Annexin A11 aggregation in FTLD-TDP type C and related neurodegenerative disease proteinopathies.

TAR DNA-binding protein 43 (TDP-43) is an RNA binding protein found within ribonucleoprotein granules tethered to lysosomes via annexin A11. TDP-43 protein forms inclusions in many neurodegenerative diseases including amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP) and limbic predominant age-related TDP-43 encephalopathy neuropathologic change (LATE-NC). Annexin A11 is also known to form aggregates in ALS cases with pathogenic variants in ANXA11. Annexin A11 aggregation has not been described in sporadic ALS, FTLD-TDP or LATE-NC cases. To explore the relationship between TDP-43 and annexin A11, genetic analysis of 822 autopsy cases was performed to identify rare ANXA11 variants. In addition, an immunohistochemical study of 368 autopsy cases was performed to identify annexin A11 aggregates. Insoluble annexin A11 aggregates which colocalize with TDP-43 inclusions were present in all FTLD-TDP Type C cases. Annexin A11 inclusions were also seen in a small proportion (3-6%) of sporadic and genetic forms of FTLD-TDP types A and B, ALS, and LATE-NC. In addition, we confirm the comingling of annexin A11 and TDP-43 aggregates in an ALS case with the pathogenic ANXA11 p.G38R variant. Finally, we found abundant annexin A11 inclusions as the primary pathologic finding in a case of progressive supranuclear palsy-like frontotemporal dementia with prominent striatal vacuolization due to a novel variant, ANXA11 p.P75S. By immunoblot, FTLD-TDP with annexinopathy and ANXA11 variant cases show accumulation of insoluble ANXA11 including a truncated fragment. These results indicate that annexin A11 forms a diverse and heterogeneous range of aggregates in both sporadic and genetic forms of TDP-43 proteinopathies. In addition, the finding of a primary vacuolar annexinopathy due to ANXA11 p.P75S suggests that annexin A11 aggregation is sufficient to cause neurodegeneration.

Amyloidogenic regions in beta-strands II and III modulate the aggregation and toxicity of SOD1 in living cells.

Mutations in the protein superoxide dismutase-1 (SOD1) promote its misfolding and aggregation, ultimately causing familial forms of the debilitating neurodegenerative disease amyotrophic lateral sclerosis (ALS). Currently, over 220 (mostly missense) ALS-causing mutations in the SOD1 protein have been identified, indicating that common structural features are responsible for aggregation and toxicity. Using in silico tools, we predicted amyloidogenic regions in the ALS-associated SOD1-G85R mutant, finding seven regions throughout the structure. Introduction of proline residues into ß-strands II (I18P) or III (I35P) reduced the aggregation propensity and toxicity of SOD1-G85R in cells, significantly more so than proline mutations in other amyloidogenic regions. The I18P and I35P mutations also reduced the capability of SOD1-G85R to template onto previously formed non-proline mutant SOD1 aggregates as measured by fluorescence recovery after photobleaching. Finally, we found that, while the I18P and I35P mutants are less structurally stable than SOD1-G85R, the proline mutants are less aggregation-prone during proteasome inhibition, and less toxic to cells overall. Our research highlights the importance of a previously underappreciated SOD1 amyloidogenic region in ß-strand II (15QGIINF20) to the aggregation and toxicity of SOD1 in ALS mutants, and suggests that ß-strands II and III may be good targets for the development of SOD1-associated ALS therapies.

HAP40 modulates mutant Huntingtin aggregation and toxicity in Huntington's disease mice.

Huntington's disease (HD) is a monogenic neurodegenerative disease, caused by the CAG trinucleotide repeat expansion in exon 1 of the Huntingtin (HTT) gene. The HTT gene encodes a large protein known to interact with many proteins. Huntingtin-associated protein 40 (HAP40) is one that shows high binding affinity with HTT and functions to maintain HTT conformation in vitro. However, the potential role of HAP40 in HD pathogenesis remains unknown. In this study, we found that the expression level of HAP40 is in parallel with HTT but inversely correlates with mutant HTT aggregates in mouse brains. Depletion of endogenous HAP40 in the striatum of HD140Q knock-in (KI) mice leads to enhanced mutant HTT aggregation and neuronal loss. Consistently, overexpression of HAP40 in the striatum of HD140Q KI mice reduced mutant HTT aggregation and ameliorated the behavioral deficits. Mechanistically, HAP40 preferentially binds to mutant HTT and promotes Lysine 48-linked ubiquitination of mutant HTT. Our results revealed that HAP40 is an important regulator of HTT protein homeostasis in vivo and hinted at HAP40 as a therapeutic target in HD treatment.

