Detecting Morphologically Distinct Oligomeric Forms of α-Synuclein

Emadi, Sharareh; Kasturirangan, Srinath; Wang, Min S.; Schulz, Philip; Sierks, Michael R.

doi:10.1074/jbc.m806559200

Cited by 92 publications

(130 citation statements)

References 65 publications

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“…Methods to detect morphologically distinct oligomeric forms of AS have been described [413]. Elevated levels of soluble AS oligomers were found in post-mortem extracts of PD [414] and DLB brains [415,416].…”

Section: α-Synuclein and Neurodegenerationmentioning

confidence: 99%

The role of α-synuclein in neurodegeneration — An update

Jellinger¹

2012

Translational Neuroscience

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Genetic, neuropathological and biochemical evidence implicates α-synuclein, a 140 amino acid presynaptic neuronal protein, in the pathogenesis of Parkinson's disease and other neurodegenerative disorders. The aggregated protein inclusions mainly containing aberrant α-synuclein are widely accepted as morphological hallmarks of α-synucleinopathies, but their composition and location vary between disorders along with neuronal networks affected. α-Synuclein exists physiologically in both soluble and membran-bound states, in unstructured and α-helical conformations, respectively, while posttranslational modifications due to proteostatic deficits are involved in β-pleated aggregation resulting in formation of typical inclusions. The physiological function of α-synuclein and its role linked to neurodegeneration, however, are incompletely understood. Soluble oligomeric, not fully fibrillar α-synuclein is thought to be neurotoxic, main targets might be the synapse, axons and glia. The effects of aberrant α-synuclein include alterations of calcium homeostasis, mitochondrial dysfunction, oxidative and nitric injuries, cytoskeletal effects, and neuroinflammation. Proteasomal dysfunction might be a common mechanism in the pathogenesis of neuronal degeneration in α-synucleinopathies. However, how α-synuclein induces neurodegeneration remains elusive as its physiological function. Genome wide association studies demonstrated the important role for genetic variants of the SNCA gene encoding α-synuclein in the etiology of Parkinson's disease, possibly through effects on oxidation, mitochondria, autophagy, and lysosomal function. The neuropathology of synucleinopathies and the role of α-synuclein as a potential biomarker are briefly summarized. Although animal models provided new insights into the pathogenesis of Parkinson disease and multiple system atrophy, most of them do not adequately reproduce the cardinal features of these disorders. Emerging evidence, in addition to synergistic interactions of α-synuclein with various pathogenic proteins, suggests that prionlike induction and seeding of α-synuclein could lead to the spread of the pathology and disease progression. Intervention in the early aggregation pathway, aberrant cellular effects, or secretion of α-synuclein might be targets for neuroprotection and disease-modifying therapy.

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Section: α-Synuclein and Neurodegenerationmentioning

confidence: 99%

The role of α-synuclein in neurodegeneration — An update

Jellinger¹

2012

Translational Neuroscience

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“…This intrabody decreased the toxicity and aggregation of α-Syn in vitro [80]. Another intrabody, 10H, recognized a later-stage oligomer and inhibited toxicity induced by extracellular α-Syn, and also inhibited in vitro aggregation of α-Syn [81]. Recently Guilliams et al [82] reported additional anti-α-Syn nanobody, NbSyn87, obtained by immunization of llama with α-Syn.…”

Section: Intrabodies Targeted To Conformational α-Syn Epitopesmentioning

confidence: 99%

“…Addition of the acidic PEST peptide to four anti-α-Syn intrabodies improved the solubility for all 4, while still retaining sufficient intracellular levels of intrabody-PEST protein. In addition, the highest affinity, but least soluble of the parent constructs, the NAC-specific [81] Human scFv Oligomeric α-Syn Tomlinson I and J antibody library D10 [76] Human scFv Pan-specific Human scFv phage Griffin I library VH14 [74] Human nanobody Part of hydrophobic NAC of α-Syn Human nonimmune yeast surface display library NAC32 [74] Human scFv Part of hydrophobic NAC of α-Syn Human nonimmune yeast surface display library 6E [28] Human scFv Fibrillar α-Syn Tomlinson I and J antibody library NbSyn2 [79] Camelid VHH Monomeric α-Syn Phage display ScFv = single chain Fv; VHH = small heavy-chain-only antibody fragments; NAC = nonamyloid component human heavy-chain only nanobody, VH14, acquired the capacity to degrade α-Syn~GFP [90]. Thus, this bifunctional approach to the intrabody engineering can be used successfully to target and direct the molecule of choice to the required compartment, and shows great promise of targeting the earliest stages of neurodegeneration.…”

Section: Synmentioning

confidence: 99%

“…Intrabody approaches can therefore include intrabodies targeted to reduce the monomeric α-Syn or those targeted to a particular neurotoxic conformation of aggregates. Emadi et al [80,81] have combined phage display and atomic force microscopy technique to isolate intrabodies that bind to different types of oligomeric α-Syn targets. Atomic force microscopy was used to ascertain the morphology of α-Syn target and to monitor the process of panning.…”

