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Mutations in the NOTCH3 gene can lead to small-vessel disease in humans, including the well-characterized cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), a condition caused by NOTCH3 mutations altering the number of cysteine residues in the extracellular domain of Notch3. Growing evidence indicates that other types of mutations in NOTCH3, including cysteine-sparing missense mutations or frameshift and premature stop codons, can lead to small-vessel disease phenotypes of variable severity or penetrance. There are currently no disease-modifying therapies for small-vessel disease, including those associated with NOTCH3 mutations. A deeper understanding of underlying molecular mechanisms and clearly defined targets are needed to promote the development of therapies. This review discusses two key pathophysiological mechanisms believed to contribute to the emergence and progression of small-vessel disease associated with NOTCH3 mutations: abnormal Notch3 aggregation and aberrant Notch3 signaling. This review offers a summary of the literature supporting and challenging the relevance of these mechanisms, together with an overview of available preclinical experiments derived from these mechanisms. It highlights knowledge gaps and future research directions. In view of recent evidence demonstrating the relatively high frequency of NOTCH3 mutations in the population, and their potential role in promoting small-vessel disease, progress in the development of therapies for NOTCH3-associated small-vessel disease is urgently needed.

Accumulation of microtubule-associated tau protein is thought to cause neuron loss in a group of neurodegenerative diseases called tauopathies. In diseased brains, tau molecules adopt pathological structures that propagate into insoluble forms with disease-specific patterns. Several types of posttranslational modifications in tau are known to modulate its aggregation propensity in vitro, but their influence on tau accumulation and toxicity at the whole-organism level has not been fully elucidated. Herein, we utilized a series of transgenic Drosophila models to compare systematically the toxicity induced by five tau constructs with mutations or deletions associated with aggregation, including substitutions at seven disease-associated phosphorylation sites (S7A and S7E), deletions of PHF6 and PHF6* sequences (ΔPHF6 and ΔPHF6*), and substitutions of cysteine residues in the microtubule binding repeats (C291/322A). We found that substitutions and deletions resulted in different patterns of neurodegeneration and accumulation, with C291/322A having a dramatic effect on both tau accumulation and neurodegeneration. These cysteines formed disulfide bonds in mouse primary cultured neurons and in the fly retina, and stabilized tau proteins. Additionally, they contributed to tau accumulation under oxidative stress. We also found that each of these cysteine residues contributes to the microtubule polymerization rate and microtubule levels at equilibrium, but none of them affected tau binding to polymerized microtubules. Since tau proteins expressed in the Drosophila retina are mostly present in the early stages of tau filaments self-assembly, our results suggest that disulfide bond formation by these cysteine residues could be attractive therapeutic targets.

Missense mutations in p53 are severely deleterious and occur in over 50% of all human cancers. The majority of these mutations are located in the inherently unstable DNA-binding domain (DBD), many of which destabilize the domain further and expose its aggregation-prone hydrophobic core, prompting self-assembly of mutant p53 into inactive cytosolic amyloid-like aggregates. Screening an oligopyridylamide library, previously shown to inhibit amyloid formation associated with Alzheimer’s disease and type II diabetes, identified a tripyridylamide, ADH-6, that abrogates self-assembly of the aggregation-nucleating subdomain of mutant p53 DBD. Moreover, ADH-6 targets and dissociates mutant p53 aggregates in human cancer cells, which restores p53’s transcriptional activity, leading to cell cycle arrest and apoptosis. Notably, ADH-6 treatment effectively shrinks xenografts harboring mutant p53, while exhibiting no toxicity to healthy tissue, thereby substantially prolonging survival. This study demonstrates the successful application of a bona fide small-molecule amyloid inhibitor as a potent anticancer agent., Amyloid aggregation of mutant p53 contributes to its loss of tumor suppressor function and oncogenic gain-of-function. Here, the authors use a protein mimetic to abrogate mutant p53 aggregation and rescue p53 function, which inhibits cancer cell proliferation in vitro and halts tumor growth in vivo.

Background: Alzheimer’s disease (AD) is a progressive neurodegenerative disease. One of the pathologies of AD is the accumulation of amyloid-β (Aβ) to form senile plaques, leading to a decline in cognitive ability and a lack of learning and memory. However, the cause leading to Aβ aggregation is not well understood. Dendritic cell factor 1 (Dcf1) shows a high expression in the entorhinal cortex neurons and neurofibrillary tangles in AD patients. Objective: Our goal is to investigate the effect of Dcf1 on Aβ aggregation and memory deficits in AD development. Methods: The mouse and Drosophila AD model were used to test the expression and aggregation of Aβ, senile plaque formation, and pathological changes in cognitive behavior during dcf1 knockout and expression. We finally explored possible drug target effects through intracerebroventricular delivery of Dcf1 antibodies. Results: Deletion of Dcf1 resulted in decreased Aβ42 level and deposition, and rescued AMPA Receptor (GluA2) levels in the hippocampus of APP-PS1-AD mice. In Aβ42 AD Drosophila, the expression of Dcf1 in Aβ42 AD flies aggravated the formation and accumulation of senile plaques, significantly reduced its climbing ability and learning-memory. Data analysis from all 20 donors with and without AD patients aged between 80 and 90 indicated a high-level expression of Dcf1 in the temporal neocortex. Dcf1 contributed to Aβ aggregation by UV spectroscopy assay. Intracerebroventricular delivery of Dcf1 antibodies in the hippocampus reduced the area of senile plaques and reversed learning and memory deficits in APP-PS1-AD mice. Conclusion: Dcf1 causes Aβ-plaque accumulation, inhibiting dcf1 expression could potentially offer therapeutic avenues.

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of human COVID-19, not only causes flu-like symptoms and gut microbiome complications but a large number of infected individuals also experience a host of neurological symptoms including loss of smell and taste, seizures, difficulty concentrating, decreased alertness, and brain inflammation. Although SARS-CoV-2 infections are not more prevalent in Parkinson's disease patients, a higher mortality rate has been reported not only associated with older age and longer disease duration, but also through several mechanisms, such as interactions with the brain dopaminergic system and through systemic inflammatory responses. Indeed, a number of the neurological symptoms seen in COVID-19 patients, as well as the alterations in the gut microbiome, are also prevalent in patients with Parkinson's disease. Furthermore, biochemical pathways such as oxidative stress, inflammation, and protein aggregation have shared commonalities between Parkinson's disease and COVID-19 disease progression. In this review, we describe and compare the numerous similarities and intersections between neurodegeneration in Parkinson's disease and RNA viral infections, emphasizing the current SARS-CoV-2 global health crisis.

The post-infection of COVID-19 includes a myriad of neurologic symptoms including neurodegeneration. Protein aggregation in brain can be considered as one of the important reasons behind the neurodegeneration. SARS-CoV-2 Spike S1 protein receptor binding domain (SARS-CoV-2 S1 RBD) binds to heparin and heparin binding proteins. Moreover, heparin binding accelerates the aggregation of the pathological amyloid proteins present in the brain. In this paper, we have shown that the SARS-CoV-2 S1 RBD binds to a number of aggregation-prone, heparin binding proteins including Aβ, α-synuclein, tau, prion, and TDP-43 RRM. These interactions suggests that the heparin-binding site on the S1 protein might assist the binding of amyloid proteins to the viral surface and thus could initiate aggregation of these proteins and finally leads to neurodegeneration in brain. The results will help us to prevent future outcomes of neurodegeneration by targeting this binding and aggregation process.

