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α-Synuclein and Its A30P Mutant Affect Actin Cytoskeletal Structure and Dynamics
2009-08-15
The function of α-synuclein, a soluble protein abundant in the brain and concentrated at presynaptic terminals, is still undefined. Yet, α-synuclein overexpression and the expression...Full Text Available
α-Synuclein and Its A30P Mutant Affect Actin Cytoskeletal Structure and Dynamics
2009-08-15
Full Text Available.The function of α-synuclein, a soluble protein abundant in the brain and concentrated at presynaptic terminals, is still undefined. Yet, α-synuclein overexpression and the expression of its A30P mutant are associated with familial Parkinson's disease. Working in cell-free conditions, in two cell lines as well as in primary neurons we demonstrate that α-synuclein and its A30P mutant have different effects on actin polymerization. Wild-type α-synuclein binds actin, slows down its polymerization and accelerates its depolymerization, probably by monomer sequestration; A30P mutant α-synuclein increases the rate of actin polymerization and disrupts the cytoskeleton during reassembly of actin filaments. Consequently, in cells expressing mutant α-synuclein, cytoskeleton-dependent processes, such as cell migration, are inhibited, while exo- and endocytic traffic is altered. In hippocampal neurons from mice carrying a deletion of the α-synuclein gene, electroporation of wild-type α-synuclein increases actin instability during remodeling, with growth of lamellipodia-like structures and apparent cell enlargement, whereas A30P α-synuclein induces discrete actin-rich foci during cytoskeleton reassembly. In conclusion, α-synuclein appears to play a major role in actin cytoskeletal dynamics and various aspects of microfilament function. Actin cytoskeletal disruption induced by the A30P mutant might alter various cellular processes and thereby play a role in the pathogenesis of neurodegeneration.
2007-01-01
The aggregation of a-synuclein is a hallmark feature of Parkinson's disease (PD) and other synucleinopathies. Metals are the significant etiological factors in PD, and their interaction with a-synuclein affect dramatically the kinetics of fibrillation in vitro and are proposed to play an important and potential neurodegenerative role in vivo. In the present study, we investigated the stoichiometry of binding of copper [Cu (II)] and iron [Fe (III)] with a-synuclein (wild recombinant type and A30P, A53T, E46K mutant forms) using isothermal titration calorimetry (ITC). a-Synuclein monomer (wild and mutant forms) titrated by Cu (II), showed two binding sites, with an apparent KB of 105M and 104M, respectively. But, a-synuclein (wild type and mutant forms) titrated with Fe (III) revealed a KB o...
2007-01-01
The aggregation of alpha-synuclein is a hallmark feature of Parkinson's disease (PD) and other synucleinopathies. Metals are the significant etiological factors in PD, and their interaction with alpha-synuclein affect dramatically the kinetics of fibrillation in vitro and are proposed to play an important and potential neurodegenerative role in vivo. In the present study, we investigated the stoichiometry of binding of copper [Cu (II)] and iron [Fe (III)] with alpha-synuclein (wild recombinant type and A30P, A53T, E46K mutant forms) using isothermal titration calorimetry (ITC). alpha-Synuclein monomer (wild and mutant forms) titrated by Cu (II), showed two binding sites, with an apparent K B of 105 M and 104 M, respectively. But, alpha-synuclein (wild type and ... >>
The aggregation of {alpha}-synuclein is a hallmark feature of Parkinson's disease (PD) and other synucleinopathies. Metals are the significant etiological factors in PD, and their interaction with {alpha}-synuclein affect dramatically the kinetics of fibrillation in vitro and are proposed to play an important and potential neurodegenerative role in vivo. In the present study, we investigated the stoichiometry of binding of copper [Cu (II)] and iron [Fe (III)] with {alpha}-synuclein (wild recombinant type and A30P, A53T, E46K mutant forms) using isothermal titration calorimetry (ITC). {alpha}-Synuclein monomer (wild and mutant forms) titrated by Cu (II), showed two binding sites, with an apparent K {sub B} of 10{sup 5} M and 10{sup 4} M, respectively. But, {alpha}-synuclein (wild type and mutant forms) titrated with Fe (III) revealed a K {sub B} of 10{sup 5} M with single binding site. The present investigation uncovers the detailed binding propensities between metals and {alpha}-synuclein and has biological implications in PD.
