Publications by authors named "Pamela J McLean"

80 Publications

Role of gut microbiota in regulating gastrointestinal dysfunction and motor symptoms in a mouse model of Parkinson's disease.

Gut Microbes 2021 01;13(1):1866974

Division of Gastroenterology and Hepatology, Mayo Clinic , Rochester, MN, USA.

Parkinson's disease (PD) is a common neurodegenerative disorder characterized primarily by motor and non-motor gastrointestinal (GI) deficits. GI symptoms' including compromised intestinal barrier function often accompanies altered gut microbiota composition and motor deficits in PD. Therefore, in this study, we set to investigate the role of gut microbiota and epithelial barrier dysfunction on motor symptom generation using a rotenone-induced mouse model of PD. We found that while six weeks of 10 mg/kg of chronic rotenone administration by oral gavage resulted in loss of tyrosine hydroxylase (TH) neurons in both germ-free (GF) and conventionally raised (CR) mice, the decrease in motor strength and coordination was observed only in CR mice. Chronic rotenone treatment did not disrupt intestinal permeability in GF mice but resulted in a significant change in gut microbiota composition and an increase in intestinal permeability in CR mice. These results highlight the potential role of gut microbiota in regulating barrier dysfunction and motor deficits in PD.
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http://dx.doi.org/10.1080/19490976.2020.1866974DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7833732PMC
January 2021

Detection of Extracellular Adenosine Triphosphate in a Mouse Model of Traumatic Brain Injury.

J Neurotrauma 2021 03 19;38(5):655-664. Epub 2020 Oct 19.

Department of Neuroscience, Mayo Clinic Graduate School of Biomedical Sciences, Mayo Clinic College of Medicine, Mayo Clinic, Jacksonville, Florida, USA.

Traumatic brain injury (TBI) is traditionally characterized by primary and secondary injury phases, both contributing to pathological and morphological changes. The mechanisms of damage and chronic consequences of TBI remain to be fully elucidated, but synaptic homeostasis disturbances and impaired energy metabolism are proposed to be a major contributor. It has been proposed that an increase of extracellular (eATP) adenosine triphosphate (ATP) in the area immediately surrounding impact may play a pivotal role in this sequence of events. After tissue injury, rupture of cell membranes allows release of intracellular ATP into the extracellular space, triggering a cascade of toxic events and inflammation. ATP is a ubiquitous messenger; however, simple and reliable techniques to measure its concentration have proven elusive. Here, we integrate a sensitive bioluminescent eATP sensor known as pmeLUC, with a controlled cortical impact mouse model to monitor eATP changes in a living animal after injury. Using the pmeLUC probe, a rapid increase of eATP is observed proximal to the point of impact within minutes of the injury. This event is significantly attenuated when animals are pretreated with an ATP hydrolyzing agent (apyrase) before surgery, confirming the contribution of eATP. This new eATP reporter could be useful for understanding the role of eATP in the pathogenesis in TBI and may identify a window of opportunity for therapeutic intervention.
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http://dx.doi.org/10.1089/neu.2020.7226DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7898407PMC
March 2021

Screening non-MAPT genes of the Chr17q21 H1 haplotype in Parkinson's disease.

Parkinsonism Relat Disord 2020 09 1;78:138-144. Epub 2020 Aug 1.

Department of Neuroscience, Mayo Clinic, Jacksonville, FL, 32224, USA; Department of Clinical Genomics, Mayo Clinic, Jacksonville, FL, 32224, USA; School of Medicine and Medical Science, University College Dublin, Dublin, Ireland; Neuroscience PhD Program, Mayo Clinic Graduate School of Biomedical Sciences, USA; Department of Biology, College of Arts and Sciences, University of North Florida, Jacksonville, FL, 32224, USA. Electronic address:

Introduction: The microtubule-associated protein tau (MAPT) gene is considered a strong genetic risk factor for Parkinson's disease (PD) in Caucasians. MAPT is located within an inversion region of high linkage disequilibrium designated as H1 and H2 haplotype, and contains eight other genes which have been implicated in neurodegeneration. The aim of the current study was to identify common coding variants in strong linkage disequilibrium (LD) within the associated loci on chr17q21 harboring MAPT.

Methods: Sanger sequencing of coding exons in 90 Caucasian late-onset PD (LOPD) patients was performed. Specific gene sequencing for LRRC37A, LRRC37A2, ARL17A and ARL17B was not possible given the high homology, presence of pseudogenes and copy number variants that are in the region, and therefore four genes (NSF, KANSL1, SPPL2C, and CRHR1) were included in the analysis. Coding variants from these four genes that did not perfectly tag (r = 1) the MAPT H1/H2 haplotype were genotyped in an independent replication series of Caucasian PD cases (N = 851) and controls (N = 730).

Results: In the 90 LOPD cases we identified 30 coding variants. Eleven non-synonymous variants tagged the MAPT H1/H2 haplotype, including two SPPL2C variants (rs12185233 and rs12373123) that had high pathogenic combined annotation dependent depletion (CADD) scores of >20. In the replication series, the non-synonymous KANSL1 rs17585974 variant was in very strong LD with MAPT H1/H2 and had a high CADD score of 24.7.

Conclusion: We have identified several non-synonymous variants across neighboring genes of MAPT that may warrant further genetic and functional investigation within the biological etiology of PD.
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http://dx.doi.org/10.1016/j.parkreldis.2020.07.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7686230PMC
September 2020

APOE4 exacerbates α-synuclein pathology and related toxicity independent of amyloid.

Sci Transl Med 2020 02;12(529)

Department of Neuroscience, Mayo Clinic, Jacksonville, FL 32224, USA.

The apolipoprotein E () ε4 allele is the strongest genetic risk factor for late-onset Alzheimer's disease mainly by driving amyloid-β pathology. Recently, has also been found to be a genetic risk factor for Lewy body dementia (LBD), which includes dementia with Lewy bodies and Parkinson's disease dementia. How drives risk of LBD and whether it has a direct effect on α-synuclein pathology are not clear. Here, we generated a mouse model of synucleinopathy using an adeno-associated virus gene delivery of α-synuclein in human APOE-targeted replacement mice expressing APOE2, APOE3, or APOE4. We found that APOE4, but not APOE2 or APOE3, increased α-synuclein pathology, impaired behavioral performances, worsened neuronal and synaptic loss, and increased astrogliosis at 9 months of age. Transcriptomic profiling in APOE4-expressing α-synuclein mice highlighted altered lipid and energy metabolism and synapse-related pathways. We also observed an effect of on α-synuclein pathology in human postmortem brains with LBD and minimal amyloid pathology. Our data demonstrate a pathogenic role of APOE4 in exacerbating α-synuclein pathology independent of amyloid, providing mechanistic insights into how increases the risk of LBD.
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http://dx.doi.org/10.1126/scitranslmed.aay1809DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8309690PMC
February 2020

Alpha-synuclein-induced mitochondrial dysfunction is mediated via a sirtuin 3-dependent pathway.