The ability of viruses to evolve several orders of magnitude faster than their host cells has enabled them to exploit host cellular machinery by selectively recruiting multiprotein complexes (MPCs) for their catalyzed assembly and replication. This hijacking may depend on alternative, 'moonlighting' functions of host proteins that deviate from their canonical functions thereby inducing cellular pathology. Here, we posit that if virus-induced cellular pathology is similar to that of other, unknown (non-viral) causes, the identification and molecular characterization of the host proteins involved in virus-mediated cellular pathology can be leveraged to decipher the non-viral disease-relevant mechanisms. We focus on how virus-induced aberrant proteostasis and protein aggregation resemble the cellular pathology of sporadic neurodegenerative diseases (NDs) and how this can be exploited for drug discovery.

The propagation of conformational strains by templated seeding is central to the prion concept. Seeded assembly of α‐synuclein into filaments is believed to underlie the prion‐like spreading of protein inclusions in a number of human neurodegenerative diseases, including Parkinson's disease, dementia with Lewy bodies (DLB) and multiple system atrophy (MSA). We previously determined the atomic structures of α‐synuclein filaments from the putamen of five individuals with MSA. Here, we used filament preparations from three of these brains for the in vitro seeded assembly of recombinant human α‐synuclein. We find that the structures of the seeded assemblies differ from those of the seeds, suggesting that additional, as yet unknown, factors play a role in the propagation of the seeds. Identification of these factors will be essential for understanding the prion‐like spreading of α‐synuclein proteinopathies., The assembly of certain proteins into amyloids underlies multiple neurodegenerative diseases. The spreading of these assemblies through the brain is thought to occur through a prion‐like mechanism. We used filaments extracted from multiple system atrophy brains to seed recombinant α‐synuclein. The resulting structures differ from those of the seeds, indicating that seeded assembly does not necessarily replicate the seed structures.

α-Synuclein aggregation at the synapse is an early event in Parkinson’s disease and is associated with impaired striatal synaptic function and dopaminergic neuronal death. The cysteine string protein (CSPα) and α-synuclein have partially overlapping roles in maintaining synaptic function and mutations in each cause neurodegenerative diseases. CSPα is a member of the DNAJ/HSP40 family of co-chaperones and like α-synuclein, chaperones the SNARE complex assembly and controls neurotransmitter release. α-Synuclein can rescue neurodegeneration in CSPαKO mice. However, whether α-synuclein aggregation alters CSPα expression and function is unknown. Here we show that α-synuclein aggregation at the synapse is associated with a decrease in synaptic CSPα and a reduction in the complexes that CSPα forms with HSC70 and STGa. We further show that viral delivery of CSPα rescues in vitro the impaired vesicle recycling in PC12 cells with α-synuclein aggregates and in vivo reduces synaptic α-synuclein aggregates increasing monomeric α-synuclein and restoring normal dopamine release in 1-120hαSyn mice. These novel findings reveal a mechanism by which α-synuclein aggregation alters CSPα at the synapse, and show that CSPα rescues α-synuclein aggregation-related phenotype in 1-120hαSyn mice similar to the effect of α-synuclein in CSPαKO mice. These results implicate CSPα as a potential therapeutic target for the treatment of early-stage Parkinson’s disease., CSPα and αsynuclein (αsyn) are involved in synaptic function and neurotransmitter release via parallel pathways. Caló et al. show that αsyn aggregation at the synapse is associated with a reduction in CSPα. An increase in CSPα reduces synaptic αsyn aggregates, rescuing impaired striatal dopamine release in αsyn transgenic mice.

Mechanical stress induced by contractions constantly threatens the integrity of muscle Z-disc, a crucial force-bearing structure in striated muscle. The PDZ-LIM proteins have been proposed to function as adaptors in transducing mechanical signals to preserve the Z-disc structure, however the underlying mechanisms remain poorly understood. Here, we show that LDB3, a well-characterized striated muscle PDZ-LIM protein, modulates mechanical stress signaling through interactions with the mechanosensing domain in filamin C, its chaperone HSPA8, and PKCα in the Z-disc of skeletal muscle. Studies of Ldb3Ala165Val/+ mice indicate that the myopathy-associated LDB3 p.Ala165Val mutation triggers early aggregation of filamin C and its chaperones at muscle Z-disc before aggregation of the mutant protein. The mutation causes protein aggregation and eventually Z-disc myofibrillar disruption by impairing PKCα and TSC2-mTOR, two important signaling pathways regulating protein stability and disposal of damaged cytoskeletal components at a major mechanosensor hub in the Z-disc of skeletal muscle., Pathak et al. identify LDB3, a striated muscle PDZ-LIM protein, as a signaling adaptor in a major mechanosensor assembly through interactions with filamin C, HSPA8, and PKCα. When LDB3 is mutated, PKCα and TSC2-mTOR mediated homeostasis is impaired, leading to protein aggregation myopathy.

In the central nervous system, microglia are regarded as the main cells responsible for phagocytosis, contributing to neural circuit refinement and homeostasis through synapse elimination. However, recent findings have shown that astrocytes also actively participate in synapse homeostasis through phagocytosing synapses, neuronal debris, axonal mitochondria, and pathological protein aggregates. In addition, it has been also suggested that astrocytes may regulate microglial phagocytosis by secreting molecules such as IL-33 and C3. Here, we have introduced key findings regarding direct and indirect astrocyte-mediated phagocytosis in CNS development, the sleep/wake cycle, and aging. We have also discussed current information about reactive astrocytes and their phagocytic function in the diseased brain, focusing on ischemia and Alzheimer's disease. Through this review, we aim to provide an overview of the current status as well as future perspectives regarding the important role of astrocytic control of phagocytosis.

Genetic cardiomyopathy is caused by mutations in various genes. The accumulation of potentially proteotoxic mutant protein aggregates due to insufficient autophagy is a possible mechanism of disease development. The objective of this study was to investigate the distribution in the myocardium of such aggregates in relation to specific pathogenic genetic mutations in cardiomyopathy hearts. Hearts from 32 genetic cardiomyopathy patients, 4 non‐genetic cardiomyopathy patients and 5 controls were studied. Microscopic slices from an entire midventricular heart slice were stained for p62 (sequestosome‐1, marker for aggregated proteins destined for autophagy). The percentage of cardiomyocytes with p62 accumulation was higher in cardiomyopathy hearts (median 3.3%) than in healthy controls (0.3%; P

Naturally occurring osmoprotectants are known to prevent aggregation of proteins under various stress factors including extreme pH and elevated temperature conditions. Here, we synthesized gold nanoparticles coated with selected osmolytes (proline, hydroxyproline, and glycine) and examined their effect on temperature-induced amyloid-formation of insulin hormone. These uniform, thermostable, and hemocompatible gold nanoparticles were capable of inhibiting both spontaneous and seed-induced amyloid aggregation of insulin. Both quenching and docking experiments suggest a direct interaction between the osmoprotectant-coated nanoparticles and aggregation-prone hydrophobic stretches of insulin. Circular-dichroism results confirmed the retention of insulin's native structure in the presence of these nanoparticles. Unlike the indirect solvent-mediated effect of free osmolytes, the inhibition effect of osmolyte-coated gold nanoparticles was observed to be mediated through their direct interaction with insulin. The results signify the protection of the exposed aggregation-prone domains of insulin from temperature-induced self-assembly through osmoprotectant-coated nanoparticles, and such effect may inspire the development of osmolyte-based antiamyloid nanoformulations.