http://hdl.handle.net/2440/34519
α-Synuclein is a pre-synaptic protein, the function of which is not completely understood, but its pathological form is involved in neurodegenerative diseases. In vitro, α-synuclein spontaneously forms amyloid fibrils. Here, we report that αB-crystallin, a molecular chaperone found in Lewy bodies that are characteristic of Parkinson's disease (PD), is a potent in vitro inhibitor of α-synuclein fibrillization, both of wild-type and the two mutant forms (A30P and A53T) that cause familial, early onset PD. In doing so, large irregular aggregates of α-synuclein and αB-crystallin are formed implying that αB-crystallin redirects α-synuclein from a fibril-formation pathway towards an amorphous aggregation pathway, thus reducing the amount of physiologically stable amyloid deposits in favor of easily degradable amorphous aggregates. α-Synuclein acts as a molecular chaperone to prevent the stress-induced, amorphous aggregation of target proteins. Compared to wild-type α-synuclein, both mutant forms have decreased chaperone activity in vitro against the aggregation of reduced insulin at 37 °C and the thermally induced aggregation of βL-crystallin at 60 °C. Wild-type α-synuclein abrogates the chaperone activity of αB-crystallin to prevent the precipitation of reduced insulin. Interaction between these two chaperones and formation of a complex are also indicated by NMR spectroscopy, size-exclusion chromatography and mass spectrometry. In summary, α-synuclein and αB-crystallin interact readily with each other and affect each other's properties, in particular α-synuclein fibril formation and αB-crystallin chaperone action.Agata Rekas, Christopher G. Adda, J. Andrew Aquilina, Kevin J. Barnham, Margaret Sunde, Denise Galatis, Nicholas A. Williamson, Colin L. Masters, Robin F. Anders, Carol V. Robinson, Roberto Cappai and John A. Carverhttp://www.elsevier.com/wps/find/journaldescription.cws_home/622890/description#description Publisher: Academic Press - Elsevier Contributor: School of Chemistry and Physics : Chemistry Other identifier: Journal of Molecular Biology, 2004; 340 (5):1167-1183; 0022-2836; RQF0000022; 10.1016/j.jmb.2004.05.054 Language: en
2009-03-20
Full Text Available.Fibrillization or conformational change of α-synuclein is central in the pathogenesis of α-synucleinopathies, such as Parkinson disease. We found that the A30P mutant accelerates nucleation-dependent fibrillization of wild type (WT) α-synuclein. Electron microscopy observation and ultracentrifugation experiments revealed that shedding of fragments occurs from A30P fibrils and that these fragments accelerate fibrillization by serving as seeds. Immunochemical analysis using epitope-specific antibodies and biochemical analyses of protease-resistant cores demonstrated that A30P fibrils have a distinct conformation. Interestingly, WT fibrils formed with A30P seeds exhibited the same character as A30P fibrils, as did A30P fibrils formed with WT seeds, indicating that the A30P mutation affects the conformation and fibrillization of both WT and A30P. These effects of A30P mutation may explain the apparent conflict between the association of A30P with Parkinson disease and the slow fibrillization of A30P itself and therefore provide new insight into the molecular mechanisms of α-synucleinopathies.
A Cellular Model To Monitor Proteasome Dysfunction by α-Synuclein†
2009-08-25
Full Text Available.Impairment of the ubiquitin-proteasome degradation system has recently been suggested to be related to the onset of neurodegenerative disorders such as Alzheimer′s disease and Parkinson's disease. In this study, we investigated whether intracellular α-synuclein affects proteasome activity in SH-SY5Y cells. To monitor intracellular proteasome activity, we used a reporter consisting of a short peptide degron fused to the carboxyl-terminus of green fluorescent protein (GFP-CL1), which is known to be degraded by proteasome. The level of intact GFP-CL1 was dramatically increased by coexpression of GFP-CL1 and α-synuclein, as judged by confocal microscopic and immunoblot analyses. Expression of two pathogenic mutants of α-synuclein, A30P and A53T, and phosphomimetic S129D mutant increased the intensities of GFP more effectively than did wild-type α-synuclein. GFP fluorescence in cells transfected with Δ73-83 mutant or β-synuclein, which does not assemble into filaments in vitro, was not changed as compared with that in cells expressing GFP-CL1 alone. Thus, the ability of α-synuclein to inhibit proteasome activity is related to its propensity to assemble into filaments. Furthermore, we observed that some compounds inhibiting α-synuclein filament formation in vitro prevented the α-synuclein-mediated proteasome dysfunction in cells transfected with both GFP-CL1 and α-synuclein. The cellular model expressing both GFP-CL1 and α-synuclein may be a useful tool to screen compounds protecting neurons from α-synuclein-mediated proteasome dysfunction.
Curvature Dynamics of α-Synuclein Familial Parkinson Disease Mutants
2009-03-13
Full Text Available.α-Synuclein remains a protein of interest due to its propensity to form fibrillar aggregates in neurodegenerative disease and its putative function in synaptic vesicle regulation. Herein, we present a series of atomistic molecular dynamics simulations of wild-type α-synuclein and three Parkinson disease familial mutants (A30P, A53T, and E46K) in two distinct environments. First, in order to match recent NMR experiments, we have simulated each protein bound to an SDS detergent micelle. Second, in order to connect more closely to the true biological environment, we have simulated the proteins bound to a 1,2-dioleoyl-sn-glycero-3-phosphoserine lipid bilayer. In the micelle-bound case, we find that the wild type and all of the variants of α-synuclein flatten the underlying micelle, decreasing its surface area. A30P is known to lessen α-synuclein/membrane affinity and, consistent with experiment, destabilizes the simulated secondary structure. In the case of A53T, our simulations reveal a range of stabilizing hydrogen bonds that form with the threonine. In both environments, the E46K mutation, which is known to increase bilayer affinity, leads to an additional hydrogen bond between the protein and either the detergent or lipid. Simulations indicate that αS and its variants are less dynamic in the bilayer than in the micelle. Furthermore, the simulations of the mutants suggest how changes in the structure and dynamics of α-synuclein may affect its biological role.
axion: Axion - Java Database - News
The Axion news feed is on hiatus, yet Axion is still being actively developed. ... Better late than never, it's the Axion database project's October 2003 ...
