bims-proned 2024-09-22 papers

bims-proned

Biomed News

on Proteostasis in neurodegeneration

Issue of 2024–09–22
23 papers selected by
Verena Kohler, Umeå University

HSPB6: A lipid-dependent molecular chaperone inhibits α-synuclein aggregation.
Live-cell visualization of tau aggregation in human neurons.
Protein tyrosine phosphatase receptor type O serves as a key regulator of insulin resistance-induced α-synuclein aggregation in Parkinson's disease.
Untangling the complexity and impact of tau protein ubiquitination.
Pathological C-terminal phosphomimetic substitutions alter the mechanism of liquid-liquid phase separation of TDP-43 low complexity domain.
Viral overexpression of human alpha-synuclein in mouse substantia nigra dopamine neurons results in hyperdopaminergia but no neurodegeneration.
Co-opting templated aggregation to degrade pathogenic tau assemblies and improve motor function.
Temporal control of acute protein aggregate turnover by UBE3C and NRF1-dependent proteasomal pathways.
Targeting ferroptosis with the lipoxygenase inhibitor PTC-041 as a therapeutic strategy for the treatment of Parkinson's disease.
Mutation/metal deficiency in the "electrostatic loop" enhanced aggregation process in apo/holo SOD1 variants: implications for ALS diseases.
Involvement of ubiquitination in Alzheimer's disease.
Atomistic Insights into the Stabilization of TDP-43 Protofibrils by ATP.
Modulation of AIE and Intramolecular Charge Transfer of a Pyrene-Based Probe for Discriminatory Detection and Imaging of Oligomers and Amyloid Fibrils.
SUMO1 modification of 0N4R-tau is regulated by PIASx, SENP1, SENP2, and TRIM11.
Membrane-assisted Aβ40 aggregation pathways.
Methods for high throughput discovery of fluoroprobes that recognize tau fibril polymorphs.
Astrocyte-neuron communication through the complement C3-C3aR pathway in Parkinson's disease.
HLH-30/TFEB modulates autophagy to improve proteostasis in Aβ transgenic Caenorhabditis elegans.
Epigenetics in the formation of pathological aggregates in amyotrophic lateral sclerosis.
Spatiotemporal formation of a single liquid-like condensate and amyloid fibrils of α-synuclein by optical trapping at solution surface.
Gigaxonin, mutated in Giant Axonal Neuropathy, interacts with TDP-43 and other RNA binding proteins.
Hallmarks of aging in age-related macular degeneration and age-related neurological disorders: novel insights into common mechanisms and clinical relevance.
Integrating amyloid and tau imaging with proteomics and genomics in Alzheimer's disease.

iScience. 2024 Sep 20. 27(9): 110657

HSPB6: A lipid-dependent molecular chaperone inhibits α-synuclein aggregation.

Valentina Secco, Tatiana Tiago, Roxine Staats, Swapan Preet, Sean Chia, Michele Vendruscolo, Serena Carra.

  The process of protein misfolding and aggregation is associated with various cytotoxic effects. Understanding how this phenomenon is regulated by the protein homeostasis system, however, is difficult, since it takes place through a complex non-linear network of coupled microscopic steps, including primary nucleation, fibril elongation, and secondary nucleation, which depend on environmental factors. To address this problem, we studied how the aggregation of α-synuclein, a protein associated with Parkinson's disease, is modulated by molecular chaperones and lipid membranes. We focused on small heat shock proteins (sHSPs/HSPBs), which interact with proteins and lipids and are upregulated during aging, a major risk factor for protein misfolding diseases. HSPBs act on different microscopic steps to prevent α-synuclein aggregation, with HSPB6 showing a lipid-dependent chaperone activity. Our findings provide an example of how HSPBs diversified their mechanisms of action to reach an efficient regulation of protein misfolding and aggregation within the complex cellular environment.

Keywords:  Biological sciences; Cell biology; Chemistry

DOI:  https://doi.org/10.1016/j.isci.2024.110657
Commun Biol. 2024 Sep 14. 7(1): 1143

Live-cell visualization of tau aggregation in human neurons.

Bryan Hurtle, Christopher J Donnelly, Xin Zhang, Amantha Thathiah.

