Structure-Toxicity Relationship in Intermediate Fibrils from α-Synuclein Condensates.

The aberrant aggregation of α-synuclein (αS) into amyloid fibrils is associated with a range of highly debilitating neurodegenerative conditions, including Parkinson's disease. Although the structural properties of mature amyloids of αS are currently understood, the nature of transient protofilaments and fibrils that appear during αS aggregation remains elusive. Using solid-state nuclear magnetic resonance (ssNMR), cryogenic electron microscopy (cryo-EM), and biophysical methods, we here characterized intermediate amyloid fibrils of αS forming during the aggregation from liquid-like spherical condensates to mature amyloids adopting the structure of pathologically observed aggregates. These transient amyloid intermediates, which induce significant levels of cytotoxicity when incubated with neuronal cells, were found to be stabilized by a small core in an antiparallel ß-sheet conformation, with a disordered N-terminal region of the protein remaining available to mediate membrane binding. In contrast, mature amyloids that subsequently appear during the aggregation showed different structural and biological properties, including low levels of cytotoxicity, a rearranged structured core embedding also the N-terminal region, and a reduced propensity to interact with the membrane. The characterization of these two fibrillar forms of αS, and the use of antibodies and designed mutants, enabled us to clarify the role of critical structural elements endowing intermediate amyloid species with the ability to interact with membranes and induce cytotoxicity.

Comparison of tau spread in people with Down syndrome versus autosomal-dominant Alzheimer's disease: a cross-sectional study.

BACKGROUND: In people with genetic forms of Alzheimer's disease, such as in Down syndrome and autosomal-dominant Alzheimer's disease, pathological changes specific to Alzheimer's disease (ie, accumulation of amyloid and tau) occur in the brain at a young age, when comorbidities related to ageing are not present. Studies including these cohorts could, therefore, improve our understanding of the early pathogenesis of Alzheimer's disease and be useful when designing preventive interventions targeted at disease pathology or when planning clinical trials. We compared the magnitude, spatial extent, and temporal ordering of tau spread in people with Down syndrome and autosomal-dominant Alzheimer's disease. METHODS: In this cross-sectional observational study, we included participants (aged ≥25 years) from two cohort studies. First, we collected data from the Dominantly Inherited Alzheimer's Network studies (DIAN-OBS and DIAN-TU), which include carriers of autosomal-dominant Alzheimer's disease genetic mutations and non-carrier familial controls recruited in Australia, Europe, and the USA between 2008 and 2022. Second, we collected data from the Alzheimer Biomarkers Consortium-Down Syndrome study, which includes people with Down syndrome and sibling controls recruited from the UK and USA between 2015 and 2021. Controls from the two studies were combined into a single group of familial controls. All participants had completed structural MRI and tau PET (18F-flortaucipir) imaging. We applied Gaussian mixture modelling to identify regions of high tau PET burden and regions with the earliest changes in tau binding for each cohort separately. We estimated regional tau PET burden as a function of cortical amyloid burden for both cohorts. Finally, we compared the temporal pattern of tau PET burden relative to that of amyloid. FINDINGS: We included 137 people with Down syndrome (mean age 38·5 years [SD 8·2], 74 [54%] male, and 63 [46%] female), 49 individuals with autosomal-dominant Alzheimer's disease (mean age 43·9 years [11·2], 22 [45%] male, and 27 [55%] female), and 85 familial controls, pooled from across both studies (mean age 41·5 years [12·1], 28 [33%] male, and 57 [67%] female), who satisfied the PET quality-control procedure for tau-PET imaging processing. 134 (98%) people with Down syndrome, 44 (90%) with autosomal-dominant Alzheimer's disease, and 77 (91%) controls also completed an amyloid PET scan within 3 years of tau PET imaging. Spatially, tau PET burden was observed most frequently in subcortical and medial temporal regions in people with Down syndrome, and within the medial temporal lobe in people with autosomal-dominant Alzheimer's disease. Across the brain, people with Down syndrome had greater concentrations of tau for a given level of amyloid compared with people with autosomal-dominant Alzheimer's disease. Temporally, increases in tau were more strongly associated with increases in amyloid for people with Down syndrome compared with autosomal-dominant Alzheimer's disease. INTERPRETATION: Although the general progression of amyloid followed by tau is similar for people Down syndrome and people with autosomal-dominant Alzheimer's disease, we found subtle differences in the spatial distribution, timing, and magnitude of the tau burden between these two cohorts. These differences might have important implications; differences in the temporal pattern of tau accumulation might influence the timing of drug administration in clinical trials, whereas differences in the spatial pattern and magnitude of tau burden might affect disease progression. FUNDING: None.