A small, fast, SQL and JDBC compliant relational database engine written in and for the Java programming language. [Open source, BSD License]
arXiv:hep-ph/9506229 v1 02 Jun 95
File Format: PDF/Adobe Acrobat - View as HTMLThe axion is the quantum of oscillation of the ? parameter of QCD. It is a particle ... ticle physics is the PQ mechanism with an ?invisible? axion. ...
Key Words thymocyte selection, T cell activation, immunosuppression, apoptosis Abstract Glucocorticoids are small lipophilic compounds that mediate their many biological effects by binding an intracellular receptor (GR) that, in turn, trans-locates to the nucleus and directly or indirectly regulates gene transcription. Perhaps the most recognized biologic effect of glucocorticoids on peripheral T cells is immu-nosuppression, which is due to inhibition of expression of a wide variety of activation-induced gene products.
The Influence of Vesicle Size and Composition on α-Synuclein Structure and Stability
2009-04-08
Full Text Available.AbstractMonomeric α-synuclein (αSN), which has no persistent structure in aqueous solution, is known to bind to anionic lipids with a resulting increase in α-helix structure. Here we show that at physiological pH and ionic strength, αSN incubated with different anionic lipid vesicles undergoes a marked increase in α-helical content at a temperature dictated either by the temperature of the lipid phase transition, or (in 1,2-DilauroylSN-Glycero-3-[Phospho-rac-(1-glycerol)] (DLPG), which is fluid down to 0°C) by an intrinsic cold denaturation that occurs around 10–20°C. This structure is subsequently lost in a thermal transition around 60°C. Remarkably, this phenomenon is only observed for vesicles >100 nm in diameter and is sensitive to lipid chain length, longer chain lengths, and larger vesicles giving more cooperative unfolding transitions and a greater degree of structure. For both vesicle size and chain length, a higher degree of compressibility or permeability in the lipid thermal transition region is associated with a higher degree of αSN folding. Furthermore, the degree of structural change is strongly reduced by an increase in ionic strength or a decrease in the amount of anionic lipid. A simple binding-and-folding model that includes the lipid phase transition, exclusive binding of αSN to the liquid disordered phase, the thermodynamics of unfolding, and the electrostatics of binding of αSN to lipids is able to reproduce the two thermal transitions as well as the effect of ionic strength and anionic lipid. Thus the nature of αSN's binding to phospholipid membranes is intimately tied to the lipids' physico-chemical properties.
The Influence of Vesicle Size and Composition on α-Synuclein Structure and Stability
2009-04-08
AbstractMonomeric α-synuclein (αSN), which has no persistent structure in aqueous solution, is known to bind to anionic lipids with a...Full Text Available
Targeting the Progression of Parkinson’s Disease
2009-03-01
Full Text Available.By the time a patient first presents with symptoms of Parkinson’s disease at the clinic, a significant proportion (50-70%) of the cells in the substantia nigra (SN) has already been destroyed. This degeneration progresses until, within a few years, most of the cells have died. Except for rare cases of familial PD, the initial trigger for cell loss is unknown. However, we do have some clues as to why the damage, once initiated, progresses unabated. It would represent a major advance in therapy to arrest cell loss at the stage when the patient first presents at the clinic. Current therapies for Parkinson’s disease focus on relieving the motor symptoms of the disease, these unfortunately lose their effectiveness as the neurodegeneration and symptoms progress. Many experimental approaches are currently being investigated attempting to alter the progression of the disease. These range from replacement of the lost neurons to neuroprotective therapies; each of these will be briefly discussed in this review. The main thrust of this review is to explore the interactions between dopamine, alpha synuclein and redox-active metals. There is abundant evidence suggesting that destruction of SN cells occurs as a result of a self-propagating series of reactions involving dopamine, alpha synuclein and redox-active metals. A potent reducing agent, the neurotransmitter dopamine has a central role in this scheme, acting through redox metallo-chemistry to catalyze the formation of toxic oligomers of alpha-synuclein and neurotoxic metabolites including 6-hydroxydopamine. It has been hypothesized that these feed the cycle of neurodegeneration by generating further oxidative stress. The goal of dissecting and understanding the observed pathological changes is to identify therapeutic targets to mitigate the progression of this debilitating disease.
