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Protein coding mutations in leucine-rich repeat kinase 2 (LRRK2) cause familial Parkinson's disease (PD), and noncoding variations around the gene increase the risk of developing sporadic PD. It is generally accepted that pathogenic LRRK2 mutations increase LRRK2 kinase activity, resulting in a toxic hyperactive protein that is inferred to lead to the PD phenotype. LRRK2 has long been linked to different membrane trafficking events, but the specific role of LRRK2 in these events has been difficult to resolve. Recently, several papers have reported the activation and translocation of LRRK2 to cellular organelles under specific conditions, which suggests that LRRK2 may influence intracellular membrane trafficking. Here, we review what is known about the role of LRRK2 at various organelle compartments.

The progressive neuropathological damage seen in Parkinson's disease (PD) is thought to be related to the spreading of aggregated forms of α‐synuclein. Clearance of extracellular α‐synuclein released by degenerating neurons may be therefore a key mechanism to control the concentration of α‐synuclein in the extracellular space. Several molecular chaperones control misfolded protein accumulation in the extracellular compartment. Among these, clusterin, a glycoprotein associated with Alzheimer's disease, binds α‐synuclein aggregated species and is present in Lewy bodies, intraneuronal aggregates mainly composed by fibrillary α‐synuclein. In this study, using murine primary astrocytes with clusterin genetic deletion, human‐induced pluripotent stem cell (iPSC)‐derived astrocytes with clusterin silencing and two animal models relevant for PD we explore how clusterin affects the clearance of α‐synuclein aggregates by astrocytes. Our findings showed that astrocytes take up α‐synuclein preformed fibrils (pffs) through dynamin‐dependent endocytosis and that clusterin levels are modulated in the culture media of cells upon α‐synuclein pffs exposure. Specifically, we found that clusterin interacts with α‐synuclein pffs in the extracellular compartment and the clusterin/α‐synuclein complex can be internalized by astrocytes. Mechanistically, using clusterin knock‐out primary astrocytes and clusterin knock‐down hiPSC‐derived astrocytes we observed that clusterin limits the uptake of α‐synuclein pffs by cells. Interestingly, we detected increased levels of clusterin in the adeno‐associated virus‐ and the α‐synuclein pffs‐ injected mouse model, suggesting a crucial role of this chaperone in the pathogenesis of PD. Overall, our observations indicate that clusterin can limit the uptake of extracellular α‐synuclein aggregates by astrocytes and, hence, contribute to the spreading of Parkinson pathology., Main Points Extracellular clusterin binds α‐synuclein fibrils thus limiting their uptake by murine and human astrocytes.Reduced levels of clusterin improve α‐synuclein aggregates clearance by astrocytes.

OBJECTIVE The goal of this study was to refine our understanding of disease risk attributable to common genetic variation in SNCA, a major locus in Parkinson disease, with potential implications for clinical trials targeting α-synuclein. We aimed to dissect the multiple independent association signals, stratify individuals by SNCA-specific risk profiles, and explore expression quantitative trait loci. METHODS We analyzed participant-level data from 12,503 patients and 12,502 controls, optimizing a risk model and assessing SNCA-specific risk scores and haplotypes as predictors of individual risk. We also explored hypotheses about functional mechanisms and correlated risk variants to gene expression in human brain and protein levels in cerebrospinal fluid. RESULTS We report and replicate a novel, third independent association signal at genome-wide significance level downstream of SNCA (rs2870004, p = 3.0*10-8 , odds ratio [OR] = 0.88, 95% confidence interval [CI] = 0.84-0.92). SNCA risk score stratification showed a 2-fold difference in disease susceptibility between top and bottom quintiles (OR = 1.99, 95% CI = 1.78-2.23). Contrary to previous reports, we provide evidence supporting top variant rs356182 as functional in itself and associated with a specific SNCA 5' untranslated region transcript isoform in frontal cortex. INTERPRETATION The SNCA locus harbors a minimum of 3 independent association signals for Parkinson disease. We demonstrate a fine-grained stratification of α-synuclein-related genetic burden in individual patients of potential future clinical relevance. Further efforts to pinpoint the functional mechanisms are warranted, including studies of the likely causal top variant rs356182 and its role in regulating levels of specific SNCA mRNA transcript variants. Ann Neurol 2018;83:117-129.