Autophagy preferentially degrades non-fibrillar polyQ aggregates.

Aggregation of proteins containing expanded polyglutamine (polyQ) repeats is the cytopathologic hallmark of a group of dominantly inherited neurodegenerative diseases, including Huntington's disease (HD). Huntingtin (Htt), the disease protein of HD, forms amyloid-like fibrils by liquid-to-solid phase transition. Macroautophagy has been proposed to clear polyQ aggregates, but the efficiency of aggrephagy is limited. Here, we used cryo-electron tomography to visualize the interactions of autophagosomes with polyQ aggregates in cultured cells in situ. We found that an amorphous aggregate phase exists next to the radially organized polyQ fibrils. Autophagosomes preferentially engulfed this amorphous material, mediated by interactions between the autophagy receptor p62/SQSTM1 and the non-fibrillar aggregate surface. In contrast, amyloid fibrils excluded p62 and evaded clearance, resulting in trapping of autophagic structures. These results suggest that the limited efficiency of autophagy in clearing polyQ aggregates is due to the inability of autophagosomes to interact productively with the non-deformable, fibrillar disease aggregates.

Neuronal glutathione depletion elevates the Aß42/Aß40 ratio and tau aggregation in Alzheimer's disease mice.

Alzheimer's disease (AD) involves reduced glutathione levels, causing oxidative stress and contributing to neuronal cell death. Our prior research identified diminished glutamate-cysteine ligase catalytic subunit (GCLC) as linked to cell death. However, the effect of GCLC on AD features such as amyloid and tau pathology remained unclear. To address this, we investigated amyloid pathology and tau pathology in mice by combining neuron-specific conditional GCLC knockout mice with amyloid precursor protein (App) knockin (KI) or microtubule-associated protein tau (MAPT) KI mice. Intriguingly, GCLC knockout resulted in an increased Aß42/40 ratio. Additionally, GCLC deficiency in MAPT KI mice accelerated the oligomerization of tau through intermolecular disulfide bonds. These findings suggest that the decline in glutathione levels, due to aging or AD pathology, may contribute to the progression of AD.

Glial fibrillary acidic protein is pathologically modified in Alexander disease.

Here, we describe pathological events potentially involved in the disease pathogenesis of Alexander disease (AxD). This is a primary genetic disorder of astrocyte caused by dominant gain-of-function mutations in the gene coding for an intermediate filament protein glial fibrillary acidic protein (GFAP). Pathologically, this disease is characterized by the upregulation of GFAP and its accumulation as Rosenthal fibers. Although the genetic basis linking GFAP mutations with Alexander disease has been firmly established, the initiating events that promote GFAP accumulation and the role of Rosenthal fibers (RFs) in the disease process remain unknown. Here, we investigate the hypothesis that disease-associated mutations promote GFAP aggregation through aberrant posttranslational modifications. We found high molecular weight GFAP species in the RFs of AxD brains, indicating abnormal GFAP crosslinking as a prominent pathological feature of this disease. In vitro and cell-based studies demonstrate that cystine-generating mutations promote GFAP crosslinking by cysteine-dependent oxidation, resulting in defective GFAP assembly and decreased filament solubility. Moreover, we found GFAP was ubiquitinated in RFs of AxD patients and rodent models, supporting this modification as a critical factor linked to GFAP aggregation. Finally, we found that arginine could increase the solubility of aggregation-prone mutant GFAP by decreasing its ubiquitination and aggregation. Our study suggests a series of pathogenic events leading to AxD, involving interplay between GFAP aggregation and abnormal modifications by GFAP ubiquitination and oxidation. More important, our findings provide a basis for investigating new strategies to treat AxD by targeting abnormal GFAP modifications.