Amyloid-β (Aβ) and tau oligomers have been identified as neurotoxic agents responsible for causing Alzheimer’s disease (AD). Clinical trials using Aβ and tau as targets have failed, giving rise to calls for new research approaches to combat AD. This paper provides such an approach. Most basic AD research has involved quiescent Aβ and tau solutions. However, studies involving laminar and extensional flow of proteins have demonstrated that mechanical agitation of proteins induces or accelerates protein aggregation. Recent MRI brain studies have revealed high energy, chaotic motion of cerebrospinal fluid (CSF) in lower brain and brainstem regions. These and studies showing CSF flow within the brain have shown that there are two energetic hot spots. These are within the third and fourth brain ventricles and in the neighborhood of the circle of Willis blood vessel region. These two regions are also the same locations as those of the earliest Aβ and tau AD pathology. In this paper, it is proposed that cardiac systolic pulse waves that emanate from the major brain arteries in the lower brain and brainstem regions and whose pulse waves drive CSF flows within the brain are responsible for initiating AD and possibly other amyloid diseases. It is further proposed that the triggering of these diseases comes about because of the strengthening of systolic pulses due to major artery hardening that generates intense CSF extensional flow stress. Such stress provides the activation energy needed to induce conformational changes of both Aβ and tau within the lower brain and brainstem region, producing unique neurotoxic oligomer molecule conformations that induce AD.

Background Treatment of neurodegenerative diseases, such as Parkinson’s disease, Huntington’s chorea, Alzheimer’s disease, is one of the priority directions in modern medicine. Thus, search and production of new physiologically active substances for the treatment of neurodegenerative disorders is one of the most important tasks for organic chemistry. The approach based on the replacement of a peptide bond in a peptide molecule with a structural isostere, non-hydrolyzable methylene phosphoryl fragment makes it possible to increase the metabolic stability of peptide molecules to the destructive action of peptidases. Methods This work is devoted to the approbation of a new synthetic approach to the production of physiologically active substances in a series of peptide-type compounds with activity by replacing the peptide bond with isosteric methylene-phosphoryl fragment with the preservation of the original amino acid sequence. Results A phosphine analog of the known physiologically active tripeptide proline-glycine-proline was obtained, cytotoxicity and neuroprotective properties of the initial tripeptide and its phosphine analog were studied. Conclusion Preliminary biological tests have shown that the obtained phosphine analog of the proline-glycine-proline tripeptide is involved in modulating the formation of sediments in the cellular system of proteinopathy, which may indicate their potential antiaggregatory properties.

Mitochondrial pathophysiology is implicated in the development of Alzheimer’s disease (AD). An integrative database of gene dysregulation suggests that the mitochondrial ubiquitin ligase MITOL/MARCH5, a fine-tuner of mitochondrial dynamics and functions, is downregulated in patients with AD. Here, we report that the perturbation of mitochondrial dynamics by MITOL deletion triggers mitochondrial impairments and exacerbates cognitive decline in a mouse model with AD-related Aβ pathology. Notably, MITOL deletion in the brain enhanced the seeding effect of Aβ fibrils, but not the spontaneous formation of Aβ fibrils and plaques, leading to excessive secondary generation of toxic and dispersible Aβ oligomers. Consistent with this, MITOL-deficient mice with Aβ etiology exhibited worsening cognitive decline depending on Aβ oligomers rather than Aβ plaques themselves. Our findings suggest that alteration in mitochondrial morphology might be a key factor in AD due to directing the production of Aβ form, oligomers or plaques, responsible for disease development., Takeda et al. investigate the role of mitochondrial ubiquitin ligase (MITOL) in Alzheimer’s disease progression. They demonstrate that MITOL has a direct effect on mitochondrial dynamics and functioning and its ablation can enhance the seeding effect of Aβ-plaques, leading to cognitive decline in the transgenic mice model of Alzheimer’s disease.

Amyotrophic lateral sclerosis (ALS) is a form of motor neuron disease (MND) that is characterized by the progressive loss of motor neurons within the spinal cord, brainstem and motor cortex. Although ALS clinically manifests as a heterogeneous disease, with varying disease onset and survival, a unifying feature is the presence of ubiquitinated cytoplasmic protein inclusion aggregates containing TDP-43. However, the precise mechanisms linking protein inclusions and aggregation to neuronal loss are currently poorly understood.Bimolecular Fluorescence Complementation (BiFC) takes advantage the association of fluorophore fragments (non-fluorescent on their own) that are attached to an aggregation prone protein of interest. Interaction of the proteins of interest allows for the fluorescent reporter protein to fold into its native state and emit a fluorescent signal. Here, we combined the power of BiFC with the advantages of the zebrafish system to validate, optimize and visualize of the formation of ALS-linked aggregates in real time in a vertebrate model. We further provide in vivo validation of the selectivity of this technique and demonstrate reduced spontaneous self-assembly of the non-fluorescent fragments in vivo by introducing a fluorophore mutation. Additionally, we report preliminary findings on the dynamic aggregation of the ALS-linked hallmark proteins Fus and TDP-43 in their corresponding nuclear and cytoplasmic compartments using BiFC.Overall, our data demonstrates the suitability of this BiFC approach to study and characterize ALS-linked aggregate formation in vivo. Importantly, the same principle can be applied in the context of other neurodegenerative diseases and has therefore critical implications to advance our understanding of pathologies that underlie aberrant protein aggregation.

The cause of most Parkinson's disease cases is unknown. However, it is well documented that mitochondrial dysfunction and misfolded α synuclein aggregation are important cellular abnormalities associated with the disease. In this paper, we use the microcompetition model to show how latent viruses, which infect the central and peripheral nervous systems, can cause the observed mitochondrial dysfunction and excess α synuclein aggregation, and eventually, Parkinson's disease.

The ubiquitin-proteasome system is the major pathway for the maintenance of protein homeostasis. Its inhibition causes accumulation of ubiquitinated proteins; this accumulation has been associated with several of the most common neurodegenerative diseases. Several genetic factors have been identified for most neurodegenerative diseases, however, most cases are considered idiopathic, thus making the study of the mechanisms of protein accumulation a relevant field of research. It is often mentioned that the biggest risk factor for neurodegenerative diseases is aging, and several groups have reported an age-related alteration of the expression of some of the 26S proteasome subunits and a reduction of its activity. Proteasome subunits interact with proteins that are known to accumulate in neurodegenerative diseases such as α-synuclein in Parkinson's, tau in Alzheimer's, and huntingtin in Huntington's diseases. These interactions have been explored for several years, but only until recently, we are beginning to understand them. In this review, we discuss the known interactions, the underlying patterns, and the phenotypes associated with the 26S proteasome subunits in the etiology and progression of neurodegenerative diseases where there is evidence of proteasome involvement. Special emphasis is made in reviewing proteasome subunits that interact with proteins known to have an age-related altered expression or to be involved in neurodegenerative diseases to explore key effectors that may trigger or augment their progression. Interestingly, while the causes of age-related reduction of some of the proteasome subunits are not known, there are specific relationships between the observed neurodegenerative disease and the affected proteasome subunits.