Background Dementia with Lewy bodies is characterized by transient clinical features, including fluctuating cognition and visual hallucinations, implicating dysfunction of cerebral hub regions, such as the pulvinar nuclei of the thalamus. However, the pulvinar is typically only mildly affected by Lewy body pathology in dementia with Lewy bodies, suggesting additional factors may account for its proposed dysfunction. Methods We conducted a comprehensive analysis of postmortem pulvinar tissue using whole-transcriptome RNA sequencing, protein expression analysis, and histological evaluation. Results We identified 321 transcripts as significantly different between dementia with Lewy bodies cases and neurologically normal controls, with gene ontology pathway analysis suggesting the enrichment of transcripts related to synapses and positive regulation of immune functioning. At the protein level, proteins related to synaptic efficiency were decreased, and general synaptic markers remained intact. Analysis of glial subpopulations revealed astrogliosis without activated microglia, which was associated with synaptic changes but not neurodegenerative pathology. Discussion These results indicate that the pulvinar, a region with relatively low Lewy body pathological burden, manifests changes at the molecular level that differ from previous reports in a more severely affected region. We speculate that these alterations result from neurodegenerative changes in regions connected to the pulvinar and likely contribute to a variety of cognitive changes resulting from decreased cortical synchrony in dementia with Lewy bodies. © 2018 International Parkinson and Movement Disorder Society.

This Preface introduces the articles of the special issue on 'Proteomics' where we survey these powerful techniques specifically in the context of applications in neurosciences. Proteomics as used here is a catch-all term for approaches where a broad survey of all protein content of a cell or tissue is performed in parallel. In the special issue, various experts outline applications as diverse as finding disease biomarkers in human samples, including living participants or post-mortem brain, as well as application of protein technologies to model systems such as animals and cells. Collectively, these articles outline the utility of current technologies for proteome interrogation and identify enhancements that will shape applications to future studies. This is the Preface for the special issue "Proteomics". Cover Image for this issue: doi: 10.1111/jnc.14530.

Leucine-rich repeat kinase 2 (LRRK2) is a causative gene for Parkinson's disease, but the physiological function and the mechanism(s) by which the cellular activity of LRRK2 is regulated are poorly understood. Here, we identified p21-activated kinase 6 (PAK6) as a novel interactor of the GTPase/ROC domain of LRRK2. p21-activated kinases are serine-threonine kinases that serve as targets for the small GTP binding proteins Cdc42 and Rac1 and have been implicated in different morphogenetic processes through remodeling of the actin cytoskeleton such as synapse formation and neuritogenesis. Using an in vivo neuromorphology assay, we show that PAK6 is a positive regulator of neurite outgrowth and that LRRK2 is required for this function. Analyses of post-mortem brain tissue from idiopathic and LRRK2 G2019S carriers reveal an increase in PAK6 activation state, whereas knock-out LRRK2 mice display reduced PAK6 activation and phosphorylation of PAK6 substrates. Taken together, these results support a critical role of LRRK2 GTPase domain in cytoskeletal dynamics in vivo through the novel interactor PAK6, and provide a valuable platform to unravel the mechanism underlying LRRK2-mediated pathophysiology. We propose p21-activated kinase 6 (PAK6) as a novel interactor of leucine-rich repeat kinase 2 (LRRK2), a kinase involved in Parkinson's disease (PD). In health, PAK6 regulates neurite complexity in the brain and LRRK2 is required for its function, (a) whereas PAK6 is aberrantly activated in LRRK2-linked PD brain (b) suggesting that LRRK2 toxicity is mediated by PAK6. ispartof: Journal of Neurochemistry vol:135 issue:6 pages:1242-56 ispartof: location:England status: published

Substantial progress has been made in the genetic basis of Parkinson's disease (PD). In particular, by identifying genes that segregate with inherited PD or show robust association with sporadic disease, and by showing the same genes are found on both lists, we have generated an outline of the cause of this condition. Here, we will discuss what those genes tell us about the underlying biology of PD. We specifically discuss the relationships between protein products of PD genes and show that common links include regulation of the autophagy-lysosome system, an important way by which cells recycle proteins and organelles. We also discuss whether all PD genes should be considered to be in the same pathway and propose that in some cases the relationships are closer, whereas in other cases the interactions are more distant and might be considered separate. Beilina and Cookson review the links between genes for Parkinson's disease (red) and the autophagy-lysosomal system. They propose the hypothesis that many of the known PD genes can be assigned to pathways that affect (I) turnover of mitochondria via mitophagy (II) turnover of several vesicular structures via macroautophagy or chaperone-mediated autophagy or (III) general lysosome function. This article is part of a special issue on Parkinson disease.