Alzheimer's disease (AD) and more than twenty other dementias, termed tauopathies, are pathologically defined by insoluble aggregates of the microtubule-associated protein tau (MAPT). Although tau aggregation correlates with AD symptomology, the specific tau species, i.e., monomers, soluble oligomers, and insoluble aggregates that induce neurotoxicity are incompletely understood. We developed a light-responsive tau protein (optoTAU) and used viscosity-sensitive AggFluor probes to investigate the consequence(s) of tau aggregation in human neurons and identify modifiers of tau aggregation in AD and other tauopathies. We determined that optoTAU reproduces biological and structural properties of tau aggregation observed in human brains and the pathophysiological transition in tau solubility in live cells. We also provide proof-of-concept for the utilization of optoTAU as a pharmacological platform to identify modifiers of tau aggregation. These findings have broad implications for the characterization of aggregation-prone proteins and investigation of the complex relationship between protein solubility, cellular function, and disease progression.

DOI: https://doi.org/10.1038/s42003-024-06840-z
Cell Mol Life Sci. 2024 Sep 14. 81(1): 403

Protein tyrosine phosphatase receptor type O serves as a key regulator of insulin resistance-induced α-synuclein aggregation in Parkinson's disease.

Shichuan Tan, Huizhong Chi, Pin Wang, Rongrong Zhao, Qinran Zhang, Zijie Gao, Hao Xue, Qilin Tang, Gang Li.

  Insulin resistance (IR) was found to be a critical element in the pathogenesis of Parkinson's disease (PD), facilitating abnormal α-synuclein (α-Syn) aggregation in neurons and thus promoting PD development. However, how IR contributes to abnormal α-Syn aggregation remains ill-defined. Here, we analyzed six PD postmortem brain transcriptome datasets to reveal module genes implicated in IR-mediated α-Syn aggregation. In addition, we induced IR in cultured dopaminergic (DA) neurons overexpressing α-Syn to identify IR-modulated differentially expressed genes (DEGs). Integrated analysis of data from PD patients and cultured neurons revealed 226 genes involved in α-Syn aggregation under IR conditions, of which 53 exhibited differential expression between PD patients and controls. Subsequently, we conducted an integrated analysis of the 53 IR-modulated genes employing transcriptome data from PD patients with different Braak stages and DA neuron subclasses with varying α-Syn aggregation scores. Protein tyrosine phosphatase receptor type O (PTPRO) was identified to be closely associated with PD progression and α-Syn aggregation. Experimental validation in a cultured PD cell model confirmed that both mRNA and protein of PTPRO were reduced under IR conditions, and the downregulation of PTPRO significantly facilitated α-Syn aggregation and cell death. Collectively, our findings identified PTPRO as a key regulator in IR-mediated α-Syn aggregation and uncovered its prospective utility as a therapeutic target in PD patients with IR.

Keywords:  Dopaminergic neurons; Insulin resistance; PTPRO; Parkinson’s disease; α-Synuclein aggregation

DOI:  https://doi.org/10.1007/s00018-024-05436-4
Chembiochem. 2024 Sep 18. e202400566

Untangling the complexity and impact of tau protein ubiquitination.

Daniele Trivellato, Francesca Munari, Michael Assfalg, Stefano Capaldi, Mariapina D'Onofrio.

  The microtubule-associated protein tau is an intrinsically disordered protein highly expressed in neuronal axons. In healthy neurons, tau regulates microtubule dynamics and neurite outgrowth. However, pathological conditions can trigger aberrant tau aggregation into insoluble filaments, a hallmark of neurodegenerative disorders known as tauopathies. Tau undergoes diverse posttranslational modifications (PTMs), suggesting complex regulation and potentially varied functions. Among PTMs, the role and mechanisms of ubiquitination in physiology and disease have remained enigmatic. The past three decades have witnessed the emergence of key studies on tau protein ubiquitination. In this concept, we discuss how these investigations have begun to shed light on the ubiquitination patterns of physiological and pathological tau, the responsible enzymatic machinery, and the influence of ubiquitination on tau aggregation. We also provide an overview of the semi-synthetic methods that have enabled in vitro investigations of conformational transitions of tau induced by ubiquitin modification. Finally, we discuss future perspectives in the field necessary to elucidate the molecular mechanisms of tau ubiquitination and clearance.