The deposition of highly ordered fibrillar-type aggregates into inclusion bodies is a hallmark of neurodegenerative diseases such as Parkinson’s disease. The high stability of such amyloid fibril aggregates makes them challenging substrates for the cellular protein quality-control machinery1,2. However, the human HSP70 chaperone and its co-chaperones DNAJB1 and HSP110 can dissolve preformed fibrils of the Parkinson’s disease-linked presynaptic protein α-synuclein in vitro3,4. The underlying mechanisms of this unique activity remain poorly understood. Here we use biochemical tools and nuclear magnetic resonance spectroscopy to determine the crucial steps of the disaggregation process of amyloid fibrils. We find that DNAJB1 specifically recognizes the oligomeric form of α-synuclein via multivalent interactions, and selectively targets HSP70 to fibrils. HSP70 and DNAJB1 interact with the fibril through exposed, flexible amino and carboxy termini of α-synuclein rather than the amyloid core itself. The synergistic action of DNAJB1 and HSP110 strongly accelerates disaggregation by facilitating the loading of several HSP70 molecules in a densely packed arrangement at the fibril surface, which is ideal for the generation of ‘entropic pulling’ forces. The cooperation of DNAJB1 and HSP110 in amyloid disaggregation goes beyond the classical substrate targeting and recycling functions that are attributed to these HSP70 co-chaperones and constitutes an active and essential contribution to the remodelling of the amyloid substrate. These mechanistic insights into the essential prerequisites for amyloid disaggregation may provide a basis for new therapeutic interventions in neurodegeneration. The molecular steps that lead to the disaggregation of amyloid fibrils are shown to involve the synergistic action of HSP70 and its co-chaperones DNAJB1 and HSP110.

Alzheimer's disease (AD) is progressive brain amyloidosis that damages brain regions associated with memory, thinking, behavioral and social skills. Neuropathologically, AD is characterized by intraneuronal hyperphosphorylated tau inclusions as neurofibrillary tangles (NFTs), and buildup of extracellular amyloid-beta (Aβ) peptide as senile plaques. Several biomarker tests capturing these pathologies have been developed. However, for the full clinical expression of the neurodegenerative events of AD, there exist other central molecular pathways. In terms of understanding the unidentified underlying processes for the progression and development of AD, a complete comprehension of the structure and composition of atypical aggregation of proteins is essential. Presently, to aid the prognosis, diagnosis, detection, and development of drug targets in AD, neuroproteomics is elected as one of the leading essential tools for the efficient exploratory discovery of prospective biomarker candidates estimated to play a crucial role. Therefore, the aim of this review is to present the role of neuroproteomics to analyze the complexity of AD.

In tauopathies, tau forms pathogenic fibrils with distinct conformations (termed “tau strains”) and acts as an aggregation “seed” templating the conversion of normal tau into isomorphic fibrils. Previous research showed that the aggregation core of tau fibril covers the C-terminal region (243–406 amino acids (aa)) and differs among the diseases. However, the mechanisms by which distinct fibrous structures are formed and inherited via templated aggregation are still unknown. Here, we sought to identify the key sequences of seed-dependent aggregation. To identify sequences for which deletion reduces tau aggregation, SH-SY5Y cells expressing a series of 10 partial deletion (Del 1–10, covering 244–400 aa) mutants of tau-CTF24 (243–441 aa) were treated with tau seeds prepared from a different tauopathy patient's brain (Alzheimer's disease, progressive supranuclear palsy, and corticobasal degeneration) or recombinant tau, and then seed-dependent tau aggregation was assessed biochemically. We found that the Del 8 mutant lacking 353–368 aa showed significantly decreased aggregation in both cellular and in vitro models. Furthermore, to identify the minimum sequence responsible for tau aggregation, we systematically repeated cellular tau aggregation assays for the delineation of shorter deletion sites and revealed that Asn-368 mutation suppressed tau aggregation triggered by an AD tau seed, but not using other tauopathy seeds. Our study suggested that 353–368 aa is a novel aggregation-responsible sequence other than PHF6 and PHF6*, and within this sequence, the Asn-368 residue plays a role in strain-specific tau aggregation in different tauopathies.

Osmolytes are organic solutes that change the protein folding landscape shifting the equilibrium towards the folded state. Herein, we use osmolytes to probe the structuring and aggregation of the intrinsically disordered mutant Huntingtin (mHtt) vis-a-vis the pathogenicity of mHtt on transcription factor function and cell survival. Using an inducible PC12 cell model of Huntington’s disease (HD), we show that stabilizing polyol osmolytes drive the aggregation of Htt103QExon1-EGFP from a diffuse ensemble into inclusion bodies (IBs), whereas the destabilizing osmolyte urea does not. This effect of stabilizing osmolytes is innate, generic, countered by urea, and unaffected by HSP70 and HSC70 knockdown. A qualitatively similar result of osmolyte-induced mHtt IB formation is observed in a conditionally immortalized striatal neuron model of HD, and IB formation correlates with improved survival under stress. Increased expression of diffuse mHtt sequesters the CREB transcription factor to repress CREB-reporter gene activity. This repression is mitigated either by stabilizing osmolytes, which deplete diffuse mHtt or by urea, which negates protein–protein interaction. Our results show that stabilizing polyol osmolytes promote mHtt aggregation, alleviate CREB dysfunction, and promote survival under stress to support the hypothesis that lower molecular weight entities of disease protein are relevant pathogenic species in neurodegeneration.

Amyloidogenic protein assembly into insoluble fibrillar aggregates is linked with several neurodegenerative disorders, such as Alzheimer’s or Parkinson’s disease, affecting millions of people worldwide. The search for a potential anti-amyloid drug has led to the discovery of hundreds of compounds, none of which have passed all clinical trials. Gallic acid has been shown to both modulate factors leading to the onset of neurodegenerative disorders, as well as directly inhibit amyloid formation. However, the conditions under which this effect is seen could lead to oxidation of this polyphenol, likely changing its properties. Here we examine the effect of gallic acid and its oxidised form on the aggregation of a model amyloidogenic protein–insulin at low pH conditions. We show a vastly higher inhibitory potential of the oxidised form, as well as an alteration in the aggregation pathway, leading to the formation of a specific fibril conformation.

Pleiotropic roles are proposed for brain extracellular vesicles (EVs) in the development of Alzheimer's disease (AD). Our previous studies have suggested a beneficial role for EVs in AD, where the endosomal system in vulnerable neurons is compromised, contributing to the removal of accumulated material from neurons. However, the involvement of EVs in propagating AD amyloidosis throughout the brain has been considered because the amyloid‐β precursor protein (APP), APP metabolites, and key APP cleaving enzymes were identified in association with EVs. Here, we undertook to determine whether the secretase machinery is actively processing APP in EVs isolated from the brains of wild‐type and APP overexpressing Tg2576 mice. We found that full‐length APP is cleaved in EVs incubated in the absence of cells. The resulting metabolites, both α‐ and β‐APP carboxyl‐terminal fragments and APP intracellular domain accumulate in EVs over time and amyloid‐β dimerizes. Thus, EVs contribute to the removal from neurons and transport of APP‐derived neurotoxic peptides. While this is potentially a venue for propagation of the pathology throughout the brain, it may contribute to efficient removal of neurotoxic peptides from the brain.