Experimental and computational investigation of the effect of Hsc70 structural variants on inhibiting amylin aggregation.

The misfolding and aggregation of human islet amyloid polypeptide (hIAPP), also known as amylin, have been implicated in the pathogenesis of type 2 diabetes (T2D). Heat shock proteins, specifically, heat shock cognate 70 (Hsc70), are molecular chaperones that protect against hIAPP misfolding and inhibits its aggregation. Nevertheless, there is an incomplete understanding of the mechanistic interactions between Hsc70 domains and hIAPP, thus limiting their potential therapeutic role in diabetes. This study investigates the inhibitory capacities of different Hsc70 variants, aiming to identify the structural determinants that strike a balance between efficacy and cytotoxicity. Our experimental findings demonstrate that the ATPase activity of Hsc70 is not a pivotal factor for inhibiting hIAPP misfolding. We underscore the significance of the C-terminal substrate-binding domain of Hsc70 in inhibiting hIAPP aggregation, emphasizing that the removal of the lid subdomain diminishes the inhibitory effect of Hsc70. Additionally, we employed atomistic discrete molecular dynamics simulations to gain deeper insights into the interaction between Hsc70 variants and hIAPP. Integrating both experimental and computational findings, we propose a mechanism by which Hsc70's interaction with hIAPP monomers disrupts protein-protein connections, primarily by shielding the ß-sheet edges of the Hsc70-ß-sandwich. The distinctive conformational dynamics of the alpha helices of Hsc70 potentially enhance hIAPP binding by obstructing the exposed edges of the ß-sandwich, particularly at the ß5-ß8 region along the alpha helix interface. This, in turn, inhibits fibril growth, and similar results were observed following hIAPP dimerization. Overall, this study elucidates the structural intricacies of Hsc70 crucial for impeding hIAPP aggregation, improving our understanding of the potential anti-aggregative properties of molecular chaperones in diabetes treatment.

Copper enhances aggregational toxicity of mutant huntingtin in a Drosophila model of Huntington's Disease.

Huntington's disease (HD) is a progressive neurodegenerative disorder with clinical presentations of moderate to severe cognitive, motor, and psychiatric disturbances. HD is caused by the trinucleotide repeat expansion of CAG of the huntingtin (HTT) gene. The mutant HTT protein containing pathological polyglutamine (polyQ) extension is prone to misfolding and aggregation in the brain. It has previously been observed that copper and iron concentrations are increased in the striata of post-mortem human HD brains. Although it has been shown that the accumulation of mutant HTT protein can interact with copper, the underlying HD progressive phenotypes due to copper overload remains elusive. Here, in a Drosophila model of HD, we showed that copper induces dose-dependent aggregational toxicity and enhancement of Htt-induced neurodegeneration. Specifically, we found that copper increases mutant Htt aggregation, enhances the accumulation of Thioflavin S positive ß-amyloid structures within Htt aggregates, and consequently alters autophagy in the brain. Administration of copper chelator D-penicillamine (DPA) through feeding significantly decreases ß-amyloid aggregates in the HD pathological model. These findings reveal a direct role of copper in potentiating mutant Htt protein-induced aggregational toxicity, and further indicate the potential impact of environmental copper exposure in the disease onset and progression of HD.

A novel pathological mutant reveals the role of torsional flexibility in the serpin breach in adoption of an aggregation-prone intermediate.

