Synuclein Aggregation Research Articles

254 - Superoxide Stimulates DOPAL Autoxidation, Lysyl Adduct Formation, and Crosslinking of α-Synuclein

Parkinson’s etiology has long been associated with redox imbalance and oxidative stress in dopaminergic neurons. The catecholaldehyde hypothesis suggests that DOPAL, an obligate product of dopamine catabolism, is a central nexus in a network of pathways leading to disease-state neurodegeneration, owing to its toxicity and potent ability to oligomerize alpha synuclein, the main component of protein aggregates in Lewy bodies. In this work we examine the connection between reactive oxygen species and DOPAL autoxidation. We show that superoxide propagates a chain reaction oxidation, and that this reaction is dramatically inhibited by superoxide dismutase. Moreover, superoxide dismutase prevents DOPAL from forming dicatechol pyrrole adducts with lysine and from covalently crosslinking alpha synuclein. Given that superoxide is the main radical byproduct of impaired cellular respiration, our results provide a plausible mechanistic link between mitochondrial dysfunction and synuclein aggregation in dopaminergic neurons.

Polymer brain-nanotherapeutics for multipronged inhibition of microglial α-synuclein aggregation, activation, and neurotoxicity.

Neuroinflammation, a common neuropathologic feature of neurodegenerative disorders including Parkinson disease (PD), is frequently exacerbated by microglial activation. The extracellular protein α-synuclein (ASYN), whose aggregation is characteristic of PD, remains a key therapeutic target, but the control of synuclein trafficking and aggregation within microglia has been challenging. First, we established that microglial internalization of monomeric ASYN was mediated by scavenger receptors (SR), CD36 and SRA1, and was rapidly accompanied by the formation of ASYN oligomers. Next, we designed a nanotechnology approach to regulate SR-mediated intracellular ASYN trafficking within microglia. We synthesized mucic acid-derivatized sugar-based amphiphilic molecules (AM) with optimal stereochemistry, rigidity, and charge for enhanced dual binding affinity to SRs and fabricated serum-stable nanoparticles via flash nanoprecipitation comprising hydrophobe cores and amphiphile shells. Treatment of microglia with AM nanoparticles decreased monomeric ASYN internalization and intracellular ASYN oligomer formation. We then engineered composite deactivating NPs with dual character, namely shell-based SR-binding amphiphiles, and core-based antioxidant poly (ferrulic acid), to investigate concerted inhibition of oxidative activation. In ASYN-challenged microglia treated with NPs, we observed decreased ASYN-mediated acute microglial activation and diminished microglial neurotoxicity caused by exposure to aggregated ASYN. When the composite NPs were administered invivo within the substantia nigra of fibrillar ASYN-challenged wild type mice, there was marked attenuation of activated microglia. Overall, SR-targeting AM nanotechnology represents a novel paradigm in alleviating microglial activation in the context of synucleinopathies like PD and other neurodegenerative diseases.

Cyclodextrins 3-Functionalized with 8-Hydroxyquinolines: Copper-Binding Ability and Inhibition of Synuclein Aggregation.

Neurodegenerative diseases such as Parkinson's and Alzheimer's diseases are multifactorial disorders related to protein aggregation, metal dyshomeostasis, and oxidative stress. To advance understanding in this area and to contribute to therapeutic development, many efforts have been directed at devising suitable agents that can target metal ions associated with relevant biomolecules such as α-synuclein. This paper presents a new cyclodextrin-8-hydroxyquinoline conjugate and discusses the properties of four cyclodextrins 3-functionalized with 8-hydroxyquinoline as copper(II) chelators and inhibitors of copper-induced synuclein aggregation. The encouraging results establish the potential of cyclodextrin-8-hydroxyquinoline conjugates as chelators for the control of copper toxicity.

LRRK2 knockdown in zebrafish causes developmental defects, neuronal loss, and synuclein aggregation.