Pleiotropic Nanostructures Built from l-Histidine Show Biologically Relevant Multicatalytic Activities.

The essential amino acid histidine plays a central role in the manifestation of several metabolic processes, including protein synthesis, enzyme-catalysis, and key biomolecular interactions. However, excess accumulation of histidine causes histidinemia, which shows brain-related medical complications, and the molecular mechanism of such histidine-linked complications is largely unknown. Here, we show that histidine undergoes a self-assembly process, leading to the formation of amyloid-like cytotoxic and catalytically active nanofibers. The kinetics of histidine self-assembly was favored in the presence of Mg(II) and Co(II) ions. Molecular dynamics data showed that preferential noncovalent interactions dominated by H-bonds between histidine molecules facilitate the formation of histidine nanofibers. The histidine nanofibers induced amyloid cross-seeding reactions in several proteins and peptides including pathogenic Aß1-42 and brain extract components. Further, the histidine nanofibers exhibited oxidase activity and enhanced the oxidation of neurotransmitters. Cell-based studies confirmed the cellular internalization of histidine nanofibers in SH-SY5Y cells and subsequent cytotoxic effects through necrosis and apoptosis-mediated cell death. Since several complications including behavioral abnormality, developmental delay, and neurological disabilities are directly linked to abnormal accumulation of histidine, our findings provide a foundational understanding of the mechanism of histidine-related complications. Further, the ability of histidine nanofibers to catalyze amyloid seeding and oxidation reactions is equally important for both biological and materials science research.

Combining Blood-Based Biomarkers and Structural MRI Measurements to Distinguish Persons with and without Significant Amyloid Plaques.

Background: Amyloid-ß (Aß) plaques play a pivotal role in Alzheimer's disease. The current positron emission tomography (PET) is expensive and limited in availability. In contrast, blood-based biomarkers (BBBMs) show potential for characterizing Aß plaques more affordably. We have previously proposed an MRI-based hippocampal morphometry measure to be an indicator of Aß plaques. Objective: To develop and validate an integrated model to predict brain amyloid PET positivity combining MRI feature and plasma Aß42/40 ratio. Methods: We extracted hippocampal multivariate morphometry statistics from MR images and together with plasma Aß42/40 trained a random forest classifier to perform a binary classification of participant brain amyloid PET positivity. We evaluated the model performance using two distinct cohorts, one from the Alzheimer's Disease Neuroimaging Initiative (ADNI) and the other from the Banner Alzheimer's Institute (BAI), including prediction accuracy, precision, recall rate, F1 score, and AUC score. Results: Results from ADNI (mean age 72.6, Aß+ rate 49.5%) and BAI (mean age 66.2, Aß+ rate 36.9%) datasets revealed the integrated multimodal (IMM) model's superior performance over unimodal models. The IMM model achieved prediction accuracies of 0.86 in ADNI and 0.92 in BAI, surpassing unimodal models based solely on structural MRI (0.81 and 0.87) or plasma Aß42/40 (0.73 and 0.81) predictors. CONCLUSIONS: Our IMM model, combining MRI and BBBM data, offers a highly accurate approach to predict brain amyloid PET positivity. This innovative multiplex biomarker strategy presents an accessible and cost-effective avenue for advancing Alzheimer's disease diagnostics, leveraging diverse pathologic features related to Aß plaques and structural MRI.