The present study was aimed to study the effect of kaempferol, on the transgenic Drosophila model of Parkinson’s disease. Kaempferol was added in the diet at final concentration of 10, 20, 30 and 40 µM and the effect was studied on various cognitive and oxidative stress markers. The results of the study showed that kaempferol, delayed the loss of climbing ability as well as the activity of PD flies in a dose dependent manner compared to unexposed PD flies. A dose-dependent reduction in oxidative stress markers was also observed. Histopathological examination of fly brains using anti-tyrosine hydroxylase immunostaining has revealed a significant dose-dependent increase in the expression of tyrosine hydroxylase in PD flies exposed to kaempferol. Molecular docking results revealed that kaempferol binds to human alpha synuclein at specific sites that might results in the inhibition of alpha synuclein aggregation and prevents the formation of Lewy bodies.

Prion diseases are fatal infectious neurodegenerative disorders in human and animals caused by misfolding of the cellular prion protein (PrPC) into the pathological isoform PrPSc. Elucidating the molecular and cellular mechanisms underlying prion propagation may help to develop disease interventions. Cell culture systems for prion propagation have greatly advanced molecular insights into prion biology, but translation of in vitro to in vivo findings is often disappointing. A wider range of cell culture systems might help overcome these shortcomings. Here, we describe an immortalized mouse neuronal astrocyte cell line (C8D1A) that can be infected with murine prions. Both PrPC protein and mRNA levels in astrocytes were comparable with those in neuronal and non-neuronal cell lines permitting persistent prion infection. We challenged astrocytes with three mouse-adapted prion strains (22L, RML, and ME7) and cultured them for six passages. Immunoblotting results revealed that the astrocytes propagated 22L prions well over all six passages, whereas ME7 prions did not replicate, and RML prions replicated only very weakly after five passages. Immunofluorescence analysis indicated similar results for PrPSc. Interestingly, when we used prion conversion activity as a readout in real-time quaking-induced conversion assays with RML-infected cell lysates, we observed a strong signal over all six passages, comparable with that for 22L-infected cells. These data indicate that the C8D1A cell line is permissive to prion infection. Moreover, the propagated prions differed in conversion and proteinase K–resistance levels in these astrocytes. We propose that the C8D1A cell line could be used to decipher prion strain biology.

Background: Loss of a-crystallin chaperone function results in the lens protein aggregation leading to cataract. In this study, we evaluated the efficacy of micellar lutein with different fatty acids in modulating a-crystallin chaperone function under selenite cataract conditions. Methods: Cataract was induced in rat pups by giving sodium selenite (25 mM/kg body weight) by IP. Lutein [(L), 1.3 mmol/kg body weight)] was given day before and five days after selenite injection as a micelle with 7.5 mM linoleic acid (LA), or 7.5 mM eicosapentaenoic acid (EPA)þdocosahexaenoic acid (DHA) or 7.5 mM oleic acid (OA). Lens a-crystallins was purified, and its chaperone function and integrity was assessed. Cholesterol, calcium, calpain-2, procaspase-3, and expression of a-A and b-B1 crystallin in the lens of cataract and micellar lutein administered rats were evaluated. Results: Cataract induction significantly (p < 0.05) decreased lens a-crystallin chaperone function. Cataract rats had increased cholesterol and calcium level, increased the expression of calpain-2, and a-A and b-B1 crystallin, and reduced the pro-caspase-3 level in the lens. However, micellar lutein administration significantly (p < 0.05) protected client proteins from aggregation via the modulation of calciumdependent calpain-2 protease activity. The chaperone function of lens a-crystallins in rats administered micellar lutein with EPA þ DHA was found to be highest when compared to OA and LA. Conclusions: Micellar lutein with unsaturated fatty acids beneficially modulates a-crystallin chaperone function. Among the fatty acids tested, micellar lutein with EPA þ DHA exhibited superior effects, thereby offering a promising strategy for cataract management.

There is growing recognition in the field of neurodegenerative diseases that mixed proteinopathies are occurring at greater frequency than originally thought. This is particularly true for three amyloid proteins defining most of these neurological disorders, amyloid-beta (Aβ), tau, and alpha-synuclein (αSyn). The co-existence and often co-localization of aggregated forms of these proteins has led to the emergence of concepts positing molecular interactions and cross-seeding between Aβ, tau, and αSyn aggregates. Amongst this trio, αSyn has received particular attention in this context during recent years due to its ability to modulate Aβ and tau aggregation in vivo, to interact at a molecular level with Aβ and tau in vivo and to cross-seed tau in mice. Here we provide a comprehensive, critical, and accessible review about the expression, role and nature of endogenous soluble αSyn oligomers because of recent developments in the understanding of αSyn multimerization, misfolding, aggregation, cross-talk, spreading and cross-seeding in neurodegenerative disorders, including Parkinson's disease, dementia with Lewy bodies, multiple system atrophy, Alzheimer's disease, and Huntington's disease. We will also discuss our current understanding about the relative toxicity of endogenous αSyn oligomers in vivo and in vitro, and introduce potential opportunities to counter their deleterious effects.

α-Synuclein fibrillation is now regarded as a major pathogenic process in Parkinson’s disease and its proteinaceous deposits are also detected in other neurological disorders including Alzheimer's disease. Therefore anti-amyloidegenic compounds may delay or prevent the progression of synucleinopathies disease. Molecular chaperones are group of proteins which mediate correct folding of proteins by preventing unsuitable interactions which may lead to aggregation. The objective of this study was to investigate the anti-amyloidogenic effect of molecular chaperone artemin on α-synuclein. As the concentration of artemin was increased up to 4 μg/ml, a decrease in fibril formation of α-synuclein was observed using thioflavin T (ThT) fluorescence and congo red (CR) assay. Transmission electron microscopy (TEM) images also demonstrated a reduction in fibrils in the presence of artemin. The secondary structure of α-synuclein was similar to its native form prior to fibrillation when incubated with artemin. A cell-based assay has shown that artemin inhibits α-synuclein aggregation and reduce cytotoxicity, apoptosis and reactive oxygen species (ROS) production. Our results revealed that artemin has efficient chaperon activity for preventing α-synuclein fibril formation and toxicity.

The presence of intraneuronal Lewy bodies (LBs) and Lewy neurites (LNs) in the substantia nigra (SN) composed of aggregated α-synuclein (α-syn) has been recognized as a hallmark of pathological changes in Parkinson’s disease (PD). Numerous studies have shown that aggregated α-syn is necessary for neurotoxicity. Meanwhile, the mitochondrial and lysosomal dysfunctions are associated with α-syn pathogenicity. The hypothesis that α-syn transmission in the human brain contributes to the instigation and progression of PD has provided insights into PD pathology. This review will provide a brief overview of increasing researches that shed light on the relationship of α-syn aggregation with mitochondrial and lysosomal dysfunctions, and highlight recent understanding of α-syn transmission in PD pathology.

Amyloid fibrils result from the aggregation of host cell-encoded proteins, many giving rise to specific human illnesses such as Alzheimer’s disease. Here we show that the major virulence factor of Rift Valley fever virus, the protein NSs, forms filamentous structures in the brain of mice and affects mortality. NSs assembles into nuclear and cytosolic disulfide bond-dependent fibrillary aggregates in infected cells. NSs structural arrangements exhibit characteristics typical for amyloids, such as an ultrastructure of 12 nm-width fibrils, a strong detergent resistance, and interactions with the amyloid-binding dye Thioflavin-S. The assembly dynamics of viral amyloid-like fibrils can be visualized in real-time. They form spontaneously and grow in an amyloid fashion within 5 hours. Together, our results demonstrate that viruses can encode amyloid-like fibril-forming proteins and have strong implications for future research on amyloid aggregation and toxicity in general., Rift Valley fever virus (RVFV) can cause severe diseases in humans, including encephalitis. Here the authors show that NSs, the major virulence factor of RVFV, is an amyloidogenic protein forming fibrils in infected mouse brains and causing increased mortality in mice.