Although mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the most common cause of genetic Parkinson's disease, their function is largely unknown. LRRK2 is pleiotropic in nature, shown to be involved in neurodegeneration and in more peripheral processes, including kidney functions, in rats and mice. Recent studies in zebrafish have shown conflicting evidence that removal of the LRRK2 WD40 domain may or may not affect dopaminergic neurons and/or locomotion. This study shows that ∼50% LRRK2 knockdown in zebrafish causes not only neuronal loss but also developmental perturbations such as axis curvature defects, ocular abnormalities, and edema in the eyes, lens, and otic vesicles. We further show that LRRK2 knockdown results in significant neuronal loss, including a reduction of dopaminergic neurons. Immunofluorescence demonstrates that endogenous LRRK2 is expressed in the lens, brain, heart, spinal cord, and kidney (pronephros), which mirror the LRRK2 morphant phenotypes observed. LRRK2 knockdown results further in the concomitant upregulation of β-synuclein, PARK13, and SOD1 and causes β-synuclein aggregation in the diencephalon, midbrain, hindbrain, and postoptic commissure. LRRK2 knockdown causes mislocalization of the Na(+) /K(+) ATPase protein in the pronephric ducts, suggesting that the edema might be linked to renal malfunction and that LRRK2 might be associated with pronephric duct epithelial cell differentiation. Combined, our study shows that LRRK2 has multifaceted roles in zebrafish and that zebrafish represent a complementary model to further our understanding of this central protein. © 2016 Wiley Periodicals, Inc.

Acacetin promotes healthy aging by altering stress response in Caenorhabditis elegans

The progression in lifespan has been associated with elevated intracellular reactive oxygen species (ROS) and oxidative stress level which contributes to development of age related disorders. The discovery of lifespan modulating phytomolecules may promote development of natural therapies against age related afflictions. Acacetin (5,7-dihydroxy-4-methoxyflavone), is a naturally occurring flavonoid known to possess therapeutic properties. To this end, the present study evaluates effect of acacetin (AC) on lifespan, stress and neurotoxicity for the first time by using well-established free living, multicellular Caenorhabditis elegans model system. The 25 μM dose of AC significantly prolonged the mean lifespan of worms by 27.31% in comparison to untreated control and other tested doses of AC. Additionally, AC enhanced stress resistance against oxidative and thermal stress in worms. Furthermore, AC attenuated age related intracellular ROS level, aggregation of age pigment lipofuscin and increased the mean survival in stress hypersensitive mev-1 mutant by 40.5%. AC supplementation also reduced the alpha synuclein aggregation in transgenic worm model of Parkinson’s disease. The enhanced stress resistance, lifespan and alleviation of age related pathology can be attributed to increment in stress modulatory enzymes like superoxide dismutase (SOD) and catalase (CAT) level. Altogether the results suggest AC exposure maintains stress level, health span and extends mean lifespan of C. elegans. The longevity promoting and neuromodulatory effects of AC are mediated by up regulation of the stress response genes sod-3 and gst-4. The present finding gives new insights of natural remedies and their future prospects in developing therapeutic interventions for managing age related diseases.

Synuclein aggregation: When the mouse's away the human will play.

Synuclein aggregation: When the mouse's away the human will play.

What pathological mutants tell us about synuclein aggregation pathways

Misfolding and pathological aggregation of proteins is a hallmark of many neurodegenerative diseases. α‐synuclein (α‐Syn) is one of the major components of the Lewy bodies associated with Parkinson's disease and other neurodegenerative disorders called synucleinopathies. Mutations in the SNCA gene were the first reported links between familial sporadic Parkinson's disease and perturbations at the molecular level.We compared the behavior of α‐synuclein and five pathological mutants (A30P, E46K, H50Q, G51D and A53T). To gain insights into the aggregagtion behavior of these proteins, we developed a method coupling single molecule detection and cell‐free expression to measure precisely the oligomerisation of proteins, without purification and denaturation steps, in completely undisturbed samples. In these conditions, α‐Syn oligomerisation and aggregation is a rapid process that occurred co‐translationally, at nM concentrations.Surprisingly, the pathogenic mutants segregated into two classes: one group forms large aggregates and fibrils while the other tends to form smaller oligomers and fewer fibrils. Strikingly, co‐expression experiments reveal that members from the different groups tend to not interact with each other, both at the fibril and monomer levels. Further biochemical analyses revealed differences of structure between the aggregates. Therefore, the different mutants could provide access to different species formed along the fibrillation path of α‐Syn.We then examined the effects of a variety of chaperones on the aggregation propensity of the different mutants. This uncovers the specificty of the different chaperones for specific species in the aggregation pathway and identifies new therapeutic targets in Parkinson's disease and multiple sceloris atrophy (MSA).

