α-synuclein aggregation and its modulation
Introduction
Parkinson’s disease (PD) is a neurodegenerative disease in the elderly affecting millions of people worldwide [1]. PD is characterized by the progressive degeneration of dopamine producing neurons (dopaminergic neurons) in the substantia nigra region of the brain, which results in impaired motor functions, displayed as compromised postural reflexes, muscular rigidity and bradykinesia in PD patients [2], [3], [4], [5]. Despite several years of intense and inter disciplinary research, the exact mechanism of selective degeneration of dopaminergic neurons is not well known probably due to the multi factorial nature of the disease. Complexity in PD arises from the involvement of several environmental and genetic risk factors, which worsen neurodegeneration and PD pathology. Interestingly, despite strong genetic contribution, PD is mostly sporadic and familial forms of the disease represent only a minor part (<10%) of all cases [6]. However, irrespective of the origin of disease (sporadic/familial), the main pathological hallmark in most PD cases is the presence of Lewy bodies (LBs) and Lewy neurites (LNs) in the PD patients’ brains [7]. These LBs and LNs are mainly composed of the aggregated form (amyloid fibrils) of α-synuclein (α-Syn), a highly conserved presynaptic 140 amino acid protein [7], [8]. This finding suggests that aggregation of α-Syn into amyloid fibrils and its subsequent accumulation in LBs and LNs (Fig. 1) is a central event in the pathogenesis of both sporadic and familial PD. In connection with this idea, several pioneer studies suggest that duplication, triplication and several mutations in the SNCA gene on chromosome 4q21-23, which encodes for the α-Syn protein, are associated with early onset familial form of PD [5], [9], [10], [11], [12], [13], [14], [15], [16], [17].
The hypothesis that accelerated fibrillation of α-Syn may worsen PD pathology and disease progression was inspired from the seminal discovery of two α-Syn mutants (A53T and E46K) with increased fibrillation propensity in vitro, which are associated with familial PD [18], [19], [20]. Furthermore, a recently discovered mutation of α-Syn (H50Q) linked to familial form of PD is also shown to accelerate the fibrillation rate of α-Syn [16], [17], [21]. On the contrary, several other mutations (A30P, G51D, and A53E) associated with familial forms of PD have been shown to decrease the fibrillation rate of α-Syn [15], [22], [23], [24], [25]. This suggests that the fibrillation rate of α-Syn is not directly correlated to disease pathogenesis. This complexity may be resolved to some extent by monitoring the effect of familial α-Syn mutations on oligomerization rate rather than the fibrillation rate. Evaluating oligomerization kinetics may be a promising approach since several studies have claimed that pre fibrillar α-Syn oligomers are probably the most potent toxic entities responsible for neuronal death in PD [26]. Consistent with this observation, it is well established that despite their differential effects on fibril formation rate, both mutations (A53T and A30P) accelerate oligomer formation, which ultimately leads to early onset PD pathology [27]. Moreover, one could explain the association of the H50Q mutation with late-onset PD because it is a faster fibril-forming mutant, thus, accumulation of toxic prefibrillar oligomers is less likely due to their rapid conversion into fibrils. However, this rationale does not hold true for E46K mutant, which also has faster fibril formation rate resulting in lesser accumulation of toxic pre-fibrillar oligomers but is still associated with early onset familial PD. Therefore, establishing a correlation between α-Syn aggregation rate and PD progression is really a puzzling task. Furthermore, favored oligomers formation and its subsequent slow fibril conversion rate are only well established for the A30P mutant and are either not clearly seen or not studied well yet for other slow fibril forming mutants (G51D and A53E).
Some of these contradictions may be partially sorted out by considering the fact that PD is a multifactorial disease, where several environmental factors such as exposure to metal ions and pesticides as well as dietary factors and altered cellular micro environment may also affect α-Syn aggregation in vivo and in turn affect PD pathology [28]. For example, the properties and functions of α-Syn would depend on the extent of its interaction with membrane, post-translational modifications, local folding in cellular crowded milieu, which will eventually affect the extent of aggregation and production toxic species of α-Syn. Additionally, the interaction/co-aggregation of α-Syn with other proteins and biomolecules such as polyamines could also be equally important for α-Syn mediated PD pathogenesis [28].
Along with these factors, various physical and chemical parameters such as aggregation surface, pH, ionic strength, metal ions, and temperature are also shown to affect the α-Syn aggregation in vitro [28], [29], [30], [31]. Thus, correlating α-Syn aggregation in test tubes with disease progression in vivo becomes a daunting task. As research on the factors associated with PD and/or modulating α-Syn aggregation is subject of active investigation by huge number of laboratories, reviewing all these factors is beyond the scope of this review. However, we tried to provide some overview of research on α-Syn aggregation pathways and its modulation by cellular factors and selected environmental factors that may modulate PD pathogenesis.