The oxidation of a key cysteine residue (Cys106) in the parkinsonism-associated protein DJ-1 regulates its ability to protect against oxidative stress and mitochondrial damage. Cys106 interacts with a neighboring protonated Glu18 residue, stabilizing the Cys106-SO2 (-) (sulfinic acid) form of DJ-1. To study this important post-translational modification, we previously designed several Glu18 mutations (E18N, E18D, E18Q) that alter the oxidative propensity of Cys106. However, recent results suggest these Glu18 mutations cause loss of DJ-1 dimerization, which would severely compromise the protein's function. The purpose of this study was to conclusively determine the oligomerization state of these mutants using X-ray crystallography, NMR spectroscopy, thermal stability analysis, circular dichroism spectroscopy, sedimentation equilibrium ultracentrifugation, and cross-linking. We found that all of the Glu18 DJ-1 mutants were dimeric. Thiol cross-linking indicates that these mutant dimers are more flexible than the wild-type protein and can form multiple cross-linked dimeric species due to the transient exposure of cysteine residues that are inaccessible in the wild-type protein. The enhanced flexibility of Glu18 DJ-1 mutants provides a parsimonious explanation for their lower observed cross-linking efficiency in cells. In addition, thiol cross-linkers may have an underappreciated value as qualitative probes of protein conformational flexibility. DJ-1 is a homodimeric protein that protects cells against oxidative stress. Designed mutations that influence the regulatory oxidation of a key cysteine residue have recently been proposed to disrupt DJ-1 dimerization. We use cysteine cross-linking and various biophysical techniques to show that these DJ-1 mutants form dimers with increased conformational flexibility.

Recent genome wide association studies have identified CLU, CR1, ABCA7 BIN1, PICALM and MS4A6A/MS4A6E in addition to the long established APOE, as loci for Alzheimer's disease. We have systematically examined each of these loci to assess whether common coding variability contributes to the risk of disease. We have also assessed the regional expression of all the genes in the brain and whether there is evidence of an eQTL explaining the risk. In agreement with other studies we find that coding variability may explain the ABCA7 association, but common coding variability does not explain any of the other loci. We were not able to show that any of the loci had eQTLs within the power of this study. Furthermore the regional expression of each of the loci did not match the pattern of brain regional distribution in Alzheimer pathology. Although these results are mainly negative, they allow us to start defining more realistic alternative approaches to determine the role of all the genetic loci involved in Alzheimer's disease. © 2013 Blackwell Publishing Ltd/University College London.

Gene expression is regulated at many levels, including after generation of the primary RNA transcript from DNA but before translation into protein. Such post-translational gene regulation occurs via the action of a multitude of RNA binding proteins and include varied actions from splicing to regulation of association with the translational machinery. Primary evidence that such processes might contribute to disease mechanisms in neurodegenerative disorders comes from the observation of mutations in RNA binding proteins, particularly in diseases in the amyotrophic lateral sclerosis-frontotemporal dementia spectrum and in some forms of ataxia and tremor. The bulk of evidence from recent surveys of the types of RNA species that are affected in these disorders suggests a global deregulation of control rather than a very small number of RNA species, although why some groups of neurons are sensitive to these changes is not well understood. Overall, these data suggest that neurodegeneration can be initiated by mutations in RNA binding proteins and, as a corollary, that neurons are particularly sensitive to loss of control of gene expression at the post-transcriptional level. Such observations have implications not only for understanding the nature of neurodegenerative disorders but also how we might intervene therapeutically in these diseases. WIREs RNA 2017, 8:e1397. doi: 10.1002/wrna.1397 For further resources related to this article, please visit the WIREs website.