Keywords:  Fibrils; Neurodegeneration; Post-translational modifications; Tau protein; Ubiquitination

DOI:  https://doi.org/10.1002/cbic.202400566
Protein Sci. 2024 Oct;33(10): e5179

Pathological C-terminal phosphomimetic substitutions alter the mechanism of liquid-liquid phase separation of TDP-43 low complexity domain.

Raza Haider, Brandon Shipley, Krystyna Surewicz, Michael Hinczewski, Witold K Surewicz.

  C-terminally phosphorylated TAR DNA-binding protein of 43 kDa (TDP-43) marks the proteinaceous inclusions that characterize a number of age-related neurodegenerative diseases, including amyotrophic lateral sclerosis, frontotemporal lobar degeneration and Alzheimer's disease. TDP-43 phosphorylation at S403/S404 and (especially) at S409/S410 is, in fact, accepted as a biomarker of proteinopathy. These residues are located within the low complexity domain (LCD), which also drives the protein's liquid-liquid phase separation (LLPS). The impact of phosphorylation at these LCD sites on phase separation of the protein is a topic of great interest, as these post-translational modifications and LLPS are both implicated in proteinopathies. Here, we employed a combination of experimental and simulation-based approaches to explore this question on a phosphomimetic model of the TDP-43 LCD. Our turbidity and fluorescence microscopy data show that phosphomimetic Ser-to-Asp substitutions at residues S403, S404, S409 and S410 alter the LLPS behavior of TDP-43 LCD. In particular, unlike the LLPS of unmodified protein, LLPS of the phosphomimetic variants displays a biphasic dependence on salt concentration. Through coarse-grained modeling, we find that this biphasic salt dependence is derived from an altered mechanism of phase separation, in which LLPS-driving short-range intermolecular hydrophobic interactions are modulated by long-range attractive electrostatic interactions. Overall, this in vitro and in silico study provides a physiochemical foundation for understanding the impact of pathologically relevant C-terminal phosphorylation on the LLPS of TDP-43 in a more complex cellular environment.

Keywords:  TDP‐43 phosphorylation; amyotrophic lateral sclerosis; coarse‐grained simulation; electrostatic forces; hydrophobic forces; liquid–liquid phase separation

DOI:  https://doi.org/10.1002/pro.5179
Exp Neurol. 2024 Sep 15. pii: S0014-4886(24)00285-1. [Epub ahead of print]382 114959

Viral overexpression of human alpha-synuclein in mouse substantia nigra dopamine neurons results in hyperdopaminergia but no neurodegeneration.

Sofia Ines Garcia Moreno, Fabian Limani, Iina Ludwig, Catherine Gilbert, Christian Pifl, Thomas S Hnasko, Thomas Steinkellner.

  Loss of select neuronal populations such as midbrain dopamine (DA) neurons is a pathological hallmark of Parkinson's disease (PD). The small neuronal protein α-synuclein has been related both genetically and neuropathologically to PD, yet how and if it contributes to selective vulnerability remains elusive. Here, we describe the generation of a novel adeno-associated viral vector (AAV) for Cre-dependent overexpression of wild-type human α-synuclein. Our strategy allows us to restrict α-synuclein to select neuronal populations and hence investigate the cell-autonomous effects of elevated α-synuclein in genetically-defined cell types. Since DA neurons in the substantia nigra pars compacta (SNc) are particularly vulnerable in PD, we investigated in more detail the effects of increased α-synuclein in these cells. AAV-mediated overexpression of wildtype human α-synuclein in SNc DA neurons increased the levels of α-synuclein within these cells and augmented phosphorylation of α-synuclein at serine-129, which is considered a pathological feature of PD and other synucleinopathies. However, despite abundant α-synuclein overexpression and hyperphosphorylation we did not observe any dopaminergic neurodegeneration up to 90 days post virus infusion. In contrast, we noticed that overexpression of α-synuclein resulted in increased locomotor activity and elevated striatal DA levels suggesting that α-synuclein enhanced dopaminergic activity. We therefore conclude that cell-autonomous effects of elevated α-synuclein are not sufficient to trigger acute dopaminergic neurodegeneration.

Keywords:  Adeno-associated viral vectors; Alpha-synuclein; Animal model of Parkinson's disease; Cre recombinase; Dopamine neuron vulnerability; Parkinson's disease

DOI:  https://doi.org/10.1016/j.expneurol.2024.114959
Cell. 2024 Sep 10. pii: S0092-8674(24)00912-7. [Epub ahead of print]

Co-opting templated aggregation to degrade pathogenic tau assemblies and improve motor function.