Mol Neurodegener 2020 01 13;15(1). Epub 2020 Jan 13.

Department of Neuroscience, Mayo Clinic, 4500 San Pablo Road, Jacksonville, FL, 32224, USA.

Background: Misfolding and aggregation of the presynaptic protein alpha-synuclein (αsyn) is a hallmark of Parkinson's disease (PD) and related synucleinopathies. Although predominantly localized in the cytosol, a body of evidence has shown that αsyn localizes to mitochondria and contributes to the disruption of key mitochondrial processes. Mitochondrial dysfunction is central to the progression of PD and mutations in mitochondrial-associated proteins are found in familial cases of PD. The sirtuins are highly conserved nicotinamide adenine dinucleotide (NAD)-dependent enzymes that play a broad role in cellular metabolism and aging. Interestingly, mitochondrial sirtuin 3 (SIRT3) plays a major role in maintaining mitochondrial function and preventing oxidative stress, and is downregulated in aging and age-associated diseases such as neurodegenerative disorders. Herein, we hypothesize that αsyn is associated with decreased SIRT3 levels contributing to impaired mitochondrial dynamics and biogenesis in PD.

Methods: The level of mitochondrial SIRT3 was assessed in cells expressing oligomeric αsyn within the cytosolic and mitochondrial-enriched fractions. Mitochondrial integrity, respiration, and health were examined using several markers of mitochondrial dynamics and stress response and by measuring the rate of oxygen consumption (OCR). Our findings were validated in a rodent model of PD as well as in human post-mortem Lewy body disease (LBD) brain tissue.

Results: Here, we demonstrate that αsyn associates with mitochondria and induces a decrease in mitochondrial SIRT3 levels and mitochondrial biogenesis. We show that SIRT3 downregulation is accompanied by decreased phosphorylation of AMPK and cAMP-response element binding protein (CREB), as well as increased phosphorylation of dynamin-related protein 1 (DRP1), indicative of impaired mitochondrial dynamics. OCR was significantly decreased suggesting a mitochondria respiratory deficit. Interestingly treatment with AMPK agonist 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside (AICAR) restores SIRT3 expression, improves mitochondrial function, and decreases αsyn oligomer formation in a SIRT3-dependent manner.

Conclusions: Together, our findings suggest that pharmacologically increasing SIRT3 levels can counteract αsyn-induced mitochondrial dysfunction by reducing αsyn oligomers and normalizing mitochondrial bioenergetics. These data support a protective role for SIRT3 in PD-associated pathways and contribute significant mechanistic insight into the interplay of SIRT3 and αsyn.
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http://dx.doi.org/10.1186/s13024-019-0349-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6956494PMC
January 2020

In Vivo Protein Complementation Demonstrates Presynaptic α-Synuclein Oligomerization and Age-Dependent Accumulation of 8-16-mer Oligomer Species.

Cell Rep 2019 Nov;29(9):2862-2874.e9

Department of Neurology, Ulm University, Ulm, Germany. Electronic address:

Intracellular accumulation of α-synuclein (α-syn) and formation of Lewy bodies are neuropathological characteristics of Parkinson's disease (PD) and related α-synucleinopathies. Oligomerization and spreading of α-syn from neuron to neuron have been suggested as key events contributing to the progression of PD. To directly visualize and characterize α-syn oligomerization and spreading in vivo, we generated two independent conditional transgenic mouse models based on α-syn protein complementation assays using neuron-specifically expressed split Gaussia luciferase or split Venus yellow fluorescent protein (YFP). These transgenic mice allow direct assessment of the quantity and subcellular distribution of α-syn oligomers in vivo. Using these mouse models, we demonstrate an age-dependent accumulation of a specific subtype of α-syn oligomers. We provide in vivo evidence that, although α-syn is found throughout neurons, α-syn oligomerization takes place at the presynapse. Furthermore, our mouse models provide strong evidence for a transsynaptic cell-to-cell transfer of de novo generated α-syn oligomers in vivo.
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http://dx.doi.org/10.1016/j.celrep.2019.10.089DOI Listing
November 2019

Increased Immune Activation by Pathologic α-Synuclein in Parkinson's Disease.

Ann Neurol 2019 10 15;86(4):593-606. Epub 2019 Aug 15.

Department of Neurology, Ulm University, Ulm, Germany.

Objective: Excessive inflammation in the central nervous system (CNS) and the periphery can result in neurodegeneration and parkinsonism. Recent evidence suggests that immune responses in Parkinson disease patients are dysregulated, leading to an increased inflammatory reaction to unspecific triggers. Although α-synuclein pathology is the hallmark of Parkinson disease, it has not been investigated whether pathologic α-synuclein is a specific trigger for excessive inflammatory responses in Parkinson disease.

Methods: We investigated the immune response of primary human monocytes and a microglial cell line to pathologic forms of α-synuclein by assessing cytokine release upon exposure.

Results: We show that pathologic α-synuclein (mutations, aggregation) results in a robust inflammatory activation of human monocytes and microglial BV2 cells. The activation is conformation- dependent, with increasing fibrillation and early onset mutations having the strongest effect on immune activation. We also found that activation of immune cells by extracellular α-synuclein is potentiated by extracellular vesicles, possibly by facilitating the uptake of α-synuclein. Blood extracellular vesicles from Parkinson disease patients induce a stronger activation of monocytes than blood extracellular vesicles from healthy controls. Most importantly, monocytes from Parkinson disease patients are dysregulated and hyperactive in response to stimulation with pathologic α-synuclein. Furthermore, we demonstrate that α-synuclein pathology in the CNS is sufficient to induce the monocyte dysregulation in the periphery of a mouse model.

Interpretation: Taken together, our data suggest that α-synuclein pathology and dysregulation of monocytes in Parkinson disease can act together to induce excessive inflammatory responses to α-synuclein. ANN NEUROL 2019;86:593-606.
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http://dx.doi.org/10.1002/ana.25557DOI Listing
October 2019

Cellular models of alpha-synuclein toxicity and aggregation.