Targeting the Progression of Parkinson’s Disease
2009-03-01
By the time a patient first presents with symptoms of Parkinson’s disease at the clinic, a significant proportion (50-70%) of the cells in the substantia nigra (SN) has already been destroyed....Full Text Available
Switch Region for Pathogenic Structural Change in Conformational Disease and Its Prediction
Many diseases are believed to be related to abnormal protein folding. In the first step of such pathogenic structural changes, misfolding occurs in regions important for the stability of the native...Full Text Available
Switch Region for Pathogenic Structural Change in Conformational Disease and Its Prediction
Full Text Available.Many diseases are believed to be related to abnormal protein folding. In the first step of such pathogenic structural changes, misfolding occurs in regions important for the stability of the native structure. This destabilizes the normal protein conformation, while exposing the previously hidden aggregation-prone regions, leading to subsequent errors in the folding pathway. Sites involved in this first stage can be deemed switch regions of the protein, and can represent perfect binding targets for drugs to block the abnormal folding pathway and prevent pathogenic conformational changes. In this study, a prediction algorithm for the switch regions responsible for the start of pathogenic structural changes is introduced. With an accuracy of 94%, this algorithm can successfully find short segments covering sites significant in triggering conformational diseases (CDs) and is the first that can predict switch regions for various CDs. To illustrate its effectiveness in dealing with urgent public health problems, the reason of the increased pathogenicity of H5N1 influenza virus is analyzed; the mechanisms of the pandemic swine-origin 2009 A(H1N1) influenza virus in overcoming species barriers and in infecting large number of potential patients are also suggested. It is shown that the algorithm is a potential tool useful in the study of the pathology of CDs because: (1) it can identify the origin of pathogenic structural conversion with high sensitivity and specificity, and (2) it provides an ideal target for clinical treatment.
Smoking's Effects on Genes May Play a Role in Lung Cancer Development and Survival
Smoking plays a role in lung cancer development and now scientists have shown that smoking also affects the way genes are expressed, leading to alterations in cell division and regulation of immune response.
Risk as Analysis and Risk as Feelings: Some Thoughts about Affect, Reason, and Risk
That intuitive feelings are still the predominant method by which human beings evaluate risk is cleverly illustrated in a cartoon by Garry Trudeau (Figure 1). Trudeau?s two characters decide whether to greet one another on a city street by employing a systematic analysis of the risks and risk-mitigating factors. We instantly recognize that no one in such a situation would ever be this analytical, even if their life was at stake.
Parkinson’s disease (PD) is the second most common progressive neurodegenerative disorder after Alzheimer’s disease (AD) and represents a large health burden to society. Genetic and...Full Text Available
Full Text Available.Parkinson’s disease (PD) is the second most common progressive neurodegenerative disorder after Alzheimer’s disease (AD) and represents a large health burden to society. Genetic and oxidative risk factors have been proposed as possible causes, but their relative contribution remains unclear. Dysfunction of alpha-synuclein (α-syn) has been associated with PD due to its increased presence, together with iron, in Lewy bodies. Brain oxidative damage caused by iron may be partly mediated by α-syn oligomerization during PD pathology. Also, α-syn gene dosage can cause familial PD and inhibition of its gene expression by blocking translation via a newly identified Iron Responsive Element-like RNA sequence in its 5’-untranslated region may provide a new PD drug target.
Parkin Reverses Intracellular β-Amyloid Accumulation and Its Negative Effects on Proteasome Function
2010-01-01
The significance of intracellular β-amyloid (Aβ42) accumulation is increasingly recognized in Alzheimer's disease (AD) pathogenesis. Aβ removal mechanisms...Full Text Available
Parkin Reverses Intracellular β-Amyloid Accumulation and Its Negative Effects on Proteasome Function
2010-01-01
Full Text Available.The significance of intracellular β-amyloid (Aβ42) accumulation is increasingly recognized in Alzheimer's disease (AD) pathogenesis. Aβ removal mechanisms that have attracted attention include IDE/neprilysin degradation and antibody-mediated uptake by immune cells. However, the role of the ubiquitin-proteasome system (UPS) in the disposal of cellular Aβ has not been fully explored. The E3 ubiquitin ligase Parkin targets several proteins for UPS degradation, and Parkin mutations are the major cause of autosomal recessive Parkinson's disease. We tested whether Parkin has cross-function to target misfolded proteins in AD for proteasome-dependent clearance in SH-SY5Y and primary neuronal cells. Wild-type Parkin greatly decreased steady-state levels of intracellular Aβ42, an action abrogated by proteasome inhibitors. Intracellular Aβ42 accumulation decreased cell viability and proteasome activity. Accordingly, Parkin reversed both effects. Changes in mitochondrial ATP production from Aβ or Parkin did not account for their effects on the proteasome. Parkin knock-down led to accumulation of Aβ. In AD brain, Parkin was found to interact with Aβ and its levels were reduced. Thus, Parkin is cytoprotective, partially by increasing the removal of cellular Aβ through a proteasome-dependent pathway.
While numerous hypotheses have been proposed to explain the molecular mechanisms underlying the pathogenesis of neurodegenerative diseases, the theory of oxidative stress has received considerable support....Full Text Available
Full Text Available.While numerous hypotheses have been proposed to explain the molecular mechanisms underlying the pathogenesis of neurodegenerative diseases, the theory of oxidative stress has received considerable support. Although many correlations have been established and encouraging evidence has been obtained, conclusive proof of causation for the oxidative stress hypothesis is lacking and potential cures have not emerged. Therefore it is likely that other factors, possibly in coordination with oxidative stress, contribute to neuron death. Using Parkinson's disease (PD) as the paradigm, this review explores the hypothesis that oxidative modifications, mitochondrial functional disruption, and impairment of protein degradation constitute three interrelated molecular pathways that execute neuron death. These intertwined events are the consequence of environmental exposure, genetic factors, and endogenous risks and constitute a "Bermuda triangle" that may be considered the underlying cause of neurodegenerative pathogenesis.