How ATP suppresses the fibrillation of amyloid peptides: analysis of the free-energy contributions.

Recent experiments have revealed that adenosine triphosphate (ATP) suppresses the fibrillation of amyloid peptides - a process closely linked to neurodegenerative diseases such as Alzheimer's and Parkinson's. Apart from the adsorption of ATP onto amyloid peptides, the molecular understanding is still limited, leaving the underlying mechanism for the fibrillation suppression by ATP largely unclear, especially in regards to the molecular energetics. Here we provide an explanation at the molecular scale by quantifying the free energies using all-atom molecular dynamics simulations. We found that the changes of the free energies due to the addition of ATP lead to a significant equilibrium shift towards monomeric peptides in agreement with experiments. Despite ATP being a highly charged species, the decomposition of the free energies reveals that the van der Waals interactions with the peptide are decisive in determining the relative stabilization of the monomeric state. While the phosphate moiety exhibits strong electrostatic interactions, the compensation by the water solvent results in a minor, overall Coulomb contribution. Our quantitative analysis of the free energies identifies which intermolecular interactions are responsible for the suppression of the amyloid fibril formation by ATP and offers a promising method to analyze the roles of similarly complex cosolvents in aggregation processes.

Antibody-mediated targeting of human microglial leukocyte Ig-like receptor B4 attenuates amyloid pathology in a mouse model.

Microglia help limit the progression of Alzheimer's disease (AD) by constraining amyloid-ß (Aß) pathology, effected through a balance of activating and inhibitory intracellular signals delivered by distinct cell surface receptors. Human leukocyte Ig-like receptor B4 (LILRB4) is an inhibitory receptor of the immunoglobulin (Ig) superfamily that is expressed on myeloid cells and recognizes apolipoprotein E (ApoE) among other ligands. Here, we find that LILRB4 is highly expressed in the microglia of patients with AD. Using mice that accumulate Aß and carry a transgene encompassing a portion of the LILR region that includes LILRB4, we corroborated abundant LILRB4 expression in microglia wrapping around Aß plaques. Systemic treatment of these mice with an anti-human LILRB4 monoclonal antibody (mAb) reduced Aß load, mitigated some Aß-related behavioral abnormalities, enhanced microglia activity, and attenuated expression of interferon-induced genes. In vitro binding experiments established that human LILRB4 binds both human and mouse ApoE and that anti-human LILRB4 mAb blocks such interaction. In silico modeling, biochemical, and mutagenesis analyses identified a loop between the two extracellular Ig domains of LILRB4 required for interaction with mouse ApoE and further indicated that anti-LILRB4 mAb may block LILRB4-mApoE by directly binding this loop. Thus, targeting LILRB4 may be a potential therapeutic avenue for AD.

Solid-state nuclear magnetic resonance in the structural study of polyglutamine aggregation.

The aggregation of proteins into amyloid-like fibrils is seen in many neurodegenerative diseases. Recent years have seen much progress in our understanding of these misfolded protein inclusions, thanks to advances in techniques such as solid-state nuclear magnetic resonance (ssNMR) spectroscopy and cryogenic electron microscopy (cryo-EM). However, multiple repeat-expansion-related disorders have presented special challenges to structural elucidation. This review discusses the special role of ssNMR analysis in the study of protein aggregates associated with CAG repeat expansion disorders. In these diseases, the misfolding and aggregation affect mutant proteins with expanded polyglutamine segments. The most common disorder, Huntington's disease (HD), is connected to the mutation of the huntingtin protein. Since the discovery of the genetic causes for HD in the 1990s, steady progress in our understanding of the role of protein aggregation has depended on the integrative and interdisciplinary use of multiple types of structural techniques. The heterogeneous and dynamic features of polyQ protein fibrils, and in particular those formed by huntingtin N-terminal fragments, have made these aggregates into challenging targets for structural analysis. ssNMR has offered unique insights into many aspects of these amyloid-like aggregates. These include the atomic-level structure of the polyglutamine core, but also measurements of dynamics and solvent accessibility of the non-core flanking domains of these fibrils' fuzzy coats. The obtained structural insights shed new light on pathogenic mechanisms behind this and other protein misfolding diseases.