Lauren V C Miller, Guido Papa, Marina Vaysburd, Shi Cheng, Paul W Sweeney, Annabel Smith, Catarina Franco, Taxiarchis Katsinelos, Melissa Huang, Sophie A I Sanford, Jonathan Benn, Jasmine Farnsworth, Katie Higginson, Holly Joyner, William A McEwan, Leo C James.

  Protein aggregation causes a wide range of neurodegenerative diseases. Targeting and removing aggregates, but not the functional protein, is a considerable therapeutic challenge. Here, we describe a therapeutic strategy called "RING-Bait," which employs an aggregating protein sequence combined with an E3 ubiquitin ligase. RING-Bait is recruited into aggregates, whereupon clustering dimerizes the RING domain and activates its E3 function, resulting in the degradation of the aggregate complex. We exemplify this concept by demonstrating the specific degradation of tau aggregates while sparing soluble tau. Unlike immunotherapy, RING-Bait is effective against both seeded and cell-autonomous aggregation. RING-Bait removed tau aggregates seeded from Alzheimer's disease (AD) and progressive supranuclear palsy (PSP) brain extracts and was also effective in primary neurons. We used a brain-penetrant adeno-associated virus (AAV) to treat P301S tau transgenic mice, reducing tau pathology and improving motor function. A RING-Bait strategy could be applied to other neurodegenerative proteinopathies by replacing the Bait sequence to match the target aggregate.

Keywords:  Alzheimer’s disease; TRIM21; antibodies; gene therapy; neurobiology; neurodegeneration; protein engineering; targeted protein degradation; tauopathy; ubiquitination

DOI:  https://doi.org/10.1016/j.cell.2024.08.024
bioRxiv. 2024 Sep 03. pii: 2024.08.30.610524. [Epub ahead of print]

Temporal control of acute protein aggregate turnover by UBE3C and NRF1-dependent proteasomal pathways.

Kelsey L Hickey, Alexandra Panov, Enya Miguel Whelan, Tillman Schäfer, Arda Mizrak, Ron R Kopito, Wolfgang Baumeister, Rubén Fernández-Busnadiego, J Wade Harper.

  A hallmark of neurodegenerative diseases is the progressive loss of proteostasis, leading to the accumulation of misfolded proteins or protein aggregates, with subsequent cytotoxicity. To combat this toxicity, cells have evolved degradation pathways (ubiquitin-proteasome system and autophagy) that detect and degrade misfolded proteins. However, studying the underlying cellular pathways and mechanisms has remained a challenge, as formation of many types of protein aggregates is asynchronous, with individual cells displaying distinct kinetics, thereby hindering rigorous time-course studies. Here, we merge a kinetically tractable and synchronous agDD-GFP system for aggregate formation with targeted gene knockdowns, to uncover degradation mechanisms used in response to acute aggregate formation. We find that agDD-GFP forms amorphous aggregates by cryo-electron tomography at both early and late stages of aggregate formation. Aggregate turnover occurs in a proteasome-dependent mechanism in a manner that is dictated by cellular aggregate burden, with no evidence of the involvement of autophagy. Lower levels of misfolded agDD-GFP, enriched in oligomers, utilizes UBE3C-dependent proteasomal degradation in a pathway that is independent of RPN13 ubiquitylation by UBE3C. Higher aggregate burden activates the NRF1 transcription factor to increase proteasome subunit transcription, and subsequent degradation capacity of cells. Loss or gain of NRF1 function alters the turnover of agDD-GFP under conditions of high aggregate burden. Together, these results define the role of UBE3C in degradation of this class of misfolded aggregation-prone proteins and reveals a role for NRF1 in proteostasis control in response to widespread protein aggregation.

Keywords:  Biological Sciences; Cell Biology; NRF1; Protein aggregates; Protein quality control; Protein turnover; UBE3C; Ubiquitin-proteasome system

DOI:  https://doi.org/10.1101/2024.08.30.610524
PLoS One. 2024 ;19(9): e0309893

Targeting ferroptosis with the lipoxygenase inhibitor PTC-041 as a therapeutic strategy for the treatment of Parkinson's disease.