Mutants of alpha-1-antitrypsin cause the protein to self-associate and form ordered aggregates ('polymers') that are retained within hepatocytes, resulting in a predisposition to the development of liver disease. The associated reduction in secretion, and for some mutants, impairment of function, leads to a failure to protect lung tissue against proteases released during the inflammatory response and an increased risk of emphysema. We report here a novel deficiency mutation (Gly192Cys), that we name the Sydney variant, identified in a patient in heterozygosity with the Z allele (Glu342Lys). Cellular analysis revealed that the novel variant was mostly retained as insoluble polymers within the endoplasmic reticulum. The basis for this behaviour was investigated using biophysical and structural techniques. The variant showed a 40% reduction in inhibitory activity and a reduced stability as assessed by thermal unfolding experiments. Polymerisation involves adoption of an aggregation-prone intermediate and paradoxically the energy barrier for transition to this state was increased by 16% for the Gly192Cys variant with respect to the wild-type protein. However, with activation to the intermediate state, polymerisation occurred at a 3.8-fold faster rate overall. X-ray crystallography provided two crystal structures of the Gly192Cys variant, revealing perturbation within the 'breach' region with Cys192 in two different orientations: in one structure it faces towards the hydrophobic core while in the second it is solvent-exposed. This orientational heterogeneity was confirmed by PEGylation. These data show the critical role of the torsional freedom imparted by Gly192 in inhibitory activity and stability against polymerisation.

Domain-specific modulatory effects of phosphomimetic substitutions on liquid-liquid phase separation of tau protein.

Aggregation of tau is one of the major pathogenic events in Alzheimer's disease and several other neurodegenerative disorders. Recent reports demonstrated that tau can condense into liquid droplets that undergo time-dependent transition to a solid-like state, suggesting that liquid condensates may be on the pathway to pathological aggregation of tau. While hyperphosphorylation is a key feature of tau isolated from brains of patients with Alzheimer's disease and other tauopathies, the mechanistic role of phosphorylation in tau liquid-liquid phase separation (LLPS) remains largely unexplored. In an attempt to bridge this gap, here we performed systematic studies by introducing phosphomimetic substitutions of Ser/Thr residues with negatively charged Asp/Glu residues in different regions of the protein. Our data indicate that the phosphorylation patterns that increase the polarization of charge distribution in full-length tau (tau441) promote protein LLPS, whereas those that decrease charge polarization have an opposite effect. Overall, this study further supports the notion that tau LLPS is driven by attractive intermolecular electrostatic interactions between the oppositely charged domains. We also show that the phosphomimetic tau variants with low intrinsic propensity for LLPS can be efficiently recruited to droplets formed by the variants with high LLPS propensity. Furthermore, the present data demonstrate that phosphomimetic substitutions have a major effect on time-dependent material properties of tau droplets, generally slowing down their aging. The latter effect is most dramatic for the tau variant with substitutions within the repeat domain, which correlates with the decreased fibrillation rate of this variant.

Enterovirus D68 Infection Induces TDP-43 Cleavage, Aggregation, and Neurotoxicity.

Enterovirus D68 (EV-D68), which causes severe respiratory diseases and irreversible central nervous system damage, has become a serious public health problem worldwide. However, the mechanisms by which EV-D68 exerts neurotoxicity remain unclear. Thus, we aimed to analyze the effects of EV-D68 infection on the cleavage, subcellular translocation, and pathogenic aggregation of TAR DNA-binding protein 43 kDa (TDP-43) in respiratory or neural cells. The results showed that EV-D68-encoded proteases 2A and 3C induced TDP-43 translocation and cleavage, respectively. Specifically, 3C cleaved residue 327Q of TDP-43. The 3C-mediated cleaved TDP-43 fragments had substantially decreased protein solubility compared with the wild-type TDP-43. Hence, 3C activity promoted TDP-43 aggregation, which exerted cytotoxicity to diverse human cells, including glioblastoma T98G cells. The effects of commercially available antiviral drugs on 3C-mediated TDP-43 cleavage were screened, and the results revealed lopinavir as a potent inhibitor of EV-D68 3C protease. Overall, these results suggested TDP-43 as a conserved host target of EV-D68 3C. This study is the first to provide evidence on the involvement of TDP-43 dysregulation in EV-D68 pathogenesis. IMPORTANCE Over the past decade, the incidence of enterovirus D68 (EV-D68) infection has increased worldwide. EV-D68 infection can cause different respiratory symptoms and severe neurological complications, including acute flaccid myelitis. Thus, elucidating the mechanisms underlying EV-D68 toxicity is important to develop novel methods to prevent EV-D68 infection-associated diseases. This study shows that EV-D68 infection triggers the translocalization, cleavage, and aggregation of TDP-43, an intracellular protein closely related to degenerative neurological disorders. The viral protease 3C decreased TDP-43 solubility, thereby exerting cytotoxicity to host cells, including human glioblastoma cells. Thus, counteracting 3C activity is an effective strategy to relieve EV-D68-triggered cell death. Cytoplasmic aggregation of TDP-43 is a hallmark of degenerative diseases, contributing to neural cell damage and central nervous system (CNS) disorders. The findings of this study on EV-D68-induced TDP-43 formation extend our understanding of virus-mediated cytotoxicity and the potential risks of TDP-43 dysfunction-related cognitive impairment and neurological symptoms in infected patients.