J Neurochem 2019 09 30;150(5):566-576. Epub 2019 Jul 30.

Department of Neuroscience, Mayo Clinic, Jacksonville, Florida, USA.

Misfolding and aggregation of alpha-synuclein (α-synuclein) with concomitant cytotoxicity is a hallmark of Lewy body related disorders such as Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy. Although it plays a pivotal role in pathogenesis and disease progression, the function of α-synuclein and the molecular mechanisms underlying α-synuclein-induced neurotoxicity in these diseases are still elusive. Many in vitro and in vivo experimental models mimicking α-synuclein pathology such as oligomerization, toxicity and more recently neuronal propagation have been generated over the years. In particular, cellular models have been crucial for our comprehension of the pathogenic process of the disease and are beneficial for screening of molecules capable of modulating α-synuclein toxicity. Here, we review α-synuclein based cell culture models that reproduce some features of the neuronal populations affected in patients, from basic unicellular organisms to mammalian cell lines and primary neurons, to the cutting edge models of patient-specific cell lines. These reprogrammed cells known as induced pluripotent stem cells (iPSCs) have garnered attention because they closely reproduce the characteristics of neurons found in patients and provide a valuable tool for mechanistic studies. We also discuss how different cell models may constitute powerful tools for high-throughput screening of molecules capable of modulating α-synuclein toxicity and prevention of its propagation. This article is part of the Special Issue "Synuclein".
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http://dx.doi.org/10.1111/jnc.14806DOI Listing
September 2019

CRISPR/Cas9 editing of APP C-terminus attenuates β-cleavage and promotes α-cleavage.

Nat Commun 2019 01 3;10(1):53. Epub 2019 Jan 3.

Department of Pathology and Laboratory Medicine, University of Wisconsin-Madison, 1111 Highland Avenue, Madison, WI, 53705, USA.

CRISPR/Cas9 guided gene-editing is a potential therapeutic tool, however application to neurodegenerative disease models has been limited. Moreover, conventional mutation correction by gene-editing would only be relevant for the small fraction of neurodegenerative cases that are inherited. Here we introduce a CRISPR/Cas9-based strategy in cell and animal models to edit endogenous amyloid precursor protein (APP) at the extreme C-terminus and reciprocally manipulate the amyloid pathway, attenuating APP-β-cleavage and Aβ production, while up-regulating neuroprotective APP-α-cleavage. APP N-terminus and compensatory APP-homologues remain intact, with no apparent effects on neurophysiology in vitro. Robust APP-editing is seen in human iPSC-derived neurons and mouse brains with no detectable off-target effects. Our strategy likely works by limiting APP and BACE-1 approximation, and we also delineate mechanistic events that abrogates APP/BACE-1 convergence in this setting. Our work offers conceptual proof for a selective APP silencing strategy.
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http://dx.doi.org/10.1038/s41467-018-07971-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6318289PMC
January 2019

14-3-3 Proteins Reduce Cell-to-Cell Transfer and Propagation of Pathogenic α-Synuclein.

J Neurosci 2018 09 9;38(38):8211-8232. Epub 2018 Aug 9.

Center for Neurodegeneration and Experimental Therapeutics, Department of Neurology,

α-Synuclein (αsyn) is the key protein that forms neuronal aggregates in the neurodegenerative disorders Parkinson's disease (PD) and dementia with Lewy bodies. Recent evidence points to the prion-like spread of αsyn from one brain region to another. Propagation of αsyn is likely dependent on release, uptake, and misfolding. Under normal circumstances, this highly expressed brain protein functions normally without promoting pathology, yet the underlying endogenous mechanisms that prevent αsyn spread are not understood. 14-3-3 proteins are highly expressed brain proteins that have chaperone function and regulate protein trafficking. In this study, we investigated the potential role of the 14-3-3 proteins in the regulation of αsyn spread using two models of αsyn spread. In a paracrine αsyn model, 14-3-3θ promoted release of αsyn complexed with 14-3-3θ. Despite higher amounts of released αsyn, extracellular αsyn showed reduced oligomerization and seeding capability, reduced internalization, and reduced toxicity in primary mixed-gender mouse neurons. 14-3-3 inhibition reduced the amount of αsyn released, yet released αsyn was more toxic and demonstrated increased oligomerization, seeding capability, and internalization. In the preformed fibril model, 14-3-3 θ reduced αsyn aggregation and neuronal death, whereas 14-3-3 inhibition enhanced αsyn aggregation and neuronal death in primary mouse neurons. 14-3-3s blocked αsyn spread to distal chamber neurons not exposed directly to fibrils in multichamber, microfluidic devices. These findings point to 14-3-3s as a direct regulator of αsyn propagation, and suggest that dysfunction of 14-3-3 function may promote αsyn pathology in PD and related synucleinopathies. Transfer of misfolded aggregates of α-synuclein from one brain region to another is implicated in the pathogenesis of Parkinson's disease and other synucleinopathies. This process is dependent on active release, internalization, and misfolding of α-synuclein. 14-3-3 proteins are highly expressed chaperone proteins that interact with α-synuclein and regulate protein trafficking. We used two different models in which toxicity is associated with cell-to-cell transfer of α-synuclein to test whether 14-3-3s impact α-synuclein toxicity. We demonstrate that 14-3-3θ reduces α-synuclein transfer and toxicity by inhibiting oligomerization, seeding capability, and internalization of α-synuclein, whereas 14-3-3 inhibition accelerates the transfer and toxicity of α-synuclein in these models. Dysfunction of 14-3-3 function may be a critical mechanism by which α-synuclein propagation occurs in disease.
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http://dx.doi.org/10.1523/JNEUROSCI.1134-18.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6146494PMC
September 2018

Bimolecular Fluorescence Complementation of Alpha-synuclein Demonstrates its Oligomerization with Dopaminergic Phenotype in Mice.

EBioMedicine 2018 Mar 31;29:13-22. Epub 2018 Jan 31.