Angela Minnella, Kevin P McCusker, Akiko Amagata, Beatrice Trias, Marla Weetall, Joey C Latham, Sloane O'Neill, Richard K Wyse, Matthew B Klein, Jeffrey K Trimmer.

Parkinson's disease is the second most common neurodegenerative disorder, affecting nearly 10 million people worldwide. Ferroptosis, a recently identified form of regulated cell death characterized by 15-lipoxygenase-mediated hydroperoxidation of membrane lipids, has been implicated in neurodegenerative disorders including amyotrophic lateral sclerosis and Parkinson's disease. Pharmacological inhibition of 15 -lipoxygenase to prevent iron- and lipid peroxidation-associated ferroptotic cell death is a rational strategy for the treatment of Parkinson's disease. We report here the characterization of PTC-041 as an anti-ferroptotic reductive lipoxygenase inhibitor developed for the treatment of Parkinson's disease. In these studies, PTC-041 potently protects primary human Parkinson's disease patient-derived fibroblasts from lipid peroxidation and subsequent ferroptotic cell death and prevents ferroptosis-related neuronal loss and astrogliosis in primary rat neuronal cultures. Additionally, PTC-041 prevents ferroptotic-mediated α-synuclein protein aggregation and nitrosylation in vitro, suggesting a potential role for anti-ferroptotic lipoxygenase inhibitors in mitigating pathogenic aspects of synucleinopathies such as Parkinson's disease. We further found that PTC-041 protects against synucleinopathy in vivo, demonstrating that PTC-041 treatment of Line 61 transgenic mice protects against α-synuclein aggregation and phosphorylation as well as prevents associated neuronal and non-neuronal cell death. Finally, we show that. PTC-041 protects against 6-hydroxydopamine-induced motor deficits in a hemiparkinsonian rat model, further validating the potential therapeutic benefits of lipoxygenase inhibitors in the treatment of Parkinson's disease.

DOI: https://doi.org/10.1371/journal.pone.0309893
BMC Chem. 2024 Sep 19. 18(1): 177

Mutation/metal deficiency in the "electrostatic loop" enhanced aggregation process in apo/holo SOD1 variants: implications for ALS diseases.

Faezeh Ashkaran, Bagher Seyedalipour, Payam Baziyar, Saman Hosseinkhani.

  Despite the many mechanisms it has created to prevent unfolding and aggregation of proteins, many diseases are caused by abnormal folding of proteins, which are called misfolding diseases. During this process, proteins undergo structural changes and become stable, insoluble beta-sheet aggregates called amyloid fibrils. Mutations/disruptions in metal ion homeostasis in the ALS-associated metalloenzyme superoxide dismutase (SOD1) reduce conformational stability, consistent with the protein aggregation hypothesis for neurodegenerative diseases. However, the exact mechanism of involvement is not well understood. Hence, to understand the role of mutation/ metal deficiency in SOD1 misfolding and aggregation, we investigated the effects of apo/holo SOD1 variants on structural properties using biophysical/experimental techniques. The MD results support the idea that the mutation/metal deficiency can lead to a change in conformation. The increased content of β-sheet structures in apo/holo SOD1 variants can be attributed to the aggregation tendency, which was confirmed by FTIR spectroscopy and dictionary of secondary structure in proteins (DSSP) results. Thermodynamic studies of GdnHCl showed that metal deficiency/mutation/intramolecular S-S reduction together are required to initiate misfolding/aggregation of SOD1. The results showed that apo/holo SOD1 variants under destabilizing conditions induced amyloid aggregates at physiological pH, which were detected by ThT/ANS fluorescence, as well as further confirmation of amyloid/amorphous species by TEM. This study confirms that mutations in the electrostatic loop of SOD1 lead to structural abnormalities, including changes in hydrophobicity, reduced disulfide bonds, and an increased propensity for protein denaturation. This process facilitates the formation of amyloid/amorphous aggregates ALS-associated.

Keywords:  Electrostatic loop; FALS; Molecular dynamics simulation; Protein aggregation; SOD1 variants

DOI:  https://doi.org/10.1186/s13065-024-01289-x
Front Neurol. 2024 ;15 1459678

Involvement of ubiquitination in Alzheimer's disease.

Nan Lin, Xi-Yan Gao, Xiao Li, Wen-Ming Chu.