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most prevalent known cause of autosomal dominant Parkinson's disease. The LRRK2 gene encodes a Roco protein featuring a Ras of complex proteins (ROC) GTPase and a kinase domain linked by the C-terminal of ROC (COR) domain. Here, we explored the effects of the Y1699C pathogenic LRRK2 mutation in the COR domain on GTPase activity and interactions within the catalytic core of LRRK2. We observed a decrease in GTPase activity for LRRK2 Y1699C comparable to the decrease observed for the R1441C pathogenic mutant and the T1348N dysfunctional mutant. To study the underlying mechanism, we explored the dimerization in the catalytic core of LRRK2. ROC-COR dimerization was significantly weakened by the Y1699C or R1441C/G mutation. Using a competition assay, we demonstrated that the intra-molecular ROC : COR interaction is favoured over ROC : ROC dimerization. Interestingly, the intra-molecular ROC : COR interaction was strengthened by the Y1699C mutation. This is supported by a 3D homology model of the ROC-COR tandem of LRRK2, showing that Y1699 is positioned at the intra-molecular ROC : COR interface. In conclusion, our data provides mechanistic insight into the mode of action of the Y1699C LRRK2 mutant: the Y1699C substitution, situated at the intra-molecular ROC : COR interface, strengthens the intra-molecular ROC : COR interaction, thereby locally weakening the dimerization of LRRK2 at the ROC-COR tandem domain resulting in decreased GTPase activity.

Genetic forms of parkinsonism are interesting for two particular reasons. First, finding a gene identifies a cause for a disease that would otherwise be unexplained. Second, finding several genes for the same disorder allows us to reconstruct molecular pathways that, in the example of Parkinson's disease, are be associated with the survival of dopamine neurons in the substantia nigra. Two rare causes of parkinsonism, DJ-1 and PINK1, are associated with mitochondria. This organelle has long been linked with Parkinson's disease, and recent results are starting to show how mutations impact mitochondrial function. In this short review, I will discuss how we can use some of this information to understand why it is that neurons become dysfunctional in PD.

Mutations in the E3 ubiquitin ligase parkin cause early-onset, autosomal-recessive juvenile parkinsonism (AJRP), presumably as a result of a lack of function that alters the level, activity, aggregation or localization of its substrates. Recently, we have reported that phospholipase Cgamma1 is a substrate for parkin. In this article, we show that parkin mutants and siRNA parkin knockdown cells possess enhanced levels of phospholipase Cgamma1 phosphorylation, basal phosphoinositide hydrolysis and intracellular Ca2+ concentration. The protein levels of Ca2+-regulated protein kinase Calpha were decreased in AJRP parkin mutant cells. Neomycin and dantrolene both decreased the intracellular Ca2+ levels in parkin mutants in comparison with those seen in wild-type parkin cells, suggesting that the differences were a consequence of altered phospholipase C activity. The protection of wild-type parkin against 6-hydroxydopamine (6OHDA) toxicity was also established in ARJP mutants on pretreatment with dantrolene, implying that a balancing Ca2+ release from ryanodine-sensitive stores decreases the toxic effects of 6OHDA. Our findings suggest that parkin is an important factor for maintaining Ca2+ homeostasis and that parkin deficiency leads to a phospholipase C-dependent increase in intracellular Ca2+ levels, which make cells more vulnerable to neurotoxins, such as 6OHDA.

Heterozygous loss-of-function mutations at the glucosecerebrosidase locus have recently been shown to be a potent risk factor for Lewy body disease. Based on this observation, we have re-evaluated the likelihood that the different PARK loci (defined using clinical criteria for disease) may be misleading attempts to find common pathways to pathogenesis. Rather, we suggest, grouping the different loci which lead to different Lewy body disease may be more revealing. Doing this, we suggest that several of the genes involved in disparate Lewy body diseases impinge on ceramide metabolism and we suggest that this may be a common theme for pathogenesis.

Until 10 years ago, conventional wisdom held that Parkinson's disease was not a genetic disorder. Since that time, there have been a plethora of genetic findings, culminating in the cloning of several genes that derive from the loci given the nomenclature PARK1-PARK12 (OMIM 168600). Recently, these research findings have begun to impact clinical practice, and this impact is likely to increase. The primary purpose of this article is to outline these genetic advances, discuss their importance for current practice in clinical and related settings, and outline briefly how they are influencing research into the causes of and possible future treatments for this prevalent disorder.