ACTN2 Mutant Causes Proteopathy in Human iPSC-Derived Cardiomyocytes.

Genetic variants in α-actinin-2 (ACTN2) are associated with several forms of (cardio)myopathy. We previously reported a heterozygous missense (c.740C>T) ACTN2 gene variant, associated with hypertrophic cardiomyopathy, and characterized by an electro-mechanical phenotype in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Here, we created with CRISPR/Cas9 genetic tools two heterozygous functional knock-out hiPSC lines with a second wild-type (ACTN2wt) and missense ACTN2 (ACTN2mut) allele, respectively. We evaluated their impact on cardiomyocyte structure and function, using a combination of different technologies, including immunofluorescence and live cell imaging, RNA-seq, and mass spectrometry. This study showed that ACTN2mut presents a higher percentage of multinucleation, protein aggregation, hypertrophy, myofibrillar disarray, and activation of both the ubiquitin-proteasome system and the autophagy-lysosomal pathway as compared to ACTN2wt in 2D-cultured hiPSC-CMs. Furthermore, the expression of ACTN2mut was associated with a marked reduction of sarcomere-associated protein levels in 2D-cultured hiPSC-CMs and force impairment in engineered heart tissues. In conclusion, our study highlights the activation of proteolytic systems in ACTN2mut hiPSC-CMs likely to cope with ACTN2 aggregation and therefore directs towards proteopathy as an additional cellular pathology caused by this ACTN2 variant, which may contribute to human ACTN2-associated cardiomyopathies.

ALS mutations in the TIA-1 prion-like domain trigger highly condensed pathogenic structures.

T cell intracellular antigen-1 (TIA-1) plays a central role in stress granule (SG) formation by self-assembly via the prion-like domain (PLD). In the TIA-1 PLD, amino acid mutations associated with neurodegenerative diseases, such as amyotrophic lateral sclerosis (ALS) or Welander distal myopathy (WDM), have been identified. However, how these mutations affect PLD self-assembly properties has remained elusive. In this study, we uncovered the implicit pathogenic structures caused by the mutations. NMR analysis indicated that the dynamic structures of the PLD are synergistically determined by the physicochemical properties of amino acids in units of five residues. Molecular dynamics simulations and three-dimensional electron crystallography, together with biochemical assays, revealed that the WDM mutation E384K attenuated the sticky properties, whereas the ALS mutations P362L and A381T enhanced the self-assembly by inducing ß-sheet interactions and highly condensed assembly, respectively. These results suggest that the P362L and A381T mutations increase the likelihood of irreversible amyloid fibrillization after phase-separated droplet formation, and this process may lead to pathogenicity.

A rare natural lipid induces neuroglobin expression to prevent amyloid oligomers toxicity and retinal neurodegeneration.

Most neurodegenerative diseases such as Alzheimer's disease are proteinopathies linked to the toxicity of amyloid oligomers. Treatments to delay or cure these diseases are lacking. Using budding yeast, we report that the natural lipid tripentadecanoin induces expression of the nitric oxide oxidoreductase Yhb1 to prevent the formation of protein aggregates during aging and extends replicative lifespan. In mammals, tripentadecanoin induces expression of the Yhb1 orthologue, neuroglobin, to protect neurons against amyloid toxicity. Tripentadecanoin also rescues photoreceptors in a mouse model of retinal degeneration and retinal ganglion cells in a Rhesus monkey model of optic atrophy. Together, we propose that tripentadecanoin affects p-bodies to induce neuroglobin expression and offers a potential treatment for proteinopathies and retinal neurodegeneration.