A coarse grained protein model with internal degrees of freedom. Application to α-synuclein aggregation.

Particles in simulations are traditionally endowed with fixed interactions. While this is appropriate for particles representing atoms or molecules, objects with significant internal dynamics--like sequences of amino acids or even an entire protein--are poorly modelled by invariable particles. We develop a highly coarse grained polymorph patchy particle with the ultimate aim of simulating proteins as chains of particles at the secondary structure level. Conformational changes, e.g., a transition between disordered and β-sheet states, are accommodated by internal coordinates that determine the shape and interaction characteristics of the particles. The internal coordinates, as well as the particle positions and orientations, are propagated by Brownian Dynamics in response to their local environment. As an example of the potential offered by polymorph particles, we model the amyloidogenic intrinsically disordered protein α-synuclein, involved in Parkinson's disease, as a single particle with two internal states. The simulations yield oligomers of particles in the disordered state and fibrils of particles in the "misfolded" cross-β-sheet state. The aggregation dynamics is complex, as aggregates can form by a direct nucleation-and-growth mechanism and by two-step-nucleation through conversions between the two cluster types. The aggregation dynamics is complex, with fibrils formed by direct nucleation-and-growth, by two-step-nucleation through the conversion of an oligomer and by auto-catalysis of this conversion.

Reducing synuclein accumulation improves neuronal survival after spinal cord injury

Spinal cord injury causes neuronal death, limiting subsequent regeneration and recovery. Thus, there is a need to develop strategies for improving neuronal survival after injury. Relative to our understanding of axon regeneration, comparatively little is known about the mechanisms that promote the survival of damaged neurons. To address this, we took advantage of lamprey giant reticulospinal neurons whose large size permits detailed examination of post-injury molecular responses at the level of individual, identified cells. We report here that spinal cord injury caused a select subset of giant reticulospinal neurons to accumulate synuclein, a synaptic vesicle-associated protein best known for its atypical aggregation and causal role in neurodegeneration in Parkinson's and other diseases. Post-injury synuclein accumulation took the form of punctate aggregates throughout the somata and occurred selectively in dying neurons, but not in those that survived. In contrast, another synaptic vesicle protein, synaptotagmin, did not accumulate in response to injury. We further show that the post-injury synuclein accumulation was greatly attenuated after single dose application of either the “molecular tweezer” inhibitor, CLR01, or a translation-blocking synuclein morpholino. Consequently, reduction of synuclein accumulation not only improved neuronal survival, but also increased the number of axons in the spinal cord proximal and distal to the lesion. This study is the first to reveal that reducing synuclein accumulation is a novel strategy for improving neuronal survival after spinal cord injury.

The Effect of Small Molecules on Early and Late Events of Alpha Synuclein Aggregation in Solution and Inside Living Cells

Parkinson disease (PD) is the most common motor-neurodegenerative disease, affecting 1-2% of the population older than 65 years and 4-5% of those aged over 85 years. PD is a movement disorder, associated with different motor and nonmotor syndromes, arising from degeneration of dopaminergic neurons of substantia nigra. The hallmark of PD is the observation of intracellular protein aggregates, Lewy Bodies (LB). Alpha-synuclein is one of the major components of LB. The heterogeneity of aggregation process and the presence of large number of triggering mechanisms results in the difficulty to devise therapeutic strategies against these toxic inclusions formation. There have been continuing efforts to search for small molecules against alpha synuclein aggregation.With the help of fluorescence correlation spectroscopy (FCS), confocal imaging, FRAP and other biophysical experiments we have studied the early and late events of alpha-synuclein aggregation. Subsequently, we have explored the possibilities of identifying possible small molecules, which would prevent early oligomer formation. For the FCS experiments, we have labeled a single cysteine mutant of alpha synuclein (G132C) using Alexa488Maleimide. Our FCS data have shown significant changes in the conformation of the protein in the presence of an inhibiting small molecule. To obtain additional insights into these molecules, we have studied their effects inside two mammalian model cell lines, namely HeLa and SH-SY5Y. For this purpose, we have transiently transfected the cells using a tetra-cysteine tagged alpha-synuclein, following which the cells have been stained using Flash-EDT2 dye for fluorescence imaging and FRAP studies. The results show that the electrostatic differences between the protein and the small molecules contribute significantly towards the ability of the small molecules to modulate the early and late stages of aggregation.