α-Synuclein (αsyn) is an abundant brain neuronal protein that can misfold and polymerize to form toxic fibrils coalescing into pathologic inclusions in neurodegenerative diseases, including Parkinson's disease, Lewy body dementia, and multiple system atrophy. These fibrils may induce further αsyn misfolding and propagation of pathologic fibrils in a prion-like process. It is unclear why αsyn initially misfolds, but a growing body of literature suggests a critical role of partial proteolytic processing resulting in various truncations of the highly charged and flexible carboxyl-terminal region. This review aims to 1) summarize recent evidence that disease-specific proteolytic truncations of αsyn occur in Parkinson's disease, Lewy body dementia, and multiple system atrophy and animal disease models; 2) provide mechanistic insights on how truncation of the amino and carboxyl regions of αsyn may modulate the propensity of αsyn to pathologically misfold; 3) compare experiments evaluating the prion-like properties of truncated forms of αsyn in various models with implications for disease progression; 4) assess uniquely toxic properties imparted to αsyn upon truncation; and 5) discuss pathways through which truncated αsyn forms and therapies targeted to interrupt them. Cumulatively, it is evident that truncation of αsyn, particularly carboxyl truncation that can be augmented by dysfunctional proteostasis, dramatically potentiates the propensity of αsyn to pathologically misfold into uniquely toxic fibrils with modulated prion-like seeding activity. Therapeutic strategies and experimental paradigms should operate under the assumption that truncation of αsyn is likely occurring in both initial and progressive disease stages, and preventing truncation may be an effective preventative strategy against pathologic inclusion formation.

Amyotrophic lateral sclerosis (ALS) is a complex neurodegenerative disease predominantly affecting upper and lower motor neurons. Diagnosis of this devastating pathology is very difficult because the high degree of clinical heterogeneity with which it occurs and until now, no truly effective treatment exists.Molecular diagnosis may be a valuable tool for dissecting out ALS complex heterogeneity and for identifying new molecular mechanisms underlying the characteristic selective degeneration and death of motor neurons. To date, pathogenic variants in ALS genes are known to be present in up to 70% of familial and 10% of apparently sporadic ALS cases and can be associated with risks for ALS only or risks for other neurodegenerative diseases. This paper shows the procedure currently used in diagnostic laboratories to investigate most frequent mutations in ALS and evaluating the utility of involved molecular techniques as potential tools to discriminate 'common mutations' in ALS patients.Genetic testing may allow for establishing an accurate pathological diagnosis and a more precise stratification of patient groups in future drug trials.

Despite the distinction between ischemic stroke and neurodegenerative disorders, they share numerous pathophysiologies particularly those mediated by inflammation and oxidative stress. Although protein aggregation is considered to be a hallmark of neurodegenerative diseases, the formation of protein aggregates can be also induced within a short time after cerebral ischemia, aggravating cerebral ischemic injury. Protein aggregation uncovers a previously unappreciated molecular overlap between neurodegenerative diseases and ischemic stroke. Unfortunately, compared with neurodegenerative disease, mechanism of protein aggregation after cerebral ischemia and how this can be averted remain unclear. This review highlights current understanding on protein aggregation and its intrinsic role in ischemic stroke.

Aging and female sex are the major risk factors for Alzheimer's disease and its associated brain amyloid-β (Aβ) neuropathology, but the mechanisms mediating these risk factors remain uncertain. Evidence indicates that Aβ aggregation by Zn2+ released from glutamatergic neurons contributes to amyloid neuropathology, so we tested whether aging and sex adversely influences this neurophysiology. Using acute hippocampal slices, we found that extracellular Zn2+-elevation induced by high K+ stimulation was significantly greater with older (65 weeks vs 10 weeks old) rats, and was exaggerated in females. This was driven by slower reuptake of extracellular Zn2+, which could be recapitulated by mitochondrial intoxication. Zn2+:Aβ aggregates were toxic to the slices, but Aβ alone was not. Accordingly, high K+ caused synthetic human Aβ added to the slices to form soluble oligomers as detected by bis-ANS, attaching to neurons and inducing toxicity, with older slices being more vulnerable. Age-dependent energy failure impairing Zn2+ reuptake, and a higher maximal capacity for Zn2+ release by females, could contribute to age and sex being major risk factors for Alzheimer's disease.

Human pancreatic islet amyloid polypeptide (hIAPP) and beta amyloid (Aβ) can accumulate in Type 2 diabetes (T2D) and Alzheimer’s disease (AD) brains and evidence suggests that interaction between the two amyloidogenic proteins can lead to the formation of heterocomplex aggregates. However, the structure and consequences of the formation of these complexes remains to be determined. The main objective of this study was to characterise the different types and morphology of Aβ-hIAPP heterocomplexes and determine if formation of such complexes exacerbate neurotoxicity. We demonstrate that hIAPP promotes Aβ oligomerization and formation of small oligomer and large aggregate heterocomplexes. Co-oligomerized Aβ42-hIAPP mixtures displayed distinct amorphous structures and a 3-fold increase in neuronal cell death as compared to Aβ and hIAPP alone. However, in contrast to hIAPP, non-amyloidogenic rat amylin (rIAPP) reduced oligomer Aβ-mediated neuronal cell death. rIAPP exhibited reductions in Aβ induced neuronal cell death that was independent of its ability to interact with Aβ and form heterocomplexes; suggesting mediation by other pathways. Our findings reveal distinct effects of IAPP peptides in modulating Aβ aggregation and toxicity and provide new insight into the potential pathogenic effects of Aβ-IAPP hetero-oligomerization and development of IAPP based therapies for AD and T2D.

The polyglutamine (polyQ) diseases are a group of inherited neurodegenerative diseases that include Huntington’s disease, various spinocerebellar ataxias, spinal and bulbar muscular atrophy, and dentatorubral pallidoluysian atrophy. They are caused by the abnormal expansion of a CAG repeat coding for the polyQ stretch in the causative gene of each disease. The expanded polyQ stretches trigger abnormal β-sheet conformational transition and oligomerization followed by aggregation of the polyQ proteins in the affected neurons, leading to neuronal toxicity and neurodegeneration. Disease-modifying therapies that attenuate both symptoms and molecular pathogenesis of polyQ diseases remain an unmet clinical need. Here we identified arginine, a chemical chaperone that facilitates proper protein folding, as a novel compound that targets the upstream processes of polyQ protein aggregation by stabilizing the polyQ protein conformation. We first screened representative chemical chaperones using an in vitro polyQ aggregation assay, and identified arginine as a potent polyQ aggregation inhibitor. Our in vitro and cellular assays revealed that arginine exerts its anti-aggregation property by inhibiting the toxic β-sheet conformational transition and oligomerization of polyQ proteins before the formation of insoluble aggregates. Arginine exhibited therapeutic effects on neurological symptoms and protein aggregation pathology in Caenorhabditis elegans, Drosophila, and two different mouse models of polyQ diseases. Arginine was also effective in a polyQ mouse model when administered after symptom onset. As arginine has been safely used for urea cycle defects and for mitochondrial myopathy, encephalopathy, lactic acid and stroke syndrome patients, and efficiently crosses the blood–brain barrier, a drug-repositioning approach for arginine would enable prompt clinical application as a promising disease-modifier drug for the polyQ diseases.