  The hallmark pathological features of Alzheimer's disease (AD) consist of senile plaques, which are formed by extracellular β-amyloid (Aβ) deposition, and neurofibrillary tangles, which are formed by the hyperphosphorylation of intra-neuronal tau proteins. With the increase in clinical studies, the in vivo imbalance of iron homeostasis and the dysfunction of synaptic plasticity have been confirmed to be involved in AD pathogenesis. All of these mechanisms are constituted by the abnormal accumulation of misfolded or conformationally altered protein aggregates, which in turn drive AD progression. Proteostatic imbalance has emerged as a key mechanism in the pathogenesis of AD. Ubiquitination modification is a major pathway for maintaining protein homeostasis, and protein degradation is primarily carried out by the ubiquitin-proteasome system (UPS). In this review, we provide an overview of the ubiquitination modification processes and related protein ubiquitination degradation pathways in AD, focusing on the microtubule-associated protein Tau, amyloid precursor protein (APP), divalent metal transporter protein 1 (DMT1), and α-amino-3-hyroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors. We also discuss recent advances in ubiquitination-based targeted therapy for AD, with the aim of contributing new ideas to the development of novel therapeutic interventions for AD.

Keywords:  AMPARs; APP; Alzheimer’s disease; DMT1; Tua; protein degradation; ubiquitination

DOI:  https://doi.org/10.3389/fneur.2024.1459678
J Chem Inf Model. 2024 Sep 18.

Atomistic Insights into the Stabilization of TDP-43 Protofibrils by ATP.

Zhengdong Xu, Jiaxing Tang, Yehong Gong, Jianxin Zhang, Yu Zou.

The aberrant accumulation of the transactive response deoxyribonucleic acid (DNA)-binding protein of 43 kDa (TDP-43) aggregates in the cytoplasm of motor neurons is the main pathological hallmark of amyotrophic lateral sclerosis (ALS). Previous experiments reported that adenosine triphosphate (ATP), the universal energy currency for all living cells, could induce aggregation and enhance the folding of TDP-43 fibrillar aggregates. However, the significance of ATP on TDP-43 fibrillation and the mechanism behind it remain elusive. In this work, we conducted multiple atomistic molecular dynamics (MD) simulations totaling 20 μs to search the critical nucleus size of TDP-43282-360 and investigate the impact of ATP molecules on preformed protofibrils. The results reveal that the trimer is the critical nucleus for TDP-43282-360 fibril formation and the tetramer is the minimal stable nucleus. When ATP molecules bind to the TDP-43282-360 trimer and tetramer, they can consolidate the TDP-43282-360 protofibrils by increasing the content of the β-sheet structure and promoting the formation of hydrogen bonds (H-bonds). Binding site analyses show that the N-terminus of TDP-43282-360 protofibrils is the main binding site of ATP, and R293 dominates the direct binding of ATP. Further analyses reveal that the π-π, cation-π, salt bridge, and H-bonding interactions together contribute to the binding of ATP to TDP-43282-360 protofibrils. This study decoded the detailed stabilization mechanism of protofibrillar TDP-43282-360 oligomers by ATP, and may provide new avenues for the development of drug design against ALS.

DOI: https://doi.org/10.1021/acs.jcim.4c01140
ACS Appl Bio Mater. 2024 Sep 18.

Modulation of AIE and Intramolecular Charge Transfer of a Pyrene-Based Probe for Discriminatory Detection and Imaging of Oligomers and Amyloid Fibrils.

Dharini Arumugam, Nidhi Anilkumar Jamuna, Adithya Kamalakshan, Sarthak Mandal.