Neurofilament pathology is a hallmark of sporadic and familial amyotrophic lateral sclerosis (SALS and FALS). The disease mechanisms underlying this pathology are presently unclear, but recent evidence in SALS patients suggest that reductions in neurofilament light subunit (NFL) mRNA may contribute to the death of motor neurones. Mutations in the gene encoding Cu-Zn superoxide dismutase (SOD1) represent the best-studied cause of FALS, and a number of laboratory models of SOD1-mediated disease exist. Here we have used microdissected lumbar spinal cord motor neurones from human SOD1 FALS patients as well as G93A SOD1 transgenic mice and demonstrated that reduced NFL mRNA levels are seen in both. To probe the molecular mechanisms underpinning these observations, we generated NSC34 motor neurone-like cell lines expressing wild-type and mutant SOD1. NSC34 cells expressing G37R or G93A SOD1 showed selective reductions in NFL and NFM mRNA and protein. These data suggest that NFL mRNA reductions are common to SALS and FALS patients, and that cells and mice expressing mutant SOD1 may enable us to characterize the molecular mechanism(s) responsible for the loss of neurofilament mRNA.

Abnormal accumulation of alpha-synuclein in Lewy bodies is a neuropathological hallmark of both sporadic and familial Parkinson's disease (PD). Although mutations in alpha-synuclein have been identified in autosomal dominant PD, the mechanism by which dopaminergic cell death occurs remains unknown. We investigated transcriptional changes in neuroblastoma cell lines transfected with either normal or mutant (A30P or A53T) alpha-synuclein using microarrays, with confirmation of selected genes by quantitative RT-PCR. Gene products whose expression was found to be significantly altered included members of diverse functional groups such as stress response, transcription regulators, apoptosis-inducing molecules, transcription factors and membrane-bound proteins. We also found evidence of altered expression of dihydropteridine reductase, which indirectly regulates the synthesis of dopamine. Because of the importance of dopamine in PD, we investigated the expression of all the known genes in dopamine synthesis. We found co-ordinated downregulation of mRNA for GTP cyclohydrolase, sepiapterin reductase (SR), tyrosine hydroxylase (TH) and aromatic acid decarboxylase by wild-type but not mutant alpha-synuclein. These were confirmed at the protein level for SR and TH. Reduced expression of the orphan nuclear receptor Nurr1 was also noted, suggesting that the co-ordinate regulation of dopamine synthesis is regulated through this transcription factor.

The reaction of superoxide and nitric oxide results in the formation of peroxynitrite, a long lived and highly reactive oxidant species. It has been suggested that the formation of peroxynitrite in vivo may contribute to cell death in some neurological conditions. We have examined the effect of peroxynitrite on cell death in the NSC34 spinal cord cell line. A brief (30 min) exposure to either peroxynitrite or hydrogen peroxide caused delayed cell death with an EC50 for both of approximately 1 mM. Cell death was prevented by the RNA synthesis inhibitor actinomycin D and included DNA damage as an early event. We sought to clarify the potential role of the DNA binding enzyme poly(ADP-ribose) polymerase (PARP) in cell death in these cells. Several PARP inhibitors [benzamide, 3-aminobenzamide, nicotinamide, and 6(5H)-phenanthridinone] prevented cell death, but the inactive analogue benzoic acid did not. However, there was no evidence of cleavage of PARP, which occurs in apoptosis via the activation of the caspase CPP32. Therefore, we suggest that PARP contributes to neuronal injury as an early event, probably by lethal NAD depletion, without any requirement for proteolytic cleavage.