Amyloid-β (Aβ) dysmetabolism is tightly associated with pathological processes in Alzheimer's disease (AD). Currently, it is thought that, in addition to Aβ fibrils that give rise to plaque formation, Aβ aggregates into non-fibrillar soluble oligomers (AβOs). Soluble AβOs have been extensively studied for their synaptotoxic and neurotoxic properties. In this review, we discuss physicochemical properties of AβOs and their impact on different brain cell types in AD. Additionally, we summarize three decades of studies with AβOs, providing a compelling bulk of evidence regarding cell-specific mechanisms of toxicity. Cellular models may lead us to a deeper understanding of the detrimental effects of AβOs in neurons and glial cells, putatively shedding light on the development of innovative therapies for AD.

In vivo, small heat-shock proteins (sHsps) are key players in maintaining a healthy proteome. αB-crystallin (αB-c) or HspB5 is one of the most widespread and populous of the ten human sHsps. Intracellularly, αB-c acts via its molecular chaperone action as the first line of defence in preventing target protein unfolding and aggregation under conditions of cellular stress. In this review, we explore how the structure of αB-c confers its function and interactions within its oligomeric self, with other sHsps, and with aggregation-prone target proteins. Firstly, the interaction between the two highly conserved regions of αB-c, the central α-crystallin domain and the C-terminal IXI motif and how this regulates αB-c chaperone activity are explored. Secondly, subunit exchange is rationalised as an integral structural and functional feature of αB-c. Thirdly, it is argued that monomeric αB-c may be its most chaperone-species active, but at the cost of increased hydrophobicity and instability. Fourthly, the reasons why hetero-oligomerisation of αB-c with other sHsps is important in regulating cellular proteostasis are examined. Finally, the interaction of αB-c with aggregation-prone, partially folded target proteins is discussed. Overall, this paper highlights the remarkably diverse capabilities of αB-c as a caretaker of the cell. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12192-020-01098-w) contains supplementary material, which is available to authorized users.

Alzheimer's disease (AD) is a chronic and irreversible neurodegenerative disorder. Abnormal aggregation of the neurotoxic amyloid‑β (Aβ) peptide is an early event in AD. The activation of astrocytic α7 nicotinic acetylcholine receptor (α7 nAChR) can inhibit Aβ aggregation; thus, the molecular mechanism between α7 nAChR activation and Aβ aggregation warrants further investigation. In the present study, Aβ oligomer levels were assessed in astrocytic cell lysates after treatment with PNU282987 (a potent agonist of α7 nAChRs) or co‑treatment with LY294002, a p‑Akt inhibitor. The levels of heat shock factor‑1 (HSF‑1), heat shock protein 70 (HSP‑70), and αB‑crystallin (Cryab) in astrocytes treated with PNU282987 at various time‑points or co‑treated with methyllycaconitine (MLA), a selective α7 nAChR antagonist, as well as co‑incubated with LY294002 were determined by western blotting. HSP‑70 and Cryab levels were determined after HSF‑1 knockdown (KD) in astrocytes. PNU282987 markedly inhibited Aβ aggregation and upregulated HSF‑1, Cryab, and HSP‑70 in primary astrocytes, while the PNU282987‑mediated neuroprotective effect was reversed by pre‑treatment with MLA or LY294002. Moreover, the HSF‑1 KD in astrocytes effectively decreased Cryab, but not HSP‑70 expression. HSF‑1 is necessary for the upregulation of Cryab expression, but not for that of HSP‑70. HSF‑1 and HSP‑70 have a neuroprotective effect. Furthermore, the neuroprotective effect of PNU282987 against Aβ aggregation was mediated by the canonical PI3K/Akt signaling pathway activation.

Expansion above a certain threshold in the polyglutamine (polyQ) tract of ataxin-3 is the main cause of neurodegeneration in Machado-Joseph disease. Ataxin-3 contains an N-terminal catalytic domain, called Josephin domain, and a highly aggregation-prone C-terminal domain containing the polyQ tract. Recent work has shown that protein aggregation inhibits clathrin-mediated endocytosis (CME). However, the effects of polyQ expansion in ataxin-3 on CME have not been investigated. We hypothesize that the expansion of the polyQ tract in ataxin-3 could impact CME. Here, we report that both the wild-type and the expanded ataxin-3 reduce transferrin internalization and expanded ataxin-3 impacts dynamics of clathrin-coated pits (CCPs) by reducing CCP nucleation and increasing short-lived abortive CCPs. Since endocytosis plays a central role in regulating receptor uptake and cargo release, our work highlights a potential mechanism linking protein aggregation to cellular dysregulation.

Parkinson's disease (PD) is one of the most common movement disorders with loss of dopaminergic neurons and the presence of Lewy bodies in certain brain areas. However, it is not clear how Lewy body (inclusion with protein aggregation) formation occurs. Mutations in leucine-rich repeat kinase 2 (LRRK2) can cause a genetic form of PD and contribute to sporadic PD with the typical Lewy body pathology. Here, we used our recently identified LRRK2 GTP-binding inhibitors as pharmacological probes to study the LRRK2-linked ubiquitination and protein aggregation. Pharmacological inhibition of GTP-binding by GTP-binding inhibitors (68 and Fx2149) increased LRRK2-linked ubiquitination predominantly via K27 linkage. Compound 68- or Fx2149 increased G2019S-LRRK2-linked ubiquitinated aggregates, which occurred through the atypical linkage types K27 and K63. Coexpression of K27R and K63R, which prevented ubiquitination via K27 and K63 linkages, reversed the effects of 68 and Fx2149. Moreover, 68 and Fx2149 also promoted G2019S-LRRK2-linked aggresome (Lewy body-like inclusion) formation via K27 and K63 linkages. These findings demonstrate that LRRK2 GTP-binding activity is critical in LRRK2-linked ubiquitination and aggregation formation. These studies provide novel insight into the LRRK2-linked Lewy body-like inclusion formation underlying PD pathogenesis.

Maintaining protein homeostasis (proteostasis) is essential for cellular health and is governed by a network of quality control machinery comprising over 800 genes. When proteostasis becomes imbalanced, proteins can abnormally aggregate or become mislocalized. Inappropriate protein aggregation and proteostasis imbalance are two of the central pathological features of common neurodegenerative diseases including Alzheimer, Parkinson, Huntington, and motor neuron diseases. How aggregation contributes to the pathogenic mechanisms of disease remains incompletely understood. Here, we integrate some of the key and emerging ideas as to how protein aggregation relates to imbalanced proteostasis with an emphasis on Huntington disease as our area of main expertise. We propose the term "aggregomics" be coined in reference to how aggregation of particular proteins concomitantly influences the spatial organization and protein-protein interactions of the surrounding proteome. Meta-analysis of aggregated interactomes from various published datasets reveals chaperones and RNA-binding proteins are common components across various disease contexts. We conclude with an examination of therapeutic avenues targeting proteostasis mechanisms.