Montreal Axion - Wikipedia, the free encyclopedia
The Montreal Axion are a National Women's Hockey League team located in Montreal, Quebec, Canada. v ? d ? e · Sports teams based in the province of Quebec, ...
Molecular Pathology of Lewy Body Diseases
Lewy body diseases are characterized by the presence of Lewy bodies, alpha-synuclein(AS)-positive inclusions in the brain. Since their main component is conformationally modified AS, aggregation of...Full Text Available
Molecular Pathology of Lewy Body Diseases
Full Text Available.Lewy body diseases are characterized by the presence of Lewy bodies, alpha-synuclein(AS)-positive inclusions in the brain. Since their main component is conformationally modified AS, aggregation of the latter is thought to be a key pathogenic event in these diseases. The analysis of inclusion body constituents gives additional information about pathways also involved in the pathology of synucleinopathies. Widespread mitochondrial dysfunction is very closely related to disease development. The impairment of protein degradation pathways, including both the ubiquitin-proteasome system and the autophagy-lysosome pathway also play an important role during the development of Lewy body diseases. Finally, differential expression changes of isoforms corresponding to genes primarily involved in Lewy body formation point to alternative splicing as another important mechanism in the development of Parkinson’s disease, as well as dementia with Lewy bodies. The present paper attempts to give an overview of recent molecular findings related to the pathogenesis of Lewy body diseases.
Recommendations of the Pancreatic Cancer Progress Review Group highlighted the need to evaluate environmental risk factors and gene-environment interactions and to identify biomarkers of early disease.
Microsoft Word - chap24.reports.final.doc
Creating Reports and Extracting Data 9/20/2009 24-1 Chapter 24: Creating Reports and Extracting Data SEER*DMS includes an integrated reporting and extract module that enables you to run pre-defined system reports and create standard extracts.
Microsoft Word - Newsletter March2005_draft2.doc
To unsubscribe from this newsletter please send an email to Dr. Betty Tarnowski tarnowsb@mail.nih.gov Send meeting announcements and other information you would like to have included in this newsletter to Ulli Wagner: ulrike@mail.nih.gov M MM MH
Microsoft Word - M&E_Archive_2003_JUN26-27_ExecSum.doc
Sciences disclaims any responsibility or liability for errors or omissions in this version of the manuscript. The final version of this manuscript, which is the citable version and the version of record, was published in the November, 2004 issue of The Journal of Nutrition.
Paul Bunn, Jr., M.D. Neil Caporaso, M.D. University of Colorado Cancer Center National Cancer Institute Neil Clendeninn, M.D., Ph.D. Walter J. Curran, Jr., M.D. Agouron Pharmaceuticals, Inc. Virginia L. Ernster, Ph.D. Bodine Center for Cancer Treatment, Philadelphia William Evans, M.D. University of California, San Francisco Cancer Care Ontario School of Medicine Dorothy Hatsukami, Ph.D. Stephen S.
Evidence that α-Synuclein Does Not Inhibit Phospholipase D†
2009-02-10
Full Text Available.α-Synuclein (αSyn) is a small cytosolic protein of unknown function, which is highly enriched in the brain. It is genetically linked to Parkinson’s disease (PD) in that missense mutations or multiplication of the gene encoding αSyn cause early-onset familial PD. Furthermore, the neuropathological hallmarks of both sporadic and familial PD, Lewy bodies and Lewy neurites, contain insoluble aggregates of αSyn. Several studies have reported evidence that αSyn can inhibit phospholipase D (PLD), which hydrolyzes phosphatidylcholine to form phosphatidic acid and choline. Although various hypotheses exist regarding the roles of αSyn in health and disease, no other specific biochemical function for this protein has been reported to date. Because PLD inhibition could represent an important function of αSyn, we sought to extend existing reports on this interaction. Using purified proteins, we tested the ability of αSyn to inhibit PLD activity in cell-free assays. We also examined several cell lines and transfection conditions to assess whether αSyn inhibits endogenous or overexpressed PLD in cultured mammalian cells. In yeast, we extended our previous report of an interaction between αSyn and PLD-dependent phenotypes, for which PLD activity is absolutely necessary. Despite testing a range of experimental conditions, including those previously published, we observed no significant inhibition of PLD by αSyn in any of these systems. We propose that the previously reported effects of αSyn on PLD activity could be due to increased endoplasmic reticulum-related stress associated with αSyn overexpression in cells, but are not likely due to a specific and direct interaction between αSyn and PLD.