The effect of mutation on an aggregation-prone protein: An in vivo, in vitro, and in silico analysis.

Aggregation of initially stably structured proteins is involved in more than 20 human amyloid diseases. Despite intense research, however, how this class of proteins assembles into amyloid fibrils remains poorly understood, principally because of the complex effects of amino acid substitutions on protein stability, solubility, and aggregation propensity. We address this question using ß2-microglobulin (ß2m) as a model system, focusing on D76N-ß2m that is involved in hereditary amyloidosis. This amino acid substitution causes the aggregation-resilient wild-type protein to become highly aggregation prone in vitro, although the mechanism by which this occurs remained elusive. Here, we identify the residues key to protecting ß2m from aggregation by coupling aggregation with antibiotic resistance in E. coli using a tripartite ß-lactamase assay (TPBLA). By performing saturation mutagenesis at three different sites (D53X-, D76X-, and D98X-ß2m) we show that residue 76 has a unique ability to drive ß2m aggregation in vivo and in vitro. Using a randomly mutated D76N-ß2m variant library, we show that all of the mutations found to improve protein behavior involve residues in a single aggregation-prone region (APR) (residues 60 to 66). Surprisingly, no correlation was found between protein stability and protein aggregation rate or yield, with several mutations in the APR decreasing aggregation without affecting stability. Together, the results demonstrate the power of the TPBLA to develop proteins that are resilient to aggregation and suggest a model for D76N-ß2m aggregation involving the formation of long-range couplings between the APR and Asn76 in a nonnative state.

α-Synuclein phosphorylation at serine 129 occurs after initial protein deposition and inhibits seeded fibril formation and toxicity.

α-Synuclein (α-syn) phosphorylation at serine 129 (pS129­α-syn) is substantially increased in Lewy body disease, such as Parkinson's disease (PD) and dementia with Lewy bodies (DLB). However, the pathogenic relevance of pS129­α-syn remains controversial, so we sought to identify when pS129 modification occurs during α-syn aggregation and its role in initiation, progression and cellular toxicity of disease. Using diverse aggregation assays, including real-time quaking-induced conversion (RT-QuIC) on brain homogenates from PD and DLB cases, we demonstrated that pS129­α-syn inhibits α-syn fibril formation and seeded aggregation. We also identified lower seeding propensity of pS129­α-syn in cultured cells and correspondingly attenuated cellular toxicity. To build upon these findings, we developed a monoclonal antibody (4B1) specifically recognizing nonphosphorylated S129­α-syn (WT­α-syn) and noted that S129 residue is more efficiently phosphorylated when the protein is aggregated. Using this antibody, we characterized the time-course of α-syn phosphorylation in organotypic mouse hippocampal cultures and mice injected with α-syn preformed fibrils, and we observed aggregation of nonphosphorylated α-syn followed by later pS129­α-syn. Furthermore, in postmortem brain tissue from PD and DLB patients, we observed an inverse relationship between relative abundance of nonphosphorylated α-syn and disease duration. These findings suggest that pS129­α-syn occurs subsequent to initial protein aggregation and apparently inhibits further aggregation. This could possibly imply a potential protective role for pS129­α-syn, which has major implications for understanding the pathobiology of Lewy body disease and the continued use of reduced pS129­α-syn as a measure of efficacy in clinical trials.

The Pathological G51D Mutation in Alpha-Synuclein Oligomers Confers Distinct Structural Attributes and Cellular Toxicity.

A wide variety of oligomeric structures are formed during the aggregation of proteins associated with neurodegenerative diseases. Such soluble oligomers are believed to be key toxic species in the related disorders; therefore, identification of the structural determinants of toxicity is of upmost importance. Here, we analysed toxic oligomers of α-synuclein and its pathological variants in order to identify structural features that could be related to toxicity and found a novel structural polymorphism within G51D oligomers. These G51D oligomers can adopt a variety of ß-sheet-rich structures with differing degrees of α-helical content, and the helical structural content of these oligomers correlates with the level of induced cellular dysfunction in SH-SY5Y cells. This structure-function relationship observed in α-synuclein oligomers thus presents the α-helical structure as another potential structural determinant that may be linked with cellular toxicity in amyloid-related proteins.