Synucleinopathies are neurological diseases that are characterized by the accumulation of aggregates of a cytosolic protein, α-synuclein, at the plasma membrane. Even though the pathological role of the protein is established, the mechanism by which it damages neurons remains unclear due to the difficulty to correctly mimic the plasma membrane in vitro. Using a microfluidic setup in which the composition of the plasma membrane, including the asymmetry of the two leaflets, is recapitulated, we demonstrate a triple action of α-synuclein on the membrane. First, it changes membrane topology by inducing pores of discrete sizes, likely nucleated from membrane-bound proteins and subsequently enlarged by proteins in solution. Second, protein binding to the cytosolic leaflet increases the membrane capacitance by thinning it and/or changing its relative permittivity. Third, α-synuclein insertion inside the membrane hydrophobic core immobilizes the lipids in both leaflets, including the opposing protein-free extracellular one., Heo and Pincet demonstrate the influence of α-synuclein aggregation on in vitro asymmetric membranes using a microfluidic setup. They show that synuclein aggregation on the asymmetric membranes leads to its thinning and formation of pores of discrete sizes and synuclein insertion immobilizes both the leaflets of the asymmetric membrane.

The aberrant aggregation of normally soluble proteins into amyloid fibrils is the pathological hallmark of several neurodegenerative disorders, including Alzheimer's and Parkinson's diseases. Understanding this process will be key to developing both diagnostic and therapeutic approaches for neurodegenerative diseases. Recent advances in biophysical techniques, coupled with kinetic analyses have enabled a thorough description of the key molecular steps involved in protein aggregation. In this review, we discuss these advances and how they have been applied to study the ability of one such protein, α-Synuclein, to form neurotoxic oligomers.

Recent studies indicated that seeded fibril formation and toxicity of α-synuclein (α-syn) play a main role in the pathogenesis of certain diseases including Parkinson's disease (PD), multiple system atrophy, and dementia with Lewy bodies. Therefore, examination of compounds that abolish the process of seeding is considered a key step towards therapy of several synucleinopathies. Using biophysical, biochemical and cell-culture-based assays, assessment of eleven compounds, extracted from Chinese medicinal herbs, was performed in this study for their effect on α-syn fibril formation and toxicity caused by the seeding process. Salvianolic acid B and dihydromyricetin were the two compounds that strongly inhibited the fibril growth and neurotoxicity of α-syn. In an in-vitro cell model, these compounds decreased the insoluble phosphorylated α-syn and aggregation. Also, in primary neuronal cells, these compounds showed a reduction in α-syn aggregates. Both compounds inhibited the seeded fibril growth with dihydromyricetin having the ability to disaggregate preformed α-syn fibrils. In order to investigate the inhibitory mechanisms of these two compounds towards fibril formation, we demonstrated that salvianolic acid B binds predominantly to monomers, while dihydromyricetin binds to oligomeric species and to a lower extent to monomers. Remarkably, these two compounds stabilized the soluble non-toxic oligomers lacking β-sheet content after subjecting them to proteinase K digestion. Eleven compounds were tested but only two showed inhibition of α-syn aggregation, seeded fibril formation and toxicity in vitro. These findings highlight an essential beginning for development of new molecules in the field of synucleinopathies treatment. The work conducted by Dr. El-Agnaf laboratory was supported by Qatar Biomedical Research Institute under the Start-up Fund SF 2017–007. Funding for this work was provided in part by NIH/NIA grant R37AG019391 to D.E. This study was made possible by NPRP grant 4–1371–1-223 from the Qatar National Research Fund (a member of Qatar Foundation). The funding bodies provided financial support for this study; they had no role in the study design, performance, data collection and analysis, decision to publish and preparation/writing of the manuscript.

Aberrant structural formations of Cu/Zn superoxide dismutase enzyme (SOD1) are the probable mechanism by which circumscribed mutations in the SOD1 gene cause familial amyotrophic lateral sclerosis (ALS1). SOD1 forms aberrant structures which can proceed by nucleation to insoluble aggregates. Here, the SOD1 aggregation reaction was investigated predominantly by time-course studies on ALS1 variants G85R, G37R, D101G, and D101N in human embryonic kidney cells (HEK293FT), with analysis by detergent ultracentrifugation extractions and high-resolution PAGE methodologies. Nucleation was found to be pseudo-zeroth order and dependent on time and concentration at constant 37.0 °C and pH 7.4. The predominant subsets of the total SOD1 expression set which comprised the nucleation phase were both soluble and insoluble inactive monomers, trimers, and hexamers with reduced intra-disulfide bonds. Superoxide exposure via paraquat initiated the formation of SOD1 trimers in untransfected SH-SY5Y cells and increased the aggregation propensity of G85R in HEK293FT. These data show the kinetic formation of aberrant SOD1 subsets implicated in ALS1 and indicate that superoxide substrate may initiate its radical polymerization. In an instance of the utility of methodological reductionism in molecular theory: though many ALS1 variants retain their global enzymatic activity, the SOD1 subsets most implicated in causing ALS1 do not retain their specific activity.

Cytoplasmic aggregation of TDP-43 characterizes degenerating neurons in most cases of amyotrophic lateral sclerosis (ALS). Here, we develop an optogenetic TDP-43 variant (opTDP-43), whose multimerization status can be modulated in vivo through external light illumination. Using the translucent zebrafish neuromuscular system, we demonstrate that short-term light stimulation reversibly induces cytoplasmic opTDP-43 mislocalization, but not aggregation, in the spinal motor neuron, leading to an axon outgrowth defect associated with myofiber denervation. In contrast, opTDP-43 forms pathological aggregates in the cytoplasm after longer-term illumination and seeds non-optogenetic TDP-43 aggregation. Furthermore, we find that an ALS-linked mutation in the intrinsically disordered region (IDR) exacerbates the light-dependent opTDP-43 toxicity on locomotor behavior. Together, our results propose that IDR-mediated TDP-43 oligomerization triggers both acute and long-term pathologies of motor neurons, which may be relevant to the pathogenesis and progression of ALS., Optogenetic approaches for inducing TDP-43 aggregation have been described previously in cellular models. Here the authors develop an approach to optogenetically induce TDP-43 aggregation in vivo using zebrafish to model ALS pathologies.

Many pathological proteins related to neurodegenerative diseases are misfolded, aggregating to form amyloid fibrils during pathogenesis. One of the pathological proteins, alpha-synuclein (α-syn), accumulates in the brains of Parkinson disease (PD), dementia with Lewy bodies (DLB) and multiple system atrophy (MSA), which are designated as synucleinopathies. Recently, structural properties of abnormal accumulated proteins are suggested to determine the disease phenotype. However, the biochemical and structural characteristics of those accumulated proteins are still poorly understood. We previously reported the sequence and seed-structure-dependent polymorphic fibrils of α-syn and the polymorphism was identified by proteinase K-resistant cores determined by mass spectrometry (MS) analysis. In this study, we applied this method to analyze α-syn aggregates of MSA and DLB. To perform MS analysis on proteinase K-resistant cores, we first performed amplification of α-syn aggregates by seeding reaction and protein misfolding cyclic amplification (PMCA) to obtain a sufficient amount of aggregates. Using SDS insoluble fraction of the disease brain, we successfully amplified enough α-syn aggregates for MS analysis. We differentiated between mouse and human α-syn aggregates by MS analysis on proteinase K-resistant cores of the aggregates before and after amplification. The results suggest that structural properties of amplified α-syn fibrils are preserved after PMCA and these methods can be applicable in the study of pathological proteins of the neurodegenerative disorders.