Evidence that α-Synuclein Does Not Inhibit Phospholipase D†
2009-02-10
α-Synuclein (αSyn) is a small cytosolic protein of unknown function, which is highly enriched in the brain. It is genetically linked to Parkinson’s disease (PD) in that...Full Text Available
2006-01-01
6-Hydroxydopamine (6-OHDA) is widely used to produce animal models of Parkinsons disease (PD) by selectively destroying the nigro-striatal dopaminergic systems, but selective toxicity of 6-OHDA towards dopaminergic cells in vitro remains controversial. Mutant (A30P and A53T) a-synuclein isoforms cause increased vulnerability of cells towards various toxic insults and enhance dopamine transporter (DAT)-mediated toxicity of the selective dopaminergic neurotoxin and mitochondrial complex I inhibitor MPP+ in vitro. Here we extend our recent studies on DAT-mediated toxicity to elucidate the mechanisms involved in selective dopaminergic toxicity of 6-OHDA. We studied the cytotoxicity as well as the toxic mechanisms of 6-OHDA in human embryonic kidney HEK-293 cells ectopically co-expressing mutan...
2010-02-09
Three familial variants of the presynaptic protein α-synuclein...Full Text Available
2010-02-09
Full Text Available.Three familial variants of the presynaptic protein α-synuclein (αS), A30P, E46K, and A53T, correlate with rare inherited Parkinson’s disease (PD), while wild-type αS is implicated in sporadic PD. The classic manifestation of both familiar and sporadic PD is the formation of fibrillar structures of αS which accumulate as the main component in intraneuronal Lewy bodies. At presynaptic termini, the partitioning of αS between disordered cytosolic and membrane-bound states likely mediates its proposed role in regulation of reserve pools of synaptic vesicles. Previously, we reported on multiple distinct phospholipid binding modes of αS with slow binding kinetics. Here, we report the phospholipid binding properties of the disease variants, viewed by solution NMR in a residue-specific manner. Our results agree qualitatively with previous biophysical studies citing overall decreased lipid affinity for the A30P mutation, comparable affinity for A53T, and an increased level of binding of E46K, relative to wild-type αS. Additionally, our NMR results describe the distribution of lipid-bound states for αS: the population of the SL1 binding mode (residues 3−25 bound as a helix) is augmented by each of the disease variants, relative to wild-type αS. We propose that the SL1 binding mode, which anchors the N-terminus of αS in the lipoprotein complex while the hydrophobic NAC region remains dynamically disordered, is prone to intermolecular interactions which progress toward disease-associated oligomers and fibrils. The elevation of the SL1 binding mode, unchecked by a proportionate population of binding modes incorporating the full N-terminal domain, may well account for the increased toxicity of the A30P, E46K, and A53T disease variants of αS.
Developing tTA Transgenic Rats for Inducible and Reversible Gene Expression
To develop transgenic lines for conditional expression of desired genes in rats, we generated several lines of the transgenic rats carrying the tetracycline-controlled transactivator (tTA) gene. Using...Full Text Available
Developing tTA Transgenic Rats for Inducible and Reversible Gene Expression
Full Text Available.To develop transgenic lines for conditional expression of desired genes in rats, we generated several lines of the transgenic rats carrying the tetracycline-controlled transactivator (tTA) gene. Using a vigorous, ubiquitous promoter to drive the tTA transgene, we obtained widespread expression of tTA in various tissues. Expression of tTA was sufficient to strongly activate its reporter gene, but was below the toxicity threshold. We examined the dynamics of Doxycycline (Dox)-regulated gene expression in transgenic rats. In the two transmittable lines, tTA-mediated activation of the reporter gene was fully subject to regulation by Dox. Dox dose-dependently suppressed tTA-activated gene expression. The washout time for the effects of Dox was dose-dependent. We tested a complex regime of Dox administration to determine the optimal effectiveness and washout duration. Dox was administered at a high dose (500 μg/ml in drinking water) for two days to reach the effective concentration, and then was given at a low dose (20 μg/ml) to maintain effectiveness. This regimen of Dox administration can achieve a quick switch between ON and OFF statuses of tTA-activated gene expression. In addition, administration of Dox to pregnant rats fully suppressed postnatal tTA-activated gene expression in their offspring. Sufficient levels of Dox are present in mother's milk to produce maximal efficacy in nursing neonates. Administration of Dox to pregnant or nursing rats can provide a continual suppression of tTA-dependent gene expression during embryonic and postnatal development. The tTA transgenic rat allows for inducible and reversible gene expression in the rat; this important tool will be valuable in the development of genetic rat models of human diseases.
Curvature Dynamics of α-Synuclein Familial Parkinson Disease Mutants
2009-03-13
α-Synuclein remains a protein of interest due to its propensity to form fibrillar aggregates in neurodegenerative disease and its putative function in synaptic vesicle regulation. Herein,...Full Text Available
Copper- and iron-induced differential fibril formation in a-synuclein: TEM study
2007-01-01
a-Synuclein filaments are the central component of intracytoplasmic inclusion bodies characteristic of Parkinsons disease (PD) and related disorders. Metals are the significant etiological factors in PD, and their interaction with a-synuclein affect dramatically the kinetics of fibrillation. Currently, we have investigated the influence of Cu(II) and Fe(III) on a-synuclein fibril formation. Cu(II) and Fe(III) selectively and differentially induced the formation of discrete a-synuclein fibrillar species. Transmission electron microscopy was used to monitor the aggregation state of a-synuclein (wild-type, A30P, A53T, and E46K) after 60h with stirring at 37degreeC in the presence and absence of metal ions. Cu(II) has induced thin long network-like fibrils with the wild-type of a-synuclein, wh...