MassGeneral Institute for Neurodegenerative Disease, Department of Neurology, Massachusetts General Hospital, Harvard Medical School, Boston, USA; Shanghai Huashan Hospital, Fudan University, Shanghai, China. Electronic address:

Alpha-synuclein (αSyn) is encoded by the first causal gene identified in Parkinson's disease (PD) and is the main component of Lewy bodies, a pathological hallmark of PD. aSyn-based animal models have contributed to our understanding of PD pathophysiology and to the development of therapeutics. Overexpression of human wildtype αSyn by viral vectors in rodents recapitulates the loss of dopaminergic neurons from the substantia nigra, another defining pathological feature of the disease. The development of a rat model exhibiting bimolecular fluorescence complementation (BiFC) of αSyn by recombinant adeno-associated virus facilitates detection of the toxic αSyn oligomers species. We report here neurochemical, neuropathological and behavioral characterization of BiFC of αSyn in mice. Overexpression and oligomerization of αSyn through BiFC is detected by conjugated fluorescence. Reduced striatal dopamine and loss of nigral dopaminergic neurons are accompanied neuroinflammation and abnormal motor activities. Our mouse model may provide a valuable tool to study the role of αSyn in PD and to explore therapeutic approaches.
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http://dx.doi.org/10.1016/j.ebiom.2018.01.035DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5925445PMC
March 2018

Impaired endo-lysosomal membrane integrity accelerates the seeding progression of α-synuclein aggregates.

Sci Rep 2017 08 9;7(1):7690. Epub 2017 Aug 9.

Department of Neuroscience, Mayo Clinic, Jacksonville, FL, 32224, USA.

In neurodegenerative diseases, seeding is a process initiated by the internalization of exogenous protein aggregates. Multiple pathways for internalization of aggregates have been proposed, including direct membrane penetration and endocytosis. To decipher the seeding mechanisms of alpha-synuclein (αS) aggregates in human cells, we visualized αS aggregation, endo-lysosome distribution, and endo-lysosome rupture in real-time. Our data suggest that exogenous αS can seed endogenous cytoplasmic αS by either directly penetrating the plasma membrane or via endocytosis-mediated endo-lysosome rupture, leading to formation of endo-lysosome-free or endo-lysosome-associated αS aggregates, respectively. Further, we demonstrate that αS aggregates isolated from postmortem human brains with diffuse Lewy body disease (DLBD) preferentially show endocytosis-mediated seeding associated with endo-lysosome rupture and have significantly reduced seeding activity compared to recombinant αS aggregates. Colocalization of αS pathology with galectin-3 (a marker of endo-lysosomal membrane rupture) in the basal forebrain of DLBD, but not in age-matched controls, suggests endo-lysosome rupture is involved in the formation of αS pathology in humans. Interestingly, cells with endo-lysosomal membrane permeabilization (LMP) are more vulnerable to the seeding effects of αS aggregates. This study suggests that endo-lysosomal impairment in neurons might play an important role in PD progression.
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http://dx.doi.org/10.1038/s41598-017-08149-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5550496PMC
August 2017

The Golgi-localized, gamma ear-containing, ARF-binding (GGA) protein family alters alpha synuclein (α-syn) oligomerization and secretion.

Aging (Albany NY) 2017 07;9(7):1677-1697

Department of Neurology, Ulm University, Ulm 89081, Germany.

Several age-related neurodegenerative disorders are associated with protein misfolding and aggregation of toxic peptides. α-synuclein (α-syn) aggregation and the resulting cytotoxicity is a hallmark of Parkinson's disease (PD) as well as dementia with Lewy bodies. Rising evidence points to oligomeric and pre-fibrillar forms as the pathogenic species, and oligomer secretion seems to be crucial for the spreading and progression of PD pathology. Recent studies implicate that dysfunctions in endolysosomal/autophagosomal pathways increase α-syn secretion. Mutation in the retromer-complex protein VPS35, which is involved in endosome to Golgi transport, was suggested to cause familial PD. GGA proteins regulate vesicular traffic between Golgi and endosomes and might work as antagonists for retromer complex mediated transport. To investigate the role of the GGAs in the α-syn oligomerization and/or secretion process we utilized protein-fragment complementation assays (PCA). We here demonstrate that GGAs alter α-syn oligomer secretion and α-syn oligomer-mediated toxicity. Specifically, we determined that GGA3 modifies extracellular α-syn species in an exosome-independent manner. Our data suggest that GGA3 drives α-syn oligomerization in endosomal compartments and thus facilitates α-syn oligomer secretion. Preventing the early events in α-syn oligomer release may be a novel approach to halt disease spreading in PD and other synucleinopathies.
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http://dx.doi.org/10.18632/aging.101261DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5559169PMC
July 2017

Neonatal AAV delivery of alpha-synuclein induces pathology in the adult mouse brain.

Acta Neuropathol Commun 2017 06 23;5(1):51. Epub 2017 Jun 23.

Department of Neuroscience, Mayo Clinic, 4500 San Pablo Rd, Jacksonville, FL, 32224, USA.

Abnormal accumulation of alpha-synuclein (αsyn) is a pathological hallmark of Lewy body related disorders such as Parkinson's disease and Dementia with Lewy body disease. During the past two decades, a myriad of animal models have been developed to mimic pathological features of synucleinopathies by over-expressing human αsyn. Although different strategies have been used, most models have little or no reliable and predictive phenotype. Novel animal models are a valuable tool for understanding neuronal pathology and to facilitate development of new therapeutics for these diseases. Here, we report the development and characterization of a novel model in which mice rapidly express wild-type αsyn via somatic brain transgenesis mediated by adeno-associated virus (AAV). At 1, 3, and 6 months of age following intracerebroventricular (ICV) injection, mice were subjected to a battery of behavioral tests followed by pathological analyses of the brains. Remarkably, significant levels of αsyn expression are detected throughout the brain as early as 1 month old, including olfactory bulb, hippocampus, thalamic regions and midbrain. Immunostaining with a phospho-αsyn (pS129) specific antibody reveals abundant pS129 expression in specific regions. Also, pathologic αsyn is detected using the disease specific antibody 5G4. However, this model did not recapitulate behavioral phenotypes characteristic of rodent models of synucleinopathies. In fact no deficits in motor function or cognition were observed at 3 or 6 months of age. Taken together, these findings show that transduction of neonatal mouse with AAV-αsyn can successfully lead to rapid, whole brain transduction of wild-type human αsyn, but increased levels of wildtype αsyn do not induce behavior changes at an early time point (6 months), despite pathological changes in several neurons populations as early as 1 month.
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http://dx.doi.org/10.1186/s40478-017-0455-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5481919PMC
June 2017

Investigation of Endocytic Pathways for the Internalization of Exosome-Associated Oligomeric Alpha-Synuclein.

Front Neurosci 2017 30;11:172. Epub 2017 Mar 30.

Department of Neuroscience, Mayo ClinicJacksonville, FL, USA.