The C-terminal α-helices of mammalian Hsc70 play a critical role in the stabilization of α-synuclein binding and inhibition of aggregation

Protein misfolding, followed by aggregation and amyloid formation is an underlying pathological hallmark in a number of prevalent diseases, including Parkinson's (PD), Alzheimer's (AD) and Type 2 diabetes (T2D). In the case of PD, the aggregation of α-synuclein protein (α-syn) has been shown to be highly cytotoxic and to play a key role in the death of dopaminergic cells. Thus, inhibition of the aggregation process may be considered as an attractive avenue for therapeutic intervention. In this respect, molecular chaperones, known to promote proper folding of proteins, are able to inhibit protein aggregation thus preventing amyloid formation. In this work, the effect of the constitutively expressed chaperone Hsc70 and its various domains on α-syn aggregation have been investigated using different approaches. The results show that the C-terminal domain alone (residues 386–646) is as efficient in inhibiting α-syn aggregation as the entire Hsc70 protein, by increasing the lag phase for α-syn oligomeric nucleus formation, suggesting that the chaperone interacts with and stabilizes α-syn monomers and/or small aggregates. Deletion of the C-terminal helices (residues 510–646), which are known to play the role of a lid locking target peptide ligands in the peptide-binding site of the chaperone, strongly reduced the efficiency of inhibition of α-syn aggregation indicating that these helices play an essential in stabilizing the interaction between Hsc70 and α-syn. Furthermore, the effects of Hsc70 and its structural domains on aggregation appear to correlate with those on cytotoxicity, by reducing the fraction of α-syn toxic species to various degrees. Together these results suggest a mechanism in which inhibition of synuclein aggregation is the result of monomeric synuclein binding to the chaperone as any monomeric target unfolded protein or peptide binding to the chaperone.

Ion mobility-mass spectrometry-based screening for inhibition of α- synuclein aggregation.

Aberrant protein folding and formation of amyloid fibrils are associated with numerous debilitating human diseases, for which there are currently no suitable therapeutic treatments. For instance, Parkinson's disease is characterised pathologically by the intraneural accumulation of the amyloid protein α- synuclein. In order to search for new therapeutic agents that are effective in preventing the early conformational changes that precede protein aggregation, it is necessary to devise new analytical screening approaches. Here we demonstrate the use of ion mobility-mass spectrometry for screening of molecules capable of inhibiting the misfolding and aggregation of α-synuclein (specifically, the A53T human mutant). Importantly, this assay allows for the analysis of conformational changes that precede aggregation, and therefore is unique in its ability to identify inhibitors working at the earliest stages of amyloid formation. In addition, we use complementary mass spectrometry methods to probe selected protein-ligand interactions responsible for fibril inhibition.

Dimerization propensities of Synucleins are not predictive for Synuclein aggregation

Aggregation and fibril formation of human alpha-Synuclein (αS) are neuropathological hallmarks of Parkinson's disease and other synucleinopathies. The molecular mechanisms of αS aggregation and fibrillogenesis are largely unknown. Several studies suggested a sequence of events from αS dimerization via oligomerization and pre-fibrillar aggregation to αS fibril formation. In contrast to αS, little evidence suggests that γS can form protein aggregates in the brain, and for βS its neurotoxic properties and aggregation propensities are controversially discussed. These apparent differences in aggregation behavior prompted us to investigate the first step in Synuclein aggregation, i.e. the formation of dimers or oligomers, by Bimolecular Fluorescence Complementation in cells. This assay showed some Synuclein-specific limitations, questioning its performance on a single cell level. Nevertheless, we unequivocally demonstrate that all Synucleins can interact with each other in a very similar way. Given the divergent aggregation properties of the three Synucleins this suggests that formation of dimers is not predictive for the aggregation of αS, βS or γS in the aged or diseased brain.