2009-03-20
Fibrillization or conformational change of α-synuclein is central in the pathogenesis of α-synucleinopathies, such as Parkinson disease. We found that the A30P mutant accelerates nucleation-dependent...Full Text Available
Cell death pathways in Parkinson’s disease: proximal triggers, distal effectors, and final steps
2009-04-01
Parkinson’s disease (PD) is a common neurodegenerative disease. Neuronal cell death in PD is still poorly understood, despite a wealth of potential pathogenic mechanisms and pathways....Full Text Available
Cell death pathways in Parkinson’s disease: proximal triggers, distal effectors, and final steps
2009-04-01
Full Text Available.Parkinson’s disease (PD) is a common neurodegenerative disease. Neuronal cell death in PD is still poorly understood, despite a wealth of potential pathogenic mechanisms and pathways. Defects in several cellular systems have been implicated as early triggers that start cells down the road towards neuronal death. These include abnormal protein accumulation, particularly of alpha-synuclein; altered protein degradation via multiple pathways; mitochondrial dysfunction; oxidative stress; neuroinflammation; and dysregulated kinase signaling. As dysfunction in these systems mounts, pathways that are more explicitly involved in cell death become recruited. These include JNK signaling, p53 activation, cell cycle re-activation, and signaling through bcl-2 family proteins. Eventually, neurons become overwhelmed and degenerate; however, even the mechanism of final cell death in PD is still unsettled. In this review, we will discuss cell death triggers and effectors that are relevant to PD, highlighting important unresolved issues and implications for the development of neuroprotective therapies.
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Axion Project Incubation Status - Apache Incubator
For general project status, see the Axion project website. ... The axion project never moved to the ASF from tigris.org. 2003-12-19: The Apache Incubator ...
Axion - Wikipedia, the free encyclopedia
The axion is a hypothetical elementary particle postulated by Peccei-Quinn theory in 1977 to resolve the strong-CP problem in quantum chromodynamics (QCD). ...
Alterations in Corticostriatal Synaptic Plasticity in Mice Over-Expressing Human α-Synuclein
2009-03-17
Most forms of Parkinson’s Disease (PD) are sporadic in nature, but some have genetic causes as first described for the α-synuclein gene. The α-synuclein protein also...Full Text Available
Alterations in Corticostriatal Synaptic Plasticity in Mice Over-Expressing Human α-Synuclein
2009-03-17
Full Text Available.Most forms of Parkinson’s Disease (PD) are sporadic in nature, but some have genetic causes as first described for the α-synuclein gene. The α-synuclein protein also accumulates as insoluble aggregates in Lewy bodies in sporadic PD as well as in most inherited forms of PD. The focus of the present study is the modulation of synaptic plasticity in the corticostriatal pathway of transgenic (Tg) mice that over express the human α-synuclein protein throughout the brain (ASOTg). Paired-pulse facilitation was detected in vitro by activation of corticostriatal afferents in ASOTg mice, consistent with a presynaptic effect of elevated human α-synuclein. However basal synaptic transmission was unchanged in ASOTg, suggesting that human α-synuclein could impact paired-pulse facilitation via a presynaptic mechanism not directly related to the probability of neurotransmitter release. Mice lacking α-synuclein or those expressing normal and A53T human α-synuclein in tyrosine hydroxylase-containing neurons showed, instead, paired-pulse depression. High-frequency stimulation induced a presynaptic form of long-term depression solely in ASOTg striatum. A presynaptic, NMDA receptor-independent form of chemical long-term potentiation induced by forskolin (FSK) was enhanced in ASOTg striatum, while FSK-induced cAMP levels were reduced in ASOTg synaptoneurosome fractions. Overall the results suggest that elevated human α-synuclein alters presynaptic plasticity in the corticostriatal pathway, possibly reflecting a reduction in glutamate at corticostriatal synapses by modulation of adenylyl cyclase signaling pathways. ASOTg mice may recapitulate an early stage in PD during which over expressed α-synuclein dampens corticostriatal synaptic transmission and reduces movement.