Misfolding and aggregation of alpha-synuclein (αsyn) resulting in cytotoxicity is a hallmark of Parkinson's disease (PD) and related synucleinopathies. The recent body of evidence indicates that αsyn can be released from neuronal cells by nonconventional exocytosis involving extracellular vesicles (EVs) such as exosomes. The transfer of αsyn between cells has been proposed to be an important mechanism of disease propagation in PD. To date, exosome trafficking mechanisms, including release and cell-cell transmission, have not been fully described. To gain insight into the mechanisms involved, exosomes were purified from conditioned media of stable cells secreting αsyn oligomers. A novel bimolecular protein complementation assay was used to detect exosomes containing αsyn oligomers. Recipient cells were treated with exosomes containing αsyn oligomers or "free" non-exosome-associated αsyn oligomers and internalization was monitored. We demonstrate that cell-derived exosome-associated αsyn oligomers can be efficiently internalized by recipient cells. Interestingly exosome-free αsyn oligomers isolated from conditioned medium were not internalized but remained bound to the extracellular surface. To investigate the endocytic pathway(s) required for the exosome uptake different pharmacological inhibitors of caveolin-dependent, clathrin-dependent, and macropinocytosis pathways were utilized. Surprisingly, none of these pathways appear to play a significant role in the internalization of exosome-associated αsyn oligomers. Finally, the role of heparin sulfate proteoglycans (HSPGs) in exosome-associated αsyn internalization was investigated using genetic approach. Despite previous studies showing HSPGs can modulate internalization of fibrillar αsyn, genetic manipulations did not attenuate internalization of exosome-associated αsyn oligomers in our hands, suggesting that exosome-associated αsyn is internalized via an alternative endocytic pathway(s) that has yet to be elucidated.
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http://dx.doi.org/10.3389/fnins.2017.00172DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5371652PMC
March 2017

Histones facilitate α-synuclein aggregation during neuronal apoptosis.

Acta Neuropathol 2017 04 21;133(4):547-558. Epub 2016 Dec 21.

Department of Neuroscience, Mayo Clinic, 4500 San Pablo Road, Jacksonville, FL, 32224, USA.

Ample in vitro and in vivo experimental evidence supports the hypothesis that intercellular transmission of α-synuclein (αS) is a mechanism underlying the spread of αS pathology in Parkinson's disease and related disorders. What remains unexplained is where and how initial transmissible αS aggregates form. In a previous study, we demonstrated that αS aggregates rapidly form in neurons with impaired nuclear membrane integrity due to the interaction between nuclear proaggregant factor(s) and αS and that such aggregates may serve as a source for αS seeding. In the present study, we identify histones as a potential nuclear proaggregant factor for αS aggregation in both apoptotic neurons and brains with αS pathology. We further demonstrate that histone-induced aggregates contain a range of αS oligomers, including protofibrils and mature fibrils, and that these αS aggregates can seed additional aggregation. Importantly, we demonstrate transmissibility in mouse brains from stereotaxic injection. This study provides new clues to the mechanism underlying initial pathological aggregation of αS in PD and related disorders, and could lead to novel diagnostic and therapeutic approaches.
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http://dx.doi.org/10.1007/s00401-016-1660-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5350017PMC
April 2017

Commentary: alpha-synuclein interacts with SOD1 and promotes its oligomerization.

J Neurol Neuromedicine 2016 ;1(7):28-30

Department of Neurology, Ulm University, Albert-Einstein-Allee 11, 89081 Ulm, Germany.

Alpha-synuclein and Cu, Zn superoxide dismutase (SOD1) are both aggregation-prone proteins that are associated with Parkinson's disease (PD) and amyotrophic lateral sclerosis (ALS), respectively. Recently, we showed that alpha-synuclein interacts with SOD1 in various cell types and tissues. Using a cell culture model, we also found that alpha-synuclein nucleates the polymerization of SOD1. Here, we discuss the current literature regarding their interaction and their co-localization in aggregates of human tissue. Furthermore we comment on the reported alpha-synuclein-induced SOD1 polymerization in terms of cross-seeding effects in neurodegeneration.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5108581PMC
http://dx.doi.org/10.29245/2572.942x/2016/7.1065DOI Listing
January 2016

The neural chaperone proSAAS blocks α-synuclein fibrillation and neurotoxicity.

Proc Natl Acad Sci U S A 2016 08 25;113(32):E4708-15. Epub 2016 Jul 25.

School of Medicine, University of Maryland, Baltimore, MD 21201;

Emerging evidence strongly suggests that chaperone proteins are cytoprotective in neurodegenerative proteinopathies involving protein aggregation; for example, in the accumulation of aggregated α-synuclein into the Lewy bodies present in Parkinson's disease. Of the various chaperones known to be associated with neurodegenerative disease, the small secretory chaperone known as proSAAS (named after four residues in the amino terminal region) has many attractive properties. We show here that proSAAS, widely expressed in neurons throughout the brain, is associated with aggregated synuclein deposits in the substantia nigra of patients with Parkinson's disease. Recombinant proSAAS potently inhibits the fibrillation of α-synuclein in an in vitro assay; residues 158-180, containing a largely conserved element, are critical to this bioactivity. ProSAAS also exhibits a neuroprotective function; proSAAS-encoding lentivirus blocks α-synuclein-induced cytotoxicity in primary cultures of nigral dopaminergic neurons, and recombinant proSAAS blocks α-synuclein-induced cytotoxicity in SH-SY5Y cells. Four independent proteomics studies have previously identified proSAAS as a potential cerebrospinal fluid biomarker in various neurodegenerative diseases. Coupled with prior work showing that proSAAS blocks β-amyloid aggregation into fibrils, this study supports the idea that neuronal proSAAS plays an important role in proteostatic processes. ProSAAS thus represents a possible therapeutic target in neurodegenerative disease.
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http://dx.doi.org/10.1073/pnas.1601091113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4987805PMC
August 2016

Intracellular formation of α-synuclein oligomers and the effect of heat shock protein 70 characterized by confocal single particle spectroscopy.

Biochem Biophys Res Commun 2016 08 7;477(1):76-82. Epub 2016 Jun 7.