Section snippets
α-Syn: a brief history and its biophysical properties
α-Syn is a protein, which belongs to the synuclein protein family found in vertebrates [8]. Synuclein was first identified in the year 1988 as a protein localized in the synaptic vesicles and nucleus of the cholinergic neurons in Pacific electric ray (Torpedo californica) [32]. Based upon its synaptic and nuclear localization, the protein was named as synuclein [8], [32]. The synucleins were present as three major forms (molecular mass ∼17.5, ∼18.5 and ∼20 kDa) in the torpedo central nervous
α-Syn aggregation in Parkinson’s disease (PD)
PD is a chronic neurodegenerative disorder, which affects mainly individuals at older ages. Initially, PD was considered solely as a sporadic disease with unknown cause and without any genetic component [51], [52]. However, in the late 1990s, this notion was challenged with the discovery of numerous genes, which were reported to be linked with rare familial forms of PD [53]. Nevertheless a minor part (<10%) represents familial forms of PD, whereas the rest 90% are sporadic cases [6]. The first
Aggregation and amyloid formation pathway of α-Syn in vitro
α-Syn is natively unstructured in physiological pH [39]. Upon incubation it aggregates into cross-β-sheet rich amyloid fibrils [39]. However, the unstructured monomeric native structure of α-Syn was contradicted by Selkoe and co-workers who suggested that the native structure of α-Syn is helix-rich tetrameric form [66]. Studies by Wang et al. also supported the helix-rich tetrameric conformation of α-Syn existing under physiological conditions [67]. The idea that α-Syn exists as helix-rich
Effect of familial PD-associated mutations on α-Syn oligomer and fibril formation in vitro
Understanding how the familial PD-associated mutations affect the aggregation properties of α-Syn will help to delineate the correlation between α-Syn aggregation and PD pathogenesis. The first two familial PD-associated mutations of α-Syn, A53T and A30P were discovered in the years 1997 and 1998, respectively. The in vitro aggregation studies for WT α-Syn and its familial PD associated mutants, A53T and A30P were performed by Lansbury and coworkers using various biophysical methods [18]. The
Role of polyamines in modulating α-Syn aggregation
Polyamines are aliphatic polycations, known to be mysterious modulators of molecular and cellular functions in humans [81], [82]. They are present in the neuronal cells [83], [84] with cellular concentration of spermine around ∼1 mM and comparatively higher concentrations reported for putrescine and spermidine [82], [85], [86]. At physiological pH, polyamines are cationic in nature and thus interact with negatively charged biomolecules like DNA, RNA, ATP, phospholipids and certain proteins [87],
Effect of glycosaminoglycans (GAGs) on α-synuclein aggregation
GAGs are charged polymers of amino sugar moiety and are important components of extra cellular matrix. They can be found either in the form of proteoglycans (coupled with protein cores of extracellular matrix) or as free macromolecules on the cell surfaces. Strikingly, they have been found to be closely associated with various amyloid diseases [100], [101], [102]. In support of this fact, substantial amount of studies have suggested that GAGs modulate the aggregation and amyloid formation of
Effect of chaperones on α-Syn aggregation
Preserving the structural identity of a protein is critical for the cell to maintain protein homeostasis. For this important role, cells recruit a group of ubiquitous highly conserved class of molecules known as chaperones. These include a family of heat shock proteins (Hsps) and many other co-chaperones, which are expressed constitutively and show increased expression under stress conditions [125]. Hsps are further classified on the basis of the molecular mass of their monomer subunit i.e
Effect of membrane on α-Syn aggregation
α-Syn is natively unstructured at physiological pH [28]. However, it can adopt helical conformation when it is bound to membranes or in membrane mimicking conditions [158], [159], [160]. α-Syn has been found to remain in equilibrium between its cytosolic free form and membrane bound form [161], [162], [163]. It has been estimated that ∼15% α-Syn remains in membrane bound within synaptic termini [163]. Upon binding to lipid vesicles with N-terminus, α-Syn forms broken helix [164]. Interaction of
Effect of α-Syn modification in its aggregation
α-Syn aggregation is a highly co-operative process where a subpopulation of post-translationally modified (PTM) protein has a significant effect on overall kinetics and amyloid formation. Several PTMs have been found to be associated with α-Syn [181], [182], [183], [184], [185], [186], [187]. Indeed LBs isolated from PD patients, transgenic models of PD and other synucleinopathies have been found to contain post translationally modified α-Syn [184], [185], [188], [189]. The major PTMs for α-Syn
Effect of environmental factors on α-Syn aggregation
Since decades PD has been considered as a sporadic disorder where non-genetic environmental factors played a major role in progression of the disease [51], [52]. This conviction was further supported in part in 1980s with the occurrence of Parkinsonism in humans caused by intravenous administration of an illicit opioid analgesic drugs, where 1-methyl 4-phenyl-1,2,3,6- tetrahydropyridine (MPTP) was formed as an accidental side product [238]. MPTP induced disease symptoms in these individuals
Conclusion and future directions
α-Syn aggregation and amyloid formation play a central role in PD pathogenesis [1]. The focal point of the molecular mechanism underlying PD pathogenesis became α-Syn centric after its discovery in 1997. Subsequently, in vitro studies along with various animal and cell model data suggest that aggregation of α-Syn into amyloid fibrils may be a pathogenic mechanism for PD [259], [260]. Further, it was hypothesized that oligomeric intermediates formed during fibrillation pathway of α-Syn are the
Conflict of interest
The authors declare no conflicts of interest.
Acknowledgement
The authors thank ICMR (5/20/9(Bio)/2011-NCD-I) Govt. of India and IRCC, IIT Bombay for the funding sources. D.G. and S.M. acknowledge UGC, Govt. of India, S.S. acknowledges SPMF, CSIR, Govt. of India, P.KS acknowledges CSIR, Govt. of India for their fellowships. The authors also acknowledge Nina Merkel for critically reading the manuscript.
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