Novel photocatalytic carbon dots: efficiently inhibiting amyloid aggregation and quickly disaggregating amyloid aggregates.

Amyloid aggregation is implicated in the pathogenesis of various neurodegenerative disorders, such as Alzheimer's disease (AD) and Parkinson's disease (PD). It is critical to develop high-performance drugs to combat amyloid-related diseases. Most identified nanomaterials exhibit limited biocompatibility and therapeutic efficacy. In this work, we used a solvent-free carbonization process to prepare new photo-responsive carbon nanodots (CNDs). The surface of the CNDs is densely packed with chemical groups. CNDs with large, conjugated domains can interact with proteins through π-π stacking and hydrophobic interactions. Furthermore, CNDs possess the ability to generate singlet oxygen species (1O2) and can be used to oxidize amyloid. The hydrophobic interaction and photo-oxidation can both influence amyloid aggregation and disaggregation. Thioflavin T (ThT) fluorescence analysis and circular dichroism (CD) spectroscopy indicate that CNDs can block the transition of amyloid from an α-helix structure to a ß-sheet structure. CNDs demonstrate efficacy in alleviating cytotoxicity induced by Aß42 and exhibit promising blood-brain barrier (BBB) permeability. CNDs have small size, low biotoxicity, good fluorescence and photocatalytic properties, and provide new ideas for the diagnosis and treatment of amyloid-related diseases.

Atomic force microscopy 3D structural reconstruction of individual particles in the study of amyloid protein assemblies.

Recent developments in atomic force microscopy (AFM) image analysis have made three-dimensional (3D) structural reconstruction of individual particles observed on 2D AFM height images a reality. Here, we review the emerging contact point reconstruction AFM (CPR-AFM) methodology and its application in 3D reconstruction of individual helical amyloid filaments in the context of the challenges presented by the structural analysis of highly polymorphous and heterogeneous amyloid protein structures. How individual particle-level structural analysis can contribute to resolving the amyloid polymorph structure-function relationships, the environmental triggers leading to protein misfolding and aggregation into amyloid species, the influences by the conditions or minor fluctuations in the initial monomeric protein structure on the speed of amyloid fibril formation, and the extent of the different types of amyloid species that can be formed, are discussed. Future perspectives in the capabilities of AFM-based 3D structural reconstruction methodology exploiting synergies with other recent AFM technology advances are also discussed to highlight the potential of AFM as an emergent general, accessible and multimodal structural biology tool for the analysis of individual biomolecules.

[New Development in Amyloidosis Research Based on Supersaturation: Biological Factors to Induce/Inhibit the Amyloid Fibril Formation].

Amyloid fibril formation is a general property of proteins and peptides. It is a physicochemical phenomenon similar to crystallization, in which amyloid precursor proteins exceeding solubility precipitate through the breakdown of supersaturation. Using the ultrasonication-forced amyloid fibril inducer HANABI, we have discovered that serum albumin acts as an inhibitor in dialysis-related amyloidosis. Exploring the factors that induce or inhibit amyloid fibril formation using HANABI can lead to the development of early diagnosis and prevention methods for amyloidosis.

[The Roles of Aß in Alzheimer's Disease: In Light of the Latest Findings].