Functional Interactions of Disease‐Linked Disordered Proteins: Alpha‐Synuclein, Tau and Complexin

The proteins alpha‐synuclein and tau are linked to Parkinson's and Alzheimer's disease, respectively, yet little is known about their normal physiological functions, while complexin, a protein more loosely linked to neurological and psychiatric disorders, has several opposing functions whose mechanisms remain poorly understood. All three proteins are completely or highly disordered when free in solution. Structural characterization of different states of alpha‐synuclein has lead us to propose a model in which the protein can bridge between the synaptic vesicle and plasma membranes. This model also has implications for membrane‐induced synuclein aggregation. These topics will be discussed in the context of one of the most recent Parkinson's disease associated synuclein mutations. Tau is normally a microtubule binding protein, but can also bind to membrane surfaces, an interaction that may be either physiologically or pathologically relevant. Recent work in the lab has confirmed that tau‐membrane interactions are mediated by short amphipathic helices, which may also play a role in mediating membrane‐ or microtubule‐induced aggregation. Complexin contains a central helix motif that binds to assembled SNARE complexes, and this interaction is key to two opposing functions, enhancing synchronous exocytosis and inhibiting spontaneous vesicle fusion. Binding of the disordered C‐terminal domain of complexin to membranes is critical for its inhibitory function. We have shown that such binding is highly dependent on membrane curvature, and that a local structural transition to a helical conformation, again necessary for function, only occurs upon binding to highly curved membranes such as synaptic vesicles. Thus complexin's function is regulated by a membrane curvature dependent structural transition.

Parkinson’s - Just another Infectious Disease

Anatomic staging of PD progression suggests that an unidentified neurotropic pathogen in the intestinal lumen triggers abnormal synuclein aggregation that, in turn, initiates a ''prion-like'' process in the enteric nervous system (ENS) eventually achieving access to the central nervous system (CNS) via the vagus nerve (cranial nerve X). This protein aggregation, manifest as , targets and destroys dopamine producing cells resulting in both the classic motor and non-motor symptoms of PD; with the non-motor symptoms predating the motor symptoms by years, or even decades.

Posttranslational modification and mutation of histidine 50 trigger alpha synuclein aggregation and toxicity.

BackgroundAggregation and aggregation-mediated formation of toxic alpha synuclein (aSyn) species have been linked to the pathogenesis of sporadic and monogenic Parkinson’s disease (PD). A novel H50Q mutation of aSyn, resulting in the substitution of histidine by glutamine, has recently been identified in PD patients. We have previously shown that the lipid peroxidation product 4-hydroxy-2-nonenal (HNE) induces the formation of HNE-aSyn adducts, thereby promoting aSyn oligomerization and increasing its extracellular toxicity to human dopaminergic neurons. Intriguingly, we identified histidine 50 (H50) of aSyn as one of the HNE modification target residues. These converging lines of evidence support the hypothesis that changes in H50 via posttranslational modification (PTM) and mutation trigger the formation of aggregated, toxic aSyn species, which interfere with cellular homeostasis. In the present study, we aim to elucidate 1) the role of H50 in HNE-mediated aSyn aggregation and toxicity, and 2) the impact of H50 mutation on aSyn pathology. Besides the PD-related H50Q, we analyze a PD-unrelated control mutation, in which H50 is replaced by an arginine residue (H50R).ResultsAnalysis of HNE-treated aSyn revealed that H50 is the most susceptible residue of aSyn to HNE modification and is crucial for HNE-mediated aSyn oligomerization. Overexpression of aSyn with substituted H50 in H4 neuroglioma cells reduced HNE-induced cell damage, indicating a pivotal role of H50 in HNE modification-induced aSyn toxicity.Furthermore, we showed in vitro that H50Q/R mutations substantially increase the formation of high density and fibrillar aSyn species, and potentiate the oligomerization propensity of aSyn in the presence of a nitrating agent. Cell-based experiments also revealed that overexpression of H50Q aSyn in H4 cells promotes aSyn oligomerization. Importantly, overexpression of both H50Q/R aSyn mutants in H4 cells significantly increased cell death when compared to wild type aSyn. This increase in cell death was further exacerbated by the application of H2O2.ConclusionA dual approach addressing alterations of H50 showed that either H50 PTM or mutation trigger aSyn aggregation and toxicity, suggesting an important role of aSyn H50 in the pathogenesis of both sporadic and monogenic PD.Electronic supplementary materialThe online version of this article (doi:10.1186/s13024-015-0004-0) contains supplementary material, which is available to authorized users.