  Oligomers and amyloid fibrils formed at different stages of protein aggregation are important biomarkers for a variety of neurodegenerative diseases including Alzheimer's and Parkinson's diseases. The development of probes for the sensitive detection of oligomeric species is important for early stage diagnosis of amyloidogenic diseases. Many small molecular dyes have been developed to probe the dynamic growth of amyloid fibrils. However, there is a lack of discriminatory detection strategies to monitor the dynamics of both oligomers and amyloid fibrils based on the differential modulation of the photophysical properties of a single dye. Here we report a pyrene-based intramolecular charge transfer (ICT) dye with large Stokes shifted red-emitting aggregation induced emission (AIE) for monitoring the dynamic populations of both oligomers and fibrils during the aggregation of hen egg white lysozyme (HEWL) protein. At the early stage of protein aggregation, the accumulation of HEWL oligomers results in a rapid and substantial increase in the red AIE intensity at 660 nm. Later, as the oligomers transform into mature fibrils, the dye exhibits a distinct photophysical change. Binding of the dye to HEWL fibrils strongly suppresses the red AIE and enhances ICT emission. This is evidenced by a gradual decrease in the AIE intensity (∼660 nm) and an increase in LE (∼490 nm) and ICT (∼540 nm) emission intensities during the later stages of protein aggregation. Thus, the dye provides simultaneous measurements of the population dynamics of both HEWL oligomers and fibrils during protein aggregation based on the discriminatory modulation of AIE and ICT of the dye. The dye also enables imaging of both HEWL oligomers and fibrils simultaneously using different emission channels in super-resolution confocal fluorescence microscopy.

Keywords:  aggregation induced emission; amyloid fibrils; donor−acceptor molecule; intramolecular charge transfer; oligomers detection

DOI:  https://doi.org/10.1021/acsabm.4c00820
Biochem Biophys Rep. 2024 Sep;39 101800

SUMO1 modification of 0N4R-tau is regulated by PIASx, SENP1, SENP2, and TRIM11.

Harmony Wada, Takuma Maruyama, Takako Niikura.

  Tau is a microtubule-associated protein that contributes to cytoskeletal stabilization. Aggregation of tau proteins is associated with neurodegenerative disorders such as Alzheimer's disease. Several types of posttranslational modifications that alter the physical properties of tau proteins have been identified. SUMOylation is a reversible modification of lysine residues by a small ubiquitin-like modifier (SUMO). In this study, we examined the enzymes that regulate the SUMOylation and deSUMOylation of tau in an alternatively spliced form, 0N4R-tau. Among SUMO E3 ligases, we found protein inhibitor of activated STAT (PIAS)xα and PIASxβ increase the levels of SUMOylated tau. The deSUMOylation enzymes sentrin-specific protease (SENP)1 and SENP2 reduced the levels of SUMO-conjugated tau. SUMO1 modification increased the level of phosphorylated tau, which was suppressed in the presence of SENP1. Furthermore, we examined the effect of tripartite motif (TRIM)11, which was recently identified as an E3 ligase for SUMO2 modification of tau. We found that TRIM11 increased the modification of both 2N4R- and 0N4R-tau by SUMO1, which was attenuated by mutation of the target lysine residue to arginine. These findings suggest that the expression and activity of SUMOylation regulatory proteins modulate the physical properties of tau proteins and may contribute to the onset and/or progression of tau-associated neurodegenerative disorders.

Keywords:  PIAS; SENP; SUMO; SUMOylation; TRIM11; Tau

DOI:  https://doi.org/10.1016/j.bbrep.2024.101800
bioRxiv. 2024 Sep 08. pii: 2024.09.05.611426. [Epub ahead of print]

Membrane-assisted Aβ40 aggregation pathways.

Fidha Nazreen Kunnath Muhammedkutty, Huan-Xiang Zhou.

Alzheimer's disease (AD) is caused by the assembly of amyloid-beta (Aβ) peptides into oligomers and fibrils. Endogenous Aβ aggregation may be assisted by cell membranes, which can accelerate the nucleation step enormously, but knowledge of membrane-assisted aggregation is still very limited. Here we used extensive MD simulations to structurally and energetically characterize key intermediates along the membrane-assisted aggregation pathways of Aβ40. Reinforcing experimental observations, the simulations reveal unique roles of GM1 ganglioside and cholesterol in stabilizing membrane-embedded β-sheets and of Y10 and K28 in the ordered release of a small oligomeric seed into solution. The same seed leads to either an open-shaped or R-shaped fibril, with significant stabilization provided by inter- or intra-subunit interfaces between a straight β- sheet (residues Q15-D23) and a bent β-sheet (residues A30-V36). This work presents the first comprehensive picture of membrane-assisted aggregation of Aβ40, with broad implications for developing AD therapies and rationalizing disease-specific polymorphisms of amyloidogenic proteins.

DOI: https://doi.org/10.1101/2024.09.05.611426
bioRxiv. 2024 Sep 02. pii: 2024.09.02.610853. [Epub ahead of print]

Methods for high throughput discovery of fluoroprobes that recognize tau fibril polymorphs.