The 'amyloid hypothesis', initially put forward in 1992, posits that amyloid ß protein (Aß) contributes to neurodegeneration through aberrant aggregation. In the process of this aberrant aggregation, Aß forms oligomers, protofibrils, and mature fibrils, ultimately developing plaques. These mature fibrils and plaques were believed to be the culprits behind the neurotoxicity and neurodegeneration seen in Alzheimer's disease (AD). However, growing evidence in recent years has led to the 'Aß oligomer hypothesis', which suggests that the intermediate forms of aggregates, such as oligomers and protofibrils, exhibit stronger neurotoxicity than the mature forms. Consequently, efforts have been made to develop anti-Aß antibody drugs that specifically target these intermediate aggregates. Such interventions hold promise as disease-modifying treatments for AD.

Preclinical evaluation of Tc-99m p5+14 peptide for SPECT detection of cardiac amyloidosis.

INTRODUCTION: Amyloid deposition is a cause of restrictive cardiomyopathy. Patients who present with cardiac disease can be evaluated for transthyretin (TTR)-associated cardiac amyloidosis using nuclear imaging with 99mTc-labeled pyrophosphate (PYP); however, light chain-associated (AL) cardiac amyloid is generally not detected using this tracer. As an alternative, the amyloid-binding peptide p5+14 radiolabeled with iodine-124 has been shown to be an effective pan-amyloid radiotracer for PET/CT imaging. Here, a 99mTc-labeled form of p5+14 peptide has been prepared to facilitate SPECT/CT imaging of cardiac amyloidosis. METHOD: A synthesis method suitable for clinical applications has been used to prepare 99mTc-labeled p5+14 and tested for peptide purity, product bioactivity, radiochemical purity and stability. The product was compared with99mTc-PYP for cardiac SPECT/CT imaging in a mouse model of AA amyloidosis and for reactivity with human tissue sections from AL and TTR patients. RESULTS: The 99mTc p5+14 tracer was produced with >95% yields in radiopurity and bioactivity with no purification steps required and retained over 95% peptide purity and >90% bioactivity for >3 h. In mice, the tracer detected hepatosplenic AA amyloid as well as heart deposits with uptake ~5 fold higher than 99mTc-PYP. 99mTc p5+14 effectively bound human amyloid deposits in the liver, kidney and both AL- and ATTR cardiac amyloid in tissue sections in which 99mTc-PYP binding was not detectable. CONCLUSION: 99mTc-p5+14 was prepared in minutes in >20 mCi doses with good performance in preclinical studies making it suitable for clinical SPECT/CT imaging of cardiac amyloidosis.

Adenosine triphosphate induces amorphous aggregation of amyloid ß by increasing Aß dynamics.

Amyloid ß (Aß) aggregates into two distinct fibril and amorphous forms in the brains of patients with Alzheimer's disease. Adenosine triphosphate (ATP) is a biological hydrotrope that causes Aß to form amorphous aggregates and inhibit fibril formation at physiological concentrations. Based on diffracted X-ray blinking (DXB) analysis, the dynamics of Aß significantly increased immediately after ATP was added compared to those in the absence and presence of ADP and AMP, and the effect diminished after 30 min as the aggregates formed. In the presence of ATP, the ß-sheet content of Aß gradually increased from the beginning, and in the absence of ATP, the content increased rapidly after 180 min incubation, as revealed by a time-dependent thioflavin T fluorescence assay. Images of an atomic force microscope revealed that ATP induces the formation of amorphous aggregates with an average diameter of less than 100 nm, preventing fibrillar formation during 4 days of incubation at 37 °C. ATP may induce amorphous aggregation by increasing the dynamics of Aß, and as a result, the other aggregation pathway is omitted. Our results also suggest that DXB analysis is a useful method to evaluate the inhibitory effect of fibrillar formation.

Remodeling mechanism of gel network structure of soy protein isolate amyloid fibrils mediated by cellulose nanocrystals.