Nutrient deprivation induces α-synuclein aggregation through endoplasmic reticulum stress response and SREBP2 pathway.

Abnormal accumulation of filamentous α-synuclein (α-syn) in neurons, regarded as Lewy bodies (LBs), are a hallmark of Parkinson disease (PD). Although the exact mechanism(s) underlying LBs formation remains unknown, autophagy and ER stress response have emerged as two important pathways affecting α-syn aggregation. In present study we tested whether cells with the tetracycline-off inducible overexpression of α-syn and accumulating α-syn aggregates can benefit from autophagy activation elicited by nutrient deprivation (ND), since this approach was reported to effectively clear cellular polyglutamine aggregates. We found that nutrient deprivation of non-induced cells did not affect cell viability, but significantly activated autophagy reflected by increasing the level of autophagy marker LC3-II and autophagic flux and decrease of endogenous α-syn. Cells with induced α-syn expression alone displayed autophagy activation in an α-syn dose-dependent manner to reach a level comparable to that found in non-induced, nutrient deprived counterparts. Nutrient deprivation also activated autophagy further in α-syn induced cells, but the extent was decreased with increase of α-syn dose, indicating α-syn overexpression reduces the responsiveness of cells to nutrient deprivation. Moreover, the nutrient deprivation enhanced α-syn aggregations concomitant with significant increase of apoptosis as well as ER stress response, SREBP2 activation and cholesterolgenesis. Importantly, α-syn aggregate accumulation and other effects caused by nutrient deprivation were counteracted by knockdown of SREBP2, treatment with cholesterol lowering agent—lovastatin, or by GRP78 overexpression, which also caused decrease of SREBP2 activity. Similar results were obtained from studies of primary neurons with α-syn overexpression under nutrient deprivation. Together our findings suggested that down-regulation of SREBP2 activity might be a means to prevent α-syn aggregation in PD via reducing cholesterol levels.

Development of targeted therapies for Parkinson's disease and related synucleinopathies

Therapeutic efforts in neurodegenerative diseases have been very challenging, particularly due to a lack of validated and mechanism-based therapeutic targets and biomarkers. The basic idea underlying the novel therapeutic approaches reviewed here is that by exploring the molecular basis of neurodegeneration in a rare lysosomal disease such as Gaucher's disease (GD), new molecular targets will be identified for therapeutic development in common synucleinopathies. Accumulation of α-synuclein plays a key role in the pathogenesis of Parkinson's disease (PD) and other synucleinopathies, suggesting that improved clearance of α-synuclein may be of therapeutic benefit. To achieve this goal, it is important to identify specific mechanisms and targets involved in the clearance of α-synuclein. Recent discovery of clinical, genetic, and pathological linkage between GD and PD offers a unique opportunity to examine lysosomal glucocerebrosidase, an enzyme mutated in GD, for development of targeted therapies in synucleinopathies. While modulation of glucocerebrosidase and glycolipid metabolism offers a viable approach to treating disorders associated with synuclein accumulation, the compounds described to date either lack the ability to penetrate the CNS or have off-target effects that may counteract or limit their capabilities to mediate the desired pharmacological action. However, recent emergence of selective inhibitors of glycosphingolipid biosynthesis and noninhibitory pharmacological chaperones of glycosphingolipid processing enzymes that gain access to the CNS provide a novel approach that may overcome some of the limitations of compounds reported to date. These new strategies may allow for development of targeted treatments for synucleinopathies that affect both children and adults.