Center for Neuropathology and Prion Research, Ludwig-Maximilians-University, Feodor-Lynen-Str. 23, 81377 Munich, Germany. Electronic address:

Synucleinopathies such as dementia with Lewy bodies or Parkinson's disease are characterized by intracellular deposition of pathologically aggregated α-synuclein. The details of the molecular pathogenesis of PD and especially the conditions that lead to intracellular aggregation of α-synuclein and the role of these aggregates in cell death remain unknown. In cell free in vitro systems considerable knowledge about the aggregation processes has been gathered. In comparison, the knowledge about these aggregation processes in cells is far behind. In cells α-synuclein aggregates can be toxic. However, the crucial particle species responsible for decisive steps in pathogenesis such as seeding a continuing aggregation process and triggering cell death remain to be identified. In order to understand the complex nature of intracellular α-synuclein aggregate formation, we analyzed fluorescent particles formed by venus and α-synuclein-venus fusion proteins and α-synuclein-hemi-venus fusion proteins derived from gently lyzed cells. With these techniques we were able to identify and characterize α-synuclein oligomers formed in cells. Especially the use of α-synuclein-hemi-venus fusion proteins enabled us to identify very small α-synuclein oligomers with high sensitivity. Furthermore, we were able to study the molecular effect of heat shock protein 70, which is known to inhibit α-synuclein aggregation in cells. Heat shock protein 70 does not only influence the size of α-synuclein oligomers, but also their quantity. In summary, this approach based on fluorescence single particle spectroscopy, that is suited for high throughput measurements, can be used to detect and characterize intracellularly formed α-synuclein aggregates and characterize the effect of molecules that interfere with α-synuclein aggregate formation.
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http://dx.doi.org/10.1016/j.bbrc.2016.06.023DOI Listing
August 2016

Proaggregant nuclear factor(s) trigger rapid formation of α-synuclein aggregates in apoptotic neurons.

Acta Neuropathol 2016 07 2;132(1):77-91. Epub 2016 Feb 2.

Neuropathology Laboratory, Department of Neuroscience, Mayo Clinic, 4500 San Pablo Road, Jacksonville, FL, 32224, USA.

Cell-to-cell transmission of α-synuclein (αS) aggregates has been proposed to be responsible for progressive αS pathology in Parkinson disease (PD) and related disorders, including dementia with Lewy bodies. In support of this concept, a growing body of in vitro and in vivo experimental evidence shows that exogenously introduced αS aggregates can spread into surrounding cells and trigger PD-like pathology. It remains to be determined what factor(s) lead to initiation of αS aggregation that is capable of seeding subsequent propagation. In this study we demonstrate that filamentous αS aggregates form in neurons in response to apoptosis induced by staurosporine or other toxins-6-hydroxy-dopamine and 1-methyl-4-phenylpyridinium (MPP+). Interaction between αS and proaggregant nuclear factor(s) is associated with disruption of nuclear envelope integrity. Knocking down a key nuclear envelop constituent protein, lamin B1, enhances αS aggregation. Moreover, in vitro and in vivo experimental models demonstrate that aggregates released upon cell breakdown can be taken up by surrounding cells. Accordingly, we suggest that at least some αS aggregation might be related to neuronal apoptosis or loss of nuclear membrane integrity, exposing cytosolic α-synuclein to proaggregant nuclear factors. These findings provide new clues to the pathogenesis of PD and related disorders that can lead to novel treatments of these disorders. Specifically, finding ways to limit the effects of apoptosis on αS aggregation, deposition, local uptake and subsequent propagation might significantly impact progression of disease.
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http://dx.doi.org/10.1007/s00401-016-1542-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4911378PMC
July 2016

A Rapid, Semi-Quantitative Assay to Screen for Modulators of Alpha-Synuclein Oligomerization Ex vivo.

Front Neurosci 2015 19;9:511. Epub 2016 Jan 19.

Department of Neuroscience, Mayo ClinicJacksonville, FL, USA; Mayo Graduate School, Mayo ClinicJacksonville, FL, USA.

Alpha synuclein (αsyn) aggregates are associated with the pathogenesis of Parkinson's disease and others related disorders. Although modulation of αsyn aggregation is an attractive therapeutic target, new powerful methodologies are desperately needed to facilitate in vivo screening of novel therapeutics. Here, we describe an in vivo rodent model with the unique ability to rapidly track αsyn-αsyn interactions and thus oligomerization using a bioluminescent protein complementation strategy that monitors spatial and temporal αsyn oligomerization ex vivo. We find that αsyn forms oligomers in vivo as early as 1 week after stereotactic AAV injection into rat substantia nigra. Strikingly, although abundant αsyn expression is also detected in striatum at 1 week, no αsyn oligomers are detected at this time point. By 4 weeks, oligomerization of αsyn is detected in both striatum and substantia nigra homogenates. Moreover, in a proof-of-principle experiment, the effect of a previously described Hsp90 inhibitor known to prevent αsyn oligomer formation, demonstrates the utility of this rapid and sensitive animal model to monitor αsyn oligomerization status in the rat brain.
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http://dx.doi.org/10.3389/fnins.2015.00511DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4717311PMC
February 2016

α-synuclein interacts with SOD1 and promotes its oligomerization.

Mol Neurodegener 2015 Dec 8;10:66. Epub 2015 Dec 8.

Department of Neurology, Ulm University, Albert-Einstein-Allee 11, 89081, Ulm, Germany.

Background: Parkinson's disease (PD) and amyotrophic lateral sclerosis (ALS) are both neurodegenerative diseases leading to impaired execution of movement. α-Synuclein plays a central role in the pathogenesis of PD whereas Cu, Zn superoxide dismutase (SOD1) is a key player in a subset of familial ALS cases. Under pathological conditions both α-synuclein and SOD1 form oligomers and fibrils. In this study we investigated the possible molecular interaction of α-synuclein and SOD1 and its functional and pathological relevance.

Results: Using a protein-fragment complementation approach and co-IP, we found that α-synuclein and SOD1 physically interact in living cells, human erythrocytes and mouse brain tissue. Additionally, our data show that disease related mutations in α-synuclein (A30P, A53T) and SOD1 (G85R, G93A) modify the binding of α-synuclein to SOD1. Notably, α-synuclein accelerates SOD1 oligomerization independent of SOD1 activity.

Conclusion: This study provides evidence for a novel interaction of α-synuclein and SOD1 that might be relevant for neurodegenerative diseases.
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http://dx.doi.org/10.1186/s13024-015-0062-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4672499PMC
December 2015

Transmission of Soluble and Insoluble α-Synuclein to Mice.

J Neuropathol Exp Neurol 2015 Dec;74(12):1158-69

From the Department of Neuroscience (DRJ, MD, ATB, MD, MEM, DWD, PJM) and Mayo Graduate School (PJM), Mayo Clinic, Jacksonville, Florida.