The differences in the gelling properties of soy protein isolate (SPI) and soy protein isolate amyloid fibrils (SAFs) as well as the role of cellulose nanocrystals (CNC) in regulating their gel behaviors were investigated in this study. The binding of CNC to ß-conglycinin (7S), glycinin (11S), and SAFs was predominantly driven by non-covalent interactions. CNC addition reduced the particle size, turbidity, subunit segments, and crystallinity of SPI and SAFs, promoted the conversion of α-helix to ß-sheet, improved the thermal stability, exposed more tyrosine and tryptophan residues, and enhanced the intermolecular interactions. A more regular and ordered lamellar network structure was formed in the SAFs-CNC composite gel, which could be conducive to the improvement of gel quality. This study would provide theoretical reference for the understanding of the regulatory mechanism of protein amyloid fibrils gelation as well as the high-value utilization of SAFs-CNC complex as a functional protein-based material or food ingredient in food field.

Amyloid PET Imaging: Standard Procedures and Semiquantification.

Amyloid plaques are a neuropathologic hallmark of Alzheimer's disease (AD), which can be imaged through positron emission tomography (PET) technology using radiopharmaceuticals that selectively bind to the fibrillar aggregates of amyloid-ß plaques (Amy-PET). Several radiotracers for amyloid PET have been validated (11C-Pittsburgh compound B and the 18F-labeled compounds such as 18F-florbetaben, 18F-florbetapir, and 18F-flutemetamol). Images can be interpreted by means of visual/qualitative, semiquantitative, and quantitative criteria. Here, we summarize the main differences between the available radiotracers for Amy-PET, the proposed interpretation criteria, and main proposed quantification methods.

Histological Typing in Patients With Cardiac Amyloidosis: JACC Review Topic of the Week.

Cardiac amyloidosis is increasingly recognized as a treatable form of heart failure. Highly effective specific therapies have recently become available for the 2 most frequent forms of cardiac amyloidosis: immunoglobulin light chain amyloidosis and transthyretin (ATTR) amyloidosis. Nevertheless, initiation of specific therapies requires recognition of cardiac amyloidosis and appropriate characterization of the amyloid type. Although noninvasive diagnosis is possible for ATTR cardiac amyloidosis, histological demonstration and typing of amyloid deposits is still required for a substantial number of patients with ATTR and in all patients with light chain amyloidosis and other rarer forms of cardiac amyloidosis. Amyloid histological typing can be performed using different techniques: mass spectrometry, immunohistochemistry, and immunoelectron microscopy. This review describes which patients require histological confirmation of cardiac amyloidosis along with when and how to type amyloid deposits in histologic specimens. Furthermore, it covers the characteristics and limitations of the different typing methods that are available in clinical practice.

Association of Neurotrophic Factors at Midlife With In Vivo Measures of ß-Amyloid and Tau Burden 15 Years Later in Dementia-Free Adults.