Protective and toxic roles of dopamine in Parkinson's disease

The molecular mechanisms causing the loss of dopaminergic neurons containing neuromelanin in the substantia nigra and responsible for motor symptoms of Parkinson's disease are still unknown. The discovery of genes associated with Parkinson's disease (such as alpha synuclein (SNCA), E3 ubiquitin protein ligase (parkin), DJ-1 (PARK7), ubiquitin carboxyl-terminal hydrolase isozyme L1 (UCHL-1), serine/threonine-protein kinase (PINK-1), leucine-rich repeat kinase 2 (LRRK2), cation-transporting ATPase 13A1 (ATP13A), etc.) contributed enormously to basic research towards understanding the role of these proteins in the sporadic form of the disease. However, it is generally accepted by the scientific community that mitochondria dysfunction, alpha synuclein aggregation, dysfunction of protein degradation, oxidative stress and neuroinflammation are involved in neurodegeneration. Dopamine oxidation seems to be a complex pathway in which dopamine o-quinone, aminochrome and 5,6-indolequinone are formed. However, both dopamine o-quinone and 5,6-indolequinone are so unstable that is difficult to study and separate their roles in the degenerative process occurring in Parkinson's disease. Dopamine oxidation to dopamine o-quinone, aminochrome and 5,6-indolequinone seems to play an important role in the neurodegenerative processes of Parkinson's disease as aminochrome induces: (i) mitochondria dysfunction, (ii) formation and stabilization of neurotoxic protofibrils of alpha synuclein, (iii) protein degradation dysfunction of both proteasomal and lysosomal systems and (iv) oxidative stress. The neurotoxic effects of aminochrome in dopaminergic neurons can be inhibited by: (i) preventing dopamine oxidation of the transporter that takes up dopamine into monoaminergic vesicles with low pH and dopamine oxidative deamination catalyzed by monoamino oxidase (ii) dopamine o-quinone, aminochrome and 5,6-indolequinone polymerization to neuromelanin and (iii) two-electron reduction of aminochrome catalyzed by DT-diaphorase. Furthermore, dopamine conversion to NM seems to have a dual role, protective and toxic, depending mostly on the cellular context. Dopamine oxidation to dopamine o-quinone, aminochrome and 5,6-indolequinone plays an important role in neurodegeneration in Parkinson's disease since they induce mitochondria and protein degradation dysfunction; formation of neurotoxic alpha synuclein protofibrils and oxidative stress. However, the cells have a protective system against dopamine oxidation composed by dopamine uptake mediated by Vesicular monoaminergic transporter-2 (VMAT-2), neuromelanin formation, two-electron reduction and GSH-conjugation mediated by Glutathione S-transferase M2-2 (GSTM2).

Biophysical groundwork as a hinge to unravel the biology of α-synuclein aggregation and toxicity

Alpha-synuclein (aS) and its aggregation properties are central in the development and spread of Parkinson's disease. Point mutations and multiplications of the SNCA gene encoding aS cause autosomal dominant forms of the disorder. Moreover, protein inclusions found in the surviving neurons of parkinsonian brains consist mainly of a fibrillar form of aS. Aggregates of aS, which form a transient, complex and heterogeneous ensemble, participate in a wide variety of toxic mechanisms that may be amplified by aS spreading among neighbouring neurons. Recently, significant effort has been directed into the study of the aS aggregation process and the impact of aS aggregates on neuron survival. In this review, we present state-of-the-art biophysical studies on the aS aggregation process in vitro and in cellular models. We comprehensively review the new insights generated by the recent biophysical investigations, which could provide a solid basis from which to design future biomedical studies. The diverse cellular models of aS toxicity and their potential use in the biophysical investigation are also discussed.