The neurodegenerative synucleinopathies, which include Parkinson disease, multiple-system atrophy, and Lewy body disease, are characterized by the presence of abundant neuronal inclusions called Lewy bodies and Lewy neurites. These disorders remain incurable, and a greater understanding of the pathologic processes is needed for effective treatment strategies to be developed. Recent data suggest that pathogenic misfolding of the presynaptic protein, α-synuclein (α-syn), and subsequent aggregation and accumulation are fundamental to the disease process. It is hypothesized that the misfolded isoform is able to induce misfolding of normal endogenous α-syn, much like what occurs in the prion diseases. Recent work highlighting the seeding effect of pathogenic α-syn has largely focused on the detergent-insoluble species of the protein. In this study, we performed intracerebral inoculations of the sarkosyl-insoluble or sarkosyl-soluble fractions of human Lewy body disease brain homogenate and show that both fractions induce CNS pathology in mice at 4 months after injection. Disease-associated deposits accumulated both near and distal to the site of the injection, suggesting a cell-to-cell spread via recruitment of α-syn. These results provide further insight into the prion-like mechanisms of α-syn and suggest that disease-associated α-syn is not homogeneous within a single patient but might exist in both soluble and insoluble isoforms.
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http://dx.doi.org/10.1097/NEN.0000000000000262DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4654695PMC
December 2015

Untangling a role for tau in synucleinopathies.

Biol Psychiatry 2015 Nov;78(10):666-7

Department of Neuroscience, Mayo Clinic, Jacksonville, Florida. Electronic address:

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http://dx.doi.org/10.1016/j.biopsych.2015.08.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5127588PMC
November 2015

Chaperones in Neurodegeneration.

J Neurosci 2015 Oct;35(41):13853-9

Mayo Clinic, Jacksonville, Florida 32224.

Unlabelled: Cellular protein homeostasis (proteostasis) maintains the integrity of the proteome and includes protein synthesis, folding, oligomerization, and turnover; chaperone proteins assist with all of these processes. Neurons appear to be especially susceptible to failures in proteostasis, and this is now increasingly recognized as a major origin of neurodegenerative disease. This review, based on a mini-symposium presented at the 2015 Society for Neuroscience meeting, describes new work in the area of neuronal proteostasis, with a specific focus on the roles and therapeutic uses of protein chaperones. We first present a brief review of protein misfolding and aggregation in neurodegenerative disease. We then discuss different aspects of chaperone control of neuronal proteostasis on topics ranging from chaperone engineering, to chaperone-mediated blockade of protein oligomerization and cytotoxicity, to the potential rescue of neurodegenerative processes using modified chaperone proteins.

Significance Statement: Aberrant protein homeostasis within neurons results in protein misfolding and aggregation. In this review, we discuss specific roles for protein chaperones in the oligomerization, assembly, and disaggregation of proteins known to be abnormally folded in neurodegenerative disease. Collectively, our goal is to identify therapeutic mechanisms to reduce the cellular toxicity of abnormal aggregates.
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http://dx.doi.org/10.1523/JNEUROSCI.2600-15.2015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4604223PMC
October 2015

Biomarkers in Parkinson's disease: Advances and strategies.

Parkinsonism Relat Disord 2016 Jan 30;22 Suppl 1:S106-10. Epub 2015 Sep 30.

Department of Neurology, Mayo Clinic, Jacksonville, FL 32224, USA. Electronic address:

Parkinson's disease (PD) is a neurodegenerative disorder characterized by progressive motor disturbances and affects more than 1% of the worldwide population. Despite considerable progress in understanding PD pathophysiology, including genetic and biochemical causes, diagnostic approaches lack accuracy and interventions are restricted to symptomatic treatments. PD is a complex syndrome with different clinical subtypes and a wide variability in disorder course. In order to deliver better clinical management of PD patients and discovery of novel therapies, there is an urgent need to find sensitive, specific, and reliable biomarkers. The development of biomarkers will not only help the scientific community to identify populations at risk, but also facilitate clinical diagnosis. Furthermore, these tools could monitor progression, which could ultimately deliver personalized therapeutic strategies. The field of biomarker discovery in PD has attracted significant attention and there have been numerous contributions in recent years. Although none of the parameters have been validated for clinical practice, some candidates hold promise. This review summarizes recent advances in the development of PD biomarkers and discusses new strategies for their utilization.
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http://dx.doi.org/10.1016/j.parkreldis.2015.09.048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5120398PMC
January 2016

Role for the microtubule-associated protein tau variant p.A152T in risk of α-synucleinopathies.

Neurology 2015 Nov 2;85(19):1680-6. Epub 2015 Sep 2.

From the Departments of Neuroscience (C.L., K.O., O.L.-B., A.I.S.-O., R.L.W., S.R., M.E.M., P.J.M., R.R., N.E.-T., D.W.D., O.A.R.), Neurology (S.F., N.E.-T., N.R.G.-R., R.J.U., Z.K.W.), and Psychiatry and Psychology (T.J.F.), Division of Biomedical Statistics and Informatics (M.G.H.), and Mayo Graduate School (P.J.M., O.A.R.), Mayo Clinic, Jacksonville, FL; Dublin Neurological Institute at the Mater Misericordiae University Hospital (A.M., T.L.), Conway; Institute of Biomolecular & Biomedical Research (A.M., T.L.), University College Dublin, Ireland; Department of Clinical Sciences (A.P.), Lund University, and Department of Neurology, Skåne University Hospital, Sweden; Department of Neurology (J.S., G.O.), Medical University of Silesia, Katowice; Department of Neurology (M.R., A.K.-W.), Jagiellonian University, Krakow; Department of Neurodegenerative Disorders (M.B.), Medical Research Centre, Polish Academy of Sciences, Warsaw; Department of Neurology (K.C.), Central Hospital of the Ministry of Interior and Administration, Warsaw, Poland; Lviv Regional Clinical Hospital (Y.S.), Ukraine; Department of Neurology and School of Medicine (I.R.), Central European Institute of Technology, Masaryk University, Brno, Czech Republic; Departments of Neurology (A.H., J.E.A., B.F.B., R.C.P.) and Pathology and Laboratory Medicine (J.E.P.), Mayo Clinic, Rochester, MN; Department of Neurology (D.M.M.), NorthShore University Health System, Evanston, IL; and Department of Neurology (C.H.A.), Mayo Clinic, Scottsdale, AZ.

Objective: To assess the importance of MAPT variant p.A152T in the risk of synucleinopathies.

Methods: In this case-control study, we screened a large global series of patients and controls, and assessed associations between p.A152T and disease risk. We included 3,229 patients with clinical Parkinson disease (PD), 442 with clinical dementia with Lewy bodies (DLB), 181 with multiple system atrophy (MSA), 832 with pathologically confirmed Lewy body disease (LBD), and 2,456 healthy controls.