Full Text Available.BackgroundThe mechanisms through which aberrant α-synuclein (ASYN) leads to neuronal death in Parkinson's disease (PD) are uncertain. In isolated liver lysosomes, mutant ASYNs impair Chaperone Mediated Autophagy (CMA), a targeted lysosomal degradation pathway; however, whether this occurs in a cellular context, and whether it mediates ASYN toxicity, is unknown. We have investigated presently the effects of WT or mutant ASYN on the lysosomal pathways of CMA and macroautophagy in neuronal cells and assessed their impact on ASYN-mediated toxicity.Methods and FindingsNovel inducible SH-SY5Y and PC12 cell lines expressing human WT and A53T ASYN, as well as two mutant forms that lack the CMA-targeting motif were generated. Such forms were also expressed in primary cortical neurons, using adenoviral transduction. In each case, effects on long-lived protein degradation, LC3 II levels (as a macroautophagy index), and cell death and survival were assessed. In both PC12 and SH-SY5Y cycling cells, induction of A53T ASYN evoked a significant decrease in lysosomal degradation, largely due to CMA impairment. In neuronally differentiated SH-SH5Y cells, both WT and A53T ASYN induction resulted in gradual toxicity, which was partly dependent on CMA impairment and compensatory macroautophagy induction. In primary neurons both WT and A53T ASYN were toxic, but only in the case of A53T ASYN did CMA dysfunction and compensatory macroautophagy induction occur and participate in death.ConclusionsExpression of mutant A53T, and, in some cases, WT ASYN in neuronal cells leads to CMA dysfunction, and this in turn leads to compensatory induction of macroautophagy. Inhibition of these lysosomal effects mitigates ASYN toxicity. Therefore, CMA dysfunction mediates aberrant ASYN toxicity, and may be a target for therapeutic intervention in PD and related disorders. Furthermore, macroautophagy induction in the context of ASYN over-expression, in contrast to other settings, appears to be a detrimental response, leading to neuronal death.
BackgroundThe mechanisms through which aberrant α-synuclein (ASYN) leads to neuronal death in Parkinson's disease (PD) are uncertain. In isolated liver lysosomes, mutant...Full Text Available
2009-04-03
Nurr1, an orphan nuclear receptor, plays an essential role in the generation and maintenance of dopaminergic neurons in the brain. Rare mutations in Nurr1 are associated with...Full Text Available
2009-04-03
Full Text Available.Nurr1, an orphan nuclear receptor, plays an essential role in the generation and maintenance of dopaminergic neurons in the brain. Rare mutations in Nurr1 are associated with familial Parkinson’s disease, but the underlying basis for this relationship has not been established. Here, we demonstrate that Nurr1 unexpectedly functions to inhibit expression of pro-inflammatory neurotoxic mediators in both microglia and astrocytes. Reduced Nurr1 expression results in exaggerated inflammatory responses in microglia that are further amplified by astrocytes, leading to the production of factors that cause death of tyrosine hydroxylase-expressing neurons. Nurr1 exerts anti-inflammatory effects by docking to NF-κB-p65 on target inflammatory gene promoters in a signal-dependent manner. Subsequently, Nurr1 recruits the CoREST corepressor complex, resulting in clearance of NF-κB-p65 and transcriptional repression. These studies suggest that Nurr1 protects against loss of dopaminergic neurons in Parkinson’s disease in part by limiting the production of neurotoxic mediators by microglia and astrocytes.
A Cellular Model To Monitor Proteasome Dysfunction by α-Synuclein†
2009-08-25
Impairment of the ubiquitin-proteasome degradation system has recently...Full Text Available
The desire to obtain maximal information from every clinical protocol offered at the Clinical Center drives our investment in genomic profiling. Analysis of every biospecimen donated by patients enrolled in our clinical trials is a major part of our information gathering. Our clinical researchers use a wealth of technologies and analysis platforms to obtain sophisticated data from each biospecimen collected.
γ-Synucleinopathy: neurodegeneration associated with overexpression of the mouse protein
2009-05-15
The role of α-synuclein in pathogenesis of familial and idiopathic forms of Parkinson’s disease, and other human disorders known as α-synucleinopathies, is well established....Full Text Available
γ-Synucleinopathy: neurodegeneration associated with overexpression of the mouse protein
2009-05-15
Full Text Available.The role of α-synuclein in pathogenesis of familial and idiopathic forms of Parkinson’s disease, and other human disorders known as α-synucleinopathies, is well established. In contrast, the involvement of two other members of the synuclein family, β-synuclein and γ-synuclein, in the development and progression of neurodegeneration is poorly studied. However, there is a growing body of evidence that α-synuclein and β-synuclein have opposite neuropathophysiological effects. Unlike α-synuclein, overexpressed β-synuclein does not cause pathological changes in the nervous system of transgenic mice and even ameliorates the pathology caused by overexpressed α-synuclein. To assess the consequences of excess expression of the third family member, γ-synuclein, on the nervous system we generated transgenic mice expressing high levels of mouse γ-synuclein under control of Thy-1 promoter. These animals develop severe age- and transgene dose-dependent neuropathology, motor deficits and die prematurely. Histopathological changes include aggregation of γ-synuclein, accumulation of various inclusions in neuronal cell bodies and processes, and astrogliosis. These changes are seen throughout the nervous system but are most prominent in the spinal cord where they lead to loss of spinal motor neurons. Our data suggest that down-regulation of small heat shock protein HSPB1 and disintegration of neurofilament network play a role in motor neurons dysfunction and death. These findings demonstrate that γ-synuclein can be involved in neuropathophysiological changes and the death of susceptible neurons suggesting the necessity of further investigations of the potential role of this synuclein in disease.
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