Compound mutations and homozygous loss of function of the parkin gene causes juvenile and early onset, autosomal recessive parkinsonism. Pathologically, the disease is associated with loss of dopaminergic neurons in the substantia nigra pars compacta and locus ceruleus, usually without Lewy body pathology. Hemizygous families have been described that may harbor mutations outside of the open reading frame. The parkin gene promoter has yet to be characterized, and therein, mutations in hemizygous families may plausibly be identified. To identify the promoter of the parkin gene, the transcription start site was defined by a combination of primer extension and 5' RACE. Five kilobases of DNA 5' to the parkin start codon were directly sequenced from a BAC containing parkin exon 1 and evaluated for promoter motifs. The parkin promoter lacks TATA or CAAT boxes and appears to share homology to the alpha-synuclein promoter. Deletion constructs demonstrated core promoter activity and tissue specific enhancing regions in HEK-293T and SH-SY5Y cells.

Emerging evidence suggests that a disturbance of the glutamate neurotransmitter system may be a contributory factor to motor neuron injury in motor neuron disease. Previous autoradiographic and immunoblotting studies have suggested that there may be reduced expression of glutamate transporter proteins in pathologically affected areas of the CNS in motor neuron disease. This study further explores the possible alteration in expression of the excitatory amino acid transporter protein EAAT2 in MND, by examining the protein expression in situ, in frozen sections, using immunohistochemistry. The aim of the study was to compare the distribution and density of EAAT2 in the motor cortex and spinal cord of MND cases (n = 16) compared with neurologically normal controls (n = 12), matched for relevant parameters. A novel, previously characterized, monoclonal antibody to EAAT2 was employed. EAAT2 immunoreactivity in motor neuron disease and control cases was compared using relative optical density measurements generated by computerized image analysis. In the motor cortex, EAAT2 immunoreactivity was laminated comprising a superficial intense band (corresponding to layers 1 and 2); a paler middle band (layer 3 and part of 5) and a more intense deep layer (layers 5 and 6). In the spinal cord, the ventral horn showed strong immunoreactivity with dense perisomatic staining around motor neuron cell bodies, the substantia gelatinosa showed moderate diffuse staining and the intermediate spinal laminae showed weak staining. This general pattern of immunoreactivity was preserved in the motor neuron disease cases. However, in the motor neuron disease cases compared with controls, the optical density values for EAAT2 immunoreactivity were significantly reduced in all grey matter regions of the lumbar spinal cord (P < 0.001) and were increased in the middle laminae of the motor cortex (P < 0.05). This study indicates that glutamate transporter pathology in motor neuron disease may be a more complex phenomenon than previously recognized.

Convergent biochemical and genetic evidence suggests that the formation of alpha-synuclein (alpha-syn) protein deposits is an important and, probably, seminal step in the development of Parkinson's disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). It has been reported that transgenic animals overexpressing human alpha-syn develop lesions similar to those found in the brain in PD, together with a progressive loss of dopaminergic cells and associated abnormalities of motor function. Inhibiting and/or reversing alpha-syn self-aggregation could, therefore, provide a novel approach to treating the underlying cause of these diseases. We synthesized a library of overlapping 7-mer peptides spanning the entire alpha-syn sequence, and identified amino acid residues 64-100 of alpha-syn as the binding region responsible for its self-association. Modified short peptides containing alpha-syn amino acid sequences from part of this binding region (residues 69-72), named alpha-syn inhibitors (ASI), were found to interact with full-length alpha-syn and block its assembly into both early oligomers and mature amyloid-like fibrils. We also developed a cell-permeable inhibitor of alpha-syn aggregation (ASID), using the polyarginine peptide delivery system. This ASID peptide was able to inhibit the DNA damage induced by Fe(II) in neuronal cells transfected with alpha-syn(A53T), a familial PD-associated mutation. ASI peptides without this delivery system did not reverse levels of Fe(II)-induced DNA damage. Furthermore, the ASID peptide increased (P