BACKGROUND AND OBJECTIVES: Neurotrophic factors (NTFs) play an important role in Alzheimer disease (AD) pathophysiology. Brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF) are important NTFs. However, a direct link of BDNF and VEGF circulating levels with in vivo measures of amyloid-ß (Aß) and tau burden remains to be elucidated. We explored the relationship of BDNF and VEGF serum levels with future brain Aß and tau pathology in a cohort of cognitively healthy, predominantly middle-aged adults and tested for possible effect modifications by sex and menopausal status. METHODS: This cross-sectional analysis was conducted using data from the Framingham Heart Study (FHS), a community-based cohort study. The study sample included cognitively healthy participants from the FHS Offspring and Third-generation cohorts. BDNF and VEGF were measured in the third-generation cohort during examination cycles 2 (2005-2008) and 1 (2002-2005), respectively, and in the offspring cohort during examination cycle 7 (1998-2001). Participants underwent 11C-Pittsburgh compound B amyloid and 18F-Flortaucipir tau-PET imaging (2015-2021). Linear regression models were used to assess the relationship of serum BDNF and VEGF levels with regional tau and global Aß, adjusting for potential confounders. Interactions with sex and menopausal status were additionally tested. RESULTS: The sample included 414 individuals (mean age = 41 ± 9 years; 51% female). Continuous measures of BDNF and VEGF were associated with tau signal in the rhinal region after adjustment for potential confounders (ß = -0.15 ± 0.06, p = 0.018 and ß = -0.19 ± 0.09, p = 0.043, respectively). High BDNF (≥32,450 pg/mL) and VEGF (≥488 pg/mL) levels were significantly related to lower rhinal tau (ß = -0.27 ± 0.11, p = 0.016 and ß = -0.40 ± 0.14, p = 0.004, respectively) and inferior temporal tau (ß = -0.24 ± 0.11, p = 0.028 and ß = -0.26 ± 0.13, p = 0.049, respectively). The BDNF-rhinal tau association was observed only among male individuals. Overall, BDNF and VEGF were not associated with global amyloid; however, high VEGF levels were associated with lower amyloid burden in postmenopausal women (ß = -1.96 ± 0.70, p = 0.013, per 1 pg/mL). DISCUSSION: This study demonstrates a robust association between BDNF and VEGF serum levels with in vivo measures of tau almost 2 decades later. These findings add to mounting evidence from preclinical studies suggesting a role of NTFs as valuable blood biomarkers for AD risk prediction.

Widespread nuclear lamina injuries defeat proteostatic purposes of α-synuclein amyloid inclusions.

Biogenesis of inclusion bodies (IBs) facilitates protein quality control (PQC). Canonical aggresomes execute degradation of misfolded proteins while non-degradable amyloids sequester into insoluble protein deposits. Lewy bodies (LBs) are filamentous amyloid inclusions of α-synuclein, but PQC benefits and drawbacks associated with LB-like IBs remain underexplored. Here, we report that crosstalk between filamentous LB-like IBs and aggresome-like IBs of α-synuclein (Syn-aggresomes) buffer the load, aggregation state, and turnover of the amyloidogenic protein in mouse primary neurons and HEK293T cells. Filamentous LB-like IBs possess unorthodox PQC capacities of self-quarantining α-synuclein amyloids and being degradable upon receding fresh amyloidogenesis. Syn-aggresomes equilibrate biogenesis of filamentous LB-like IBs by facilitating spontaneous degradation of α-synuclein and conditional turnover of disintegrated α-synuclein amyloids. Thus, both types of IB primarily contribute to PQC. Incidentally, the overgrown perinuclear LB-like IBs become degenerative once these are misidentified by BICD2, a cargo-adapter for the cytosolic motor-protein dynein. Microscopy indicates that microtubules surrounding the perinuclear filamentous inclusions are also distorted, misbalancing the cytoskeleton-nucleoskeleton tension leading to widespread lamina injuries. Together, nucleocytoplasmic mixing, DNA damage, and deregulated transcription of stress chaperones defeat the proteostatic purposes of the filamentous amyloids of α-synuclein.

Hairpin trimer transition state of amyloid fibril.

Protein fibril self-assembly is a universal transition implicated in neurodegenerative diseases. Although fibril structure/growth are well characterized, fibril nucleation is poorly understood. Here, we use a computational-experimental approach to resolve fibril nucleation. We show that monomer hairpin content quantified from molecular dynamics simulations is predictive of experimental fibril formation kinetics across a tau motif mutant library. Hairpin trimers are predicted to be fibril transition states; one hairpin spontaneously converts into the cross-beta conformation, templating subsequent fibril growth. We designed a disulfide-linked dimer mimicking the transition state that catalyzes fibril formation, measured by ThT fluorescence and TEM, of wild-type motif - which does not normally fibrillize. A dimer compatible with extended conformations but not the transition-state fails to nucleate fibril at any concentration. Tau repeat domain simulations show how long-range interactions sequester this motif in a mutation-dependent manner. This work implies that different fibril morphologies could arise from disease-dependent hairpin seeding from different loci.