Elevated constitutive expression of Hsp40 chaperone Sis1 reduces TDP-43 aggregation-induced oxidative stress in Ire1 pathway dependent-manner in yeast TDP-43 proteinopathy model of amyotrophic lateral sclerosis.

Oxidative stress is a therapeutic target in TDP-43 proteinopathies like amyotrophic lateral sclerosis (ALS) and FTLD-TDP. TDP-43 over-expression causes oxidative stress in yeast model of ALS. Previously, we developed a red/white color conversion reporter assay using ade1 or ade2 mutant yeast to examine oxidative stress induced by expression of amyloidogenic proteins. Also, a previous study showed that overexpression of yeast Hsp40 chaperone Sis1 could mitigate the toxicity and proteosomal blockage induced by TDP-43 over-expression. Here, using the red/white reporter yeast assay and also by CellROX-staining, we found that an elevated expression of Sis1 mitigates the TDP-43-induced oxidative stress. Furthermore, as redox signalling and the ER stress response pathways cross-talk, we checked if the Sis1-mediated mitigation of the TDP-43-induced oxidative stress can also be observed in yeast deleted for ER stress response gene, IRE1. We find that in the yeast deleted for the IRE1 gene, the elevated expression of Sis1 fails to neutralize the TDP-43-induced oxidative stress. Taken together, Hsp40 chaperone modulation can be further examined towards therapeutic research on the TDP-43 proteinopathies.

Congenital cataract-causing mutation ßB1-L116P is prone to amyloid fibrils aggregation and protease degradation with low structural stability.

Congenital cataract, a common disease with lens opacification, causes blindness in the newborn worldwide and is mainly caused by abnormal aggregation of crystallin. As the main structural protein in the mammalian lens, ßB1-crystallin has an important role in the maintenance of lens transparency. Recently, the L116P mutation in ßB1-CRY was found in a Chinese family with congenital nuclear cataracts, while its underlying pathogenic mechanism remains unclear. In the current study, the ßB1 wild-type protein was purified, and the mutated form, ßB1-L116P, was examined for examining the effect on structural stability and susceptibility against environmental stresses. Our results reveal low solubility and structural stability of ßB1-L116P at physiological temperature, which markedly impaired the protein structure and the oligomerization of ßB1-crystallin. Under guanidine hydrochloride-induced denaturing conditions, ßB1-L116P mutation perturbed the protein unfolding process, making it prone to amyloid fibrils aggregation. More importantly, the L116P mutation increased susceptibility of ßB1-crystallin against UV radiation. ßB1-L116P overexpression led to the formation of more serious intracellular aggresomes under UV radiation or oxidative stress. Furthermore, the ßB1-L116P mutation increased the sensitivity to the proteolysis process. These results indicate that the low structural stability, susceptibility to amyloid fibrils aggregation, and protease degradation of ßB1-L116P may contribute to cataract development and associated symptoms.

Neurotoxic or neuroprotective: Post-translational modifications of α-synuclein at the cross-roads of functions.

Parkinson's disease is the second most prevalent neurodegenerative disease. The loss of dopaminergic neurons in the substantia nigra is one of the pathological hallmarks of PD. PD also belongs to the class of neurodegenerative disease known as 'Synucleinopathies' as α-synuclein is responsible for disease development. The presence of aggregated α-synuclein associated with other proteins found in the Lewy bodies and Lewy neurites in the substantia nigra and other regions of the brain including locus ceruleus, dorsal vagal nucleus, nucleus basalis of Meynert and cerebral cortex is one of the central events for PD development. The complete biological function of α-synuclein is still debated. Besides its ability to propagate, it undergoes various post-translational modifications which play a paramount role in PD development and progression. Also, the aggregation of α-synuclein is modulated by various post-translational modifications. Here, we present a summary of multiple PTMs involved in the modulation of α-synuclein directly or indirectly and to identify their neuroprotective or neurotoxic roles, which might act as potential therapeutic targets for Parkinson's disease.