In this issue of the Journal of Neurochemistry, Rotermund et al report that manipulation of fats in the brain promotes pathology that is related to Parkinson’s disease (Rotermund, Truckenmuller, Schell, & Kahle, 2014). This observation has impact not only on how we think about disease but also practical implications for disease modeling in rodents. The essential manipulation reported by Rotermund et al is to change the diet of mice expressing the a-synuclein A30P mutation that causes Parkinson’s disease in humans, from standard lab chow to a diet containing about ten times the amount of total fat. These mice have brainstem pathology that is reminiscent of Parkinson’s disease and die a little earlier than wild type littermates. The essential results of Rotermund et al are that the high fat diet caused the A30P mice to die earlier and have greater numbers of phosphorylated a-synuclein pathology in the brainstem. Although all the details of the mechanism(s) are not completely worked out, the authors suggest that Akt phosphorylation might be involved (Rotermund et al., 2014). To understand why the experiment might have worked, it might be helpful to consider the rationale for manipulating fats. One advanced by the authors is the idea that age is a major driver of PD risk and also a driver of metabolic disorders, specifically diabetes (reviewed in: Zhang & Tian, 2014). The effects in humans are not huge. For example, in one study there was an ~80% increase in risk of PD if a person had diabetes prior to development of their motor symptoms, although the number of cases with diabetes is small and therefore the confidence interval around the estimates are broad (Hu, Jousilahti, Bidel, Antikainen, & Tuomilehto, 2007). Also, whether the human risk is related to diet, obesity, diabetes, aging or all (or none) of the above is hard to disentangle from the extant epidemiological data. Nonetheless, the available data suggests that manipulating diet in animals might be a reasonable way to manipulate pathology in animals. Additionally, alpha-synuclein, the main pathological protein in PD, is a lipid binding protein and there are some reasons to think that lipid binding is important in this proteins normal function (Antonny, 2011) and pathogenesis (Cookson, 2005). Specifically, the form of alpha-synuclein used here, A30P, has lower lipid binding ability compared to the normal protein. Hence, manipulation of dietary lipids might affect PD-like pathology by mimicking a pro-pathological syndrome in the whole body and/or by changing brain lipid composition that might then change synuclein function. Determining which of these are critical is something that can be tested in future experiments. For example, increasing caloric content of food without changing lipid composition should impact the whole body with lesser effects on the lipid composition of membranes. However, at least one other recent study suggested that expression of a different mutant of a-synuclein (A53T, which does bind lipids) is associated with a resistant to diet induced obesity. Therefore, the interactions between synuclein and diets are complex and need further resolution. The other aspect of the paper that is pragmatically important is the idea that our animal models can be improved, that is pathology can be promoted by factors other than gene manipulation. Many transgenic models only partially recapitulate human disease pathology within the lifespan of mice (Bezard, Yue, Kirik, & Spillantini, 2013). Therefore, having ways in which we can manipulate animals to promote neuronal damage might be particularly helpful. One limitation to simply making animals obese is that the manipulation itself might easily impact some motor phenotypes. However, if studying pathology is the main focus, then diet would be a reasonable and easy to perform adjustment to standard mouse housing conditions. It would be very interesting to see if a similar dietary manipulation provokes dopaminergic cell death in models such as viral over-expression of a-synuclein (Fiandaca & Federoff, 2014). Such an approach might allow for the comparison of different mutant forms of a-synuclein. One area where I am not certain how the results should be interpreted is whether this animal model supports a diabetes-PD link specifically, or a environment-gene link generally. On the one hand, both this model and the reported epidemiological data point in the same direction (i.e., more diabesity/obesity with higher PD risk) even if both are relatively small effects. On the other hand, human epidemiology and mouse models are quite different and should each be considered separately in terms of their need to be replicated – and with small effect sizes the need for replication is very high. An additional argument against a common mechanism is that there have been genomewide association studies in both type 2 diabetes and PD that have nominated different genetic loci, suggesting independent mechanisms of risk of these two diseases. Identifying the detailed mechanisms of how diet induces a-synuclein pathology in the mouse model should be instructive for future directions in identifying disease mechanisms in human PD. For example, if there is a link between diet and kinase activation then one ought to be able to see the same activation in human samples, perhaps separating PD patients who had diabetes in lifetime from those that did not. Overall, the paper by Rotermund et al provides some important new directions in understanding perhaps the mechanisms involved in human Parkinson’s disease and provides an interesting way to make phenotypes in mice somewhat worse. How these types of approaches might be useful to understand other neurodegenerative diseases remains to be established.