Section: Intrabodies Targeted To Conformational α-Syn Epitopesmentioning

confidence: 99%

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Intrabodies as Neuroprotective Therapeutics

Messer

Joshi

2013

Neurotherapeutics

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The process of misfolding of proteins that can trigger a pathogenic cascade leading to neurodegenerative diseases largely originates intracellularly. It is possible to harness the specificity and affinity of antibodies to counteract either protein misfolding itself, or the aberrant interactions and excess stressors immediately downstream of the primary insult. This review covers the emerging field of engineering intracellular antibody fragments, intrabodies and nanobodies, in neurodegeneration. Huntington's disease has provided the clearest proof of concept for this approach. The model systems and readouts for this disorder power the studies, and the potential to intervene therapeutically at early stages in known carriers with projected ages of onset increases the chances of meaningful clinical trials. Both single-chain Fv and single-domain nanobodies have been identified against specific targets; data have allowed feedback for rational design of bifunctional constructs, as well as target validation. Intrabodies that can modulate the primary accumulating protein in Parkinson's disease, alphasynuclein, are also reviewed, covering a range of domains and conformers. Recombinant antibody technology has become a major player in the therapeutic pipeline for cancer, infectious diseases, and autoimmunity. There is also tremendous potential for applying this powerful biotechnology to neurological diseases.Keywords Intrabodies . Huntington's disease . Immunotherapy . Intrabody-PEST fusions . α-synuclein . Parkinson's disease . Polyglutamine Biological Antibody Therapies for Misfolding ProteinsThe problem of misfolding proteins appears to underlie a significant fraction of neurodegenerative diseases. Protein homeostasis, often referred to as proteostasis, is an especially critical process in postmitotic neurons. The balance of pathways for protein folding, interactions, intracellular localizations, and degradation is a complex one, and can be dysregulated by combinations of mutations, environmental stressors, and aging. We have taken the neurotherapeutic approach of normalizing or manipulating proteostasis using engineered intracellular antibody fragments-intrabodies-that bind with high specificity to selected targets. These intrabodies can be selected and manipulated as genes, allowing the full spectrum of genetic engineering to produce multifunctional constructs that can alter the folding, interactions, intracellular localization, and turnover kinetics/ levels of the target protein. Intrabodies are also valuable molecular tools, which can be used to dissect the functional and pathogenic motifs in these proteins and that could be useful for the development of alternative therapeutics. In this review, we will cover the development of this approach for Huntington's disease (HD), which has a well-described target, evaluating effects in several cell culture and in vivo systems with strong readouts for therapeutic efficacy, and advances in engineering anti-HD intrabodies. We also cover a small, but growing, literature...

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“…Recent studies have implicated small soluble oligomeric and protofibrillar forms of αS as the most neurotoxic species. [9][10][11][12] The influence of nanoparticles (NPs) on amyloid formation is of great interest due to their small sizes and high surface area-to-volume ratios. The small size of NPs allows them to access almost all parts of the human body; they even pass through the blood-brain barrier (BBB), which is the homeostatic defense mechanism of the brain.…”

Section: Introductionmentioning

confidence: 99%

Investigation of the effects of carbon-based nanomaterials on A53T alpha-synuclein aggregation using a whole-cell recombinant biosensor

Mohammadi¹,

Nikkhah²,

Hosseinkhani³

2017

IJN

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The aggregation of alpha-synuclein (αS), natively unstructured presynaptic protein, is a crucial factor leading to the pathogenesis of Parkinson’s disease (PD) and other related disorders. Recent studies have shown prefibrillar and oligomeric intermediates of αS as toxic to the cells. Herein, split-luciferase complementation assay is used to design a “signal-on” biosensor to monitor oligomerization of A53T αS inside the cells. Then, the effect of carbon-based nanomaterials, such as graphene quantum dots (GQDs) and graphene oxide quantum dots (GOQDs), on A53T αS oligomerization in vitro and in living cells is investigated. In this work, for the first time, it was found that GQDs at a concentration of 0.5 μg/mL can promote A53T αS aggregation by shortening the nucleation process, which is the key rate-determining step of fibrillation, thereby making a signal-on biosensor. While these nanomaterials may cross the blood–brain barrier because of their small sizes, the interaction between αS and GQDs may contribute to PD etiology.

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Detecting Morphologically Distinct Oligomeric Forms of α-Synuclein

Cited by 92 publications

References 65 publications

The role of α-synuclein in neurodegeneration — An update

The role of α-synuclein in neurodegeneration — An update

Intrabodies as Neuroprotective Therapeutics

Investigation of the effects of carbon-based nanomaterials on A53T alpha-synuclein aggregation using a whole-cell recombinant biosensor

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