels, despite the easeof obtaining the material, do not appear to havepotential as biomarkers.

CONCLUDING REMARKS

It is clear from the above thata-synuclein repre-sents a valid therapeutic target in PD and possi-bly in related synucleinpathies. Possible thera-peutic strategies that could be envisioned aredepicted in Figure 2. Perhaps the most promis-ing strategy would involve small molecules thatwould have the potential to alter the confor-mation of protofibrillar forms of a-synucleinand render them nonpathogenic. An exampleof such natural products could be the greentea derivative EGCG (Bieschke et al. 2010) orthe natural product scyllo inositol (Vekrelliset al. 2009). It may be that multiple therapeuticapproaches will need to be used, in view of thefeed-forward amplification loops involved inthe pathogenic effects of a-synuclein.

ACKNOWLEDGMENTS

L.S. was funded for PD-related research by ECFP7 Programmes NEURASYNC and MEFOPA.

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Preventingspecific aberrant

functions

Synapticvesicles

Proteasomes

Lysosomes

ER-Golgi

Cell membrane/Calcium

Mitochondria

Axonal transport

Promoting fibrilformation

Promoting off-pathwayoligomers

Inhibitingoligomer/protofibril

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EnhancingdegradationModulating

transcription

Inhibitingproaggregationposttranslational

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December 13, 20112012; doi: 10.1101/cshperspect.a009399 originally published onlineCold Spring Harb Perspect Med

Leonidas Stefanis

-Synuclein in Parkinson's Diseaseα

Subject Collection Parkinson's Disease

Functional Neuroanatomy of the Basal Ganglia

ObesoJosé L. Lanciego, Natasha Luquin and José A. Dysfunction in Parkinson's Disease

as a Model to Study MitochondrialDrosophila

Ming Guo

GeneticsAnimal Models of Parkinson's Disease: Vertebrate

DawsonYunjong Lee, Valina L. Dawson and Ted M. Pathways

and DJ-1 and Oxidative Stress and Mitochondrial Parkinsonism Due to Mutations in PINK1, Parkin,

Mark R. CooksonInnate Inflammation in Parkinson's Disease

V. Hugh PerryProgrammed Cell Death in Parkinson's Disease

Katerina Venderova and David S. Park

NeuropathologyParkinson's Disease and Parkinsonism:

Dennis W. DicksonDiseaseGenomics and Bioinformatics of Parkinson's

al.Sonja W. Scholz, Tim Mhyre, Habtom Ressom, et

Parkinson's DiseasePhysiological Phenotype and Vulnerability in

Sanchez, et al.D. James Surmeier, Jaime N. Guzman, Javier

DiseaseMotor Control Abnormalities in Parkinson's

CortésPietro Mazzoni, Britne Shabbott and Juan Camilo

ManagementFeatures, Diagnosis, and Principles of Clinical Approach to Parkinson's Disease:

João Massano and Kailash P. Bhatia

Parkinson's Disease: Gene Therapies

Patrick AebischerPhilippe G. Coune, Bernard L. Schneider and

The Role of Autophagy in Parkinson's DiseaseMelinda A. Lynch-Day, Kai Mao, Ke Wang, et al.

Functional Neuroimaging in Parkinson's Disease

EidelbergMartin Niethammer, Andrew Feigin and David

Parkinson's DiseaseDisruption of Protein Quality Control in

PetrucelliCasey Cook, Caroline Stetler and Leonard

Key QuestionsLeucine-Rich Repeat Kinase 2 for Beginners: Six

Lauren R. Kett and William T. Dauer

http://perspectivesinmedicine.cshlp.org/cgi/collection/ For additional articles in this collection, see

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