Since the German anatomist Rudolf Virchow first coined the term neuroglia in 1846, our ideas about glial cells have moved from viewing them simply as the glue that holds neurons in place. We now know that glia are intimately involved in the lives of neurons, with functions ranging from nurturing developing synapses to actually facilitating the transmission of neuronal information (Eroglu & Barres 2010). Furthermore, glial cells may play detrimental roles in the pathogenesis of nervous system diseases. As an example of this, work from Hinkle and colleagues in this issue of the Journal of Neurochemistry highlight a potential role of astrocytes in Parkinson’s disease associated with mutations in DJ-1 (Mullett & Hinkle 2011). Parkinson’s disease (PD) has been considered a disease of neuronal origin due to the degeneration of groups of neurons, including dopaminergic cells in the substantia nigra pars compacta (SNpc). Mitochondrial complex I dysfunction, associated with accumulation of reactive oxygen species, has been suggested to be important in the loss of neurons in PD. This hypothesis is supported by multiple lines of evidence including the observations that; (1) post mortem biochemical studies have demonstrated lower complex I activity in the SNpc of PD brains compared to controls (Schapira 2007); (2) inhibition of complex I with rotenone or MPTP reproduces the pathological degeneration of the SNpc in mammalian models (Cannon & Greenamyre 2010); and (3) the protein products of genes known to cause familial forms of parkinsonism have clear roles in the maintenance of functional mitochondria (Thomas et al. 2011). A rare form of autosomal recessive parkinsonism is caused by mutations or deletions of the gene coding for the protein DJ-1 (Bonifati et al. 2003). DJ-1 is known to have a role in oxidative stress responses in cells and this is thought to be important in the preservation of neuronal viability. Some of the mechanisms involved are known. For example, the oxidation of a specific cysteine residue in DJ-1 is required to protect cells against mitochondrial damage (Blackinton et al. 2009). However, previous studies have focused on cell-autonomous effects of DJ-1. The work presented here by the Hinkle group further supports their previous findings which suggest a novel mode of action for DJ-1 in which it protects neurons via an astrocyte-mediated mechanism (Mullett & Hinkle 2009). Using a new method of detecting neuronal death in astrocyte/neuron co-cultures, the authors demonstrate that the down-regulation of astrocytic DJ-1 diminishes their ability to protect neurons from stressors. In particular, they show that neuroprotection is selective for drugs that inhibit mitochondrial complex I. However, the authors also found that the neuronal death in their DJ-1 deficient co-cultures treated with these inhibitors could not be rescued by the addition of antioxidants. They also demonstrate that the mechanism underlying astrocyte-mediated protection is not due to the release of the antioxidant glutathione or an up-regulation of astrocytic heme oxygenase-1. Taken together, these findings suggest that astrocytic DJ-1 is necessary for astrocyte-mediated neuroprotection and that the mechanism underlying this phenomenon is selective for complex I inhibitors and is not mediated by an oxidative stress response. The notion that dysfunctional astrocytes may contribute to the death of dopaminergic neurons in the SNpc has several consequences on how we view the pathogenesis of PD and possible treatments. Predominantly, these results say that even where there is a single gene cause of parkinsonism, we should consider models that involve more than just dopamine neurons. In fact, we should consider astrocytes and other non-neuronal cells as possible contributors to the early pathogenic mechanisms. A smaller, but also important point, is that when considering ways in which to treat PD we may need to consider whether the glial environment is supportive or not; treating only neurons may provide only part of the answer. Although the findings presented in this issue by the Hinkle group convincingly show a role for astrocytic DJ-1 in astrocyte-mediated neuroprotection, there are a number of areas that would merit further investigation. If DJ-1 is fundamentally an oxidative stress response protein, then is the neuroprotection also related to oxidative stress? The data presented by Mullet and Hinkle suggests that the effects are not due to, for example, antioxidant release from astrocytes, which may have been an obvious link. The authors propose that soluble factors are released from astrocytes, but these still need to be identified before more detailed mechanistic studies are undertaken. Furthermore, most of the current experiments use dissociated cultured cells. It is known that astrocytes express DJ-1 in vivo, and so it would seem important to determine whether astrocytic DJ-1 is important for neuroprotection in the intact brain. Overall, the data presented here identifies an exciting possibility, that neurodegeneration in a defined form of genetic parkinsonism may involve non-cell autonomous events. Identifying the underlying mechanisms is an important next step and can be facilitated by the types of models presented by Mullet and Hinkle.