Results: The minor allele frequencies (MAF) in clinical PD cases (0.28%) and in controls (0.2%) were not found to be significantly different (odds ratio [OR] 1.37, 95% confidence interval [CI] 0.63-2.98, p = 0.42). However, a significant association was observed with clinical DLB (MAF 0.68%, OR 5.76, 95% CI 1.62-20.51, p = 0.007) and LBD (MAF 0.42%, OR 3.55, 95% CI 1.04-12.17, p = 0.04). Additionally, p.A152T was more common in patients with MSA compared to controls (MAF 0.55%, OR 4.68, 95% CI 0.85-25.72, p = 0.08) but this was not statistically significant and therefore should be interpreted with caution.

Conclusions: Overall, our findings suggest that MAPT p.A152T is a rare low penetrance variant likely associated with DLB that may be influenced by coexisting LBD and AD pathology. Given the rare nature of the variant, further studies with greater sample size are warranted and will help to fully explain the role of p.A152T in the pathogenesis of the synucleinopathies.
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http://dx.doi.org/10.1212/WNL.0000000000001946DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4653108PMC
November 2015

Targeting α-synuclein oligomers by protein-fragment complementation for drug discovery in synucleinopathies.

Expert Opin Ther Targets 2015 May 18;19(5):589-603. Epub 2015 Mar 18.

Mayo Clinic Florida, Neuroscience , 4500 San Pablo road, Jacksonville, 32224, FL , USA

Objective: Reducing the burden of α-synuclein oligomeric species represents a promising approach for disease-modifying therapies against synucleinopathies such as Parkinson's disease and dementia with Lewy bodies. However, the lack of efficient drug discovery strategies that specifically target α-synuclein oligomers has been a limitation to drug discovery programs.

Research Design And Methods: Here we describe an innovative strategy that harnesses the power of bimolecular protein-fragment complementation to monitor synuclein-synuclein interactions. We have developed two robust models to monitor α-synuclein oligomerization by generating novel stable cell lines expressing α-synuclein fusion proteins for either fluorescent or bioluminescent protein-fragment complementation under the tetracycline-controlled transcriptional activation system.

Main Outcome Measures: A pilot screen was performed resulting in the identification of two potential hits, a p38 MAPK inhibitor and a casein kinase 2 inhibitor, thereby demonstrating the suitability of our protein-fragment complementation assay for the measurement of α-synuclein oligomerization in living cells at high throughput.

Conclusions: The application of the strategy described herein to monitor α-synuclein oligomer formation in living cells with high throughput will facilitate drug discovery efforts for disease-modifying therapies against synucleinopathies and other proteinopathies.
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http://dx.doi.org/10.1517/14728222.2015.1009448DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4608017PMC
May 2015

Extracellular ATP induces intracellular alpha-synuclein accumulation via P2X1 receptor-mediated lysosomal dysfunction.

Neurobiol Aging 2015 Feb 5;36(2):1209-20. Epub 2014 Nov 5.

Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA; Mayo Graduate School, Mayo Clinic, Jacksonville, FL, USA. Electronic address:

The pathologic hallmark of Parkinson's disease (PD) is the accumulation of alpha-synuclein (αsyn) in susceptible neurons in the form of Lewy bodies and Lewy neurites. The etiology of PD remains unclear. Because brain injury has been suggested to facilitate αsyn aggregation, we investigated whether cellular breakdown products from damaged cells can act on neighboring healthy cells and cause intracellular αsyn accumulation and/or aggregation. Using 2 neuronal cell models, we found that extracellular adenosine triphosphate (ATP) induced a significant increase in intracellular αsyn levels between 24 and 48 hours after treatment. Further investigation revealed that the observed αsyn accumulation is a result of lysosome dysfunction caused by extracellular ATP-induced elevation of lysosomal pH. Interestingly, P2X1 receptor appears to mediate the cells' response to extracellular ATP. Although Ca(2+) influx via P2X1 receptor is necessary for αsyn accumulation, Ca(2+) influx per se is not sufficient for increased αsyn accumulation. These findings provide new insight into our knowledge of the role of P2X receptors in PD pathogenesis and may be helpful in identifying new therapeutic targets for PD.
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http://dx.doi.org/10.1016/j.neurobiolaging.2014.10.037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4315767PMC
February 2015

Mutual exacerbation of peroxisome proliferator-activated receptor γ coactivator 1α deregulation and α-synuclein oligomerization.

Ann Neurol 2015 Jan 19;77(1):15-32. Epub 2014 Dec 19.

Department of Neurology, Ulm University, Ulm, Germany; Inoviem Scientific, Strasbourg, France.

Objective: Aggregation of α-synuclein (α-syn) and α-syn cytotoxicity are hallmarks of sporadic and familial Parkinson disease (PD), with accumulating evidence that prefibrillar oligomers and protofibrils are the pathogenic species in PD and related synucleinopathies. Peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), a key regulator of mitochondrial biogenesis and cellular energy metabolism, has recently been associated with the pathophysiology of PD. Despite extensive effort on studying the function of PGC-1α in mitochondria, no studies have addressed whether PGC-1α directly influences oligomerization of α-syn or whether α-syn oligomers impact PGC-1α expression.

Materials And Methods: We tested whether pharmacological or genetic activation of PGC-1α or PGC-11α knockdown could modulate the oligomerization of α-syn in vitro by using an α-syn -fragment complementation assay.

Results: In this study, we found that both PGC-1α reference gene (RG-PGC-1α) and the central nervous system (CNS)-specific PGC-1α (CNS-PGC-1α) are downregulated in human PD brain, in A30P α-syn transgenic animals, and in a cell culture model for α-syn oligomerization. Importantly, downregulation of both RG-PGC-1α and CNS-PGC-1α in cell culture or neurons from RG-PGC-1α-deficient mice leads to a strong induction of α-syn oligomerization and toxicity. In contrast, pharmacological activation or genetic overexpression of RG-PGC-1α reduced α-syn oligomerization and rescued α-syn-mediated toxicity.

Interpretation: Based on our results, we propose that PGC-1α downregulation and α-syn oligomerization form a vicious circle, thereby influencing and/or potentiating each other. Our data indicate that restoration of PGC-1α is a promising approach for development of effective drugs for the treatment of PD and related synucleinopathies.
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http://dx.doi.org/10.1002/ana.24294DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4293280PMC
January 2015
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