Emma C Carroll, Hyunjun Yang, Julia G Jones, Abby Oehler, Annemarie F Charvat, Kelly M Montgomery, Anthony Yung, Zoe Millbern, Nelson R Vinueza, William F DeGrado, Daniel A Mordes, Carlo Condello, Jason E Gestwicki.

Aggregation of microtubule-associated protein tau (MAPT/tau) into conformationally distinct fibrils underpins neurodegenerative tauopathies. Fluorescent probes (fluoroprobes), such as thioflavin T (ThT), have been essential tools for studying tau aggregation; however, most of them do not discriminate between amyloid fibril conformations (polymorphs). This gap is due, in part, to a lack of high-throughput methods for screening large, diverse chemical collections. Here, we leverage advances in protein adaptive differential scanning fluorimetry (paDSF) to screen the Aurora collection of 300+ fluorescent dyes against multiple synthetic tau fibril polymorphs. This screen, coupled with orthogonal secondary assays, revealed pan-fibril binding chemotypes, as well as fluoroprobes selective for subsets of fibrils. One fluoroprobe recognized tau pathology in ex vivo brain slices from Alzheimer's disease patients. We propose that these scaffolds represent entry points for development of selective fibril ligands and, more broadly, that high throughput, fluorescence-based dye screening is a platform for their discovery.

DOI: https://doi.org/10.1101/2024.09.02.610853
Brain Behav Immun. 2024 Sep 15. pii: S0889-1591(24)00625-1. [Epub ahead of print]123 229-243

Astrocyte-neuron communication through the complement C3-C3aR pathway in Parkinson's disease.

Xiaosa Chi, Sijia Yin, Yadi Sun, Liang Kou, Wenkai Zou, Yiming Wang, Zongjie Jin, Tao Wang, Yun Xia.

  Neuroinflammation and autoimmunity are pivotal in the pathogenesis of neurodegenerative diseases. Complement activation and involvement of astrocyte-neuron C3/C3aR pathway have been observed, yet the mechanisms influencing α-synuclein (α-syn) pathology and neurodegeneration remain unclear. In this study, elevated levels of complement C3 were detected in the plasma of α-syn PFF-induced mice and the substantia nigra of A53T transgenic mice. Colocalization of complement C3 with astrocytes was also observed. Overexpression of complement C3 exacerbated motor dysfunction, dopaminergic neuron loss, and phosphorylated α-syn expression in mice injected with α-syn preformed fibrils (α-syn PFFs). Conversely, downregulation of complement C3 protected α-syn PFF-induced mice. Molecular investigations revealed that inhibition of Toll-like receptor 2 (TLR2) or NF-κB reduced complement C3 expression in primary astrocytes following α-syn PFF treatment. Astrocyte-neuron communication via the C3/C3aR pathway influenced α-syn PFF-induced neuronal apoptosis and α-syn pathology, potentially through modulation of GSK3β. These findings underscore the critical role of astrocyte-neuron communication via the C3/C3aR pathway in PD pathogenesis, highlighting its potential as a therapeutic target.

Keywords:  Astrocytes; Complement C3; Parkinson’s disease; α-synuclein

DOI:  https://doi.org/10.1016/j.bbi.2024.09.022
Front Pharmacol. 2024 ;15 1433030

HLH-30/TFEB modulates autophagy to improve proteostasis in Aβ transgenic Caenorhabditis elegans.

Hongru Lin, Chen Zhang, Yehui Gao, Yi Zhou, Botian Ma, Jinyun Jiang, Xue Long, Nuerziya Yimamu, Kaixin Zhong, Yingzi Li, Xianghuan Cui, Hongbing Wang.

  Alzheimer's disease (AD) is a complex neurodegenerative disease that affects elderly individuals, characterized by senile plaques formed by extracellular amyloid beta (Aβ). Autophagy dysfunction is a manifestation of protein homeostasis imbalance in patients with AD, but its relationship with Aβ remains unclear. Here, we showed that in Aβ transgenic Caenorhabditis elegans, Aβ activated the TOR pathway and reduced the nuclear entry of HLH-30, leading to autophagy dysfunction characterized by autophagosome accumulation. Then, utilizing RNA-seq, we investigated the regulatory mechanisms by which HLH-30 modulates autophagy in C. elegans. We found that HLH-30 elevated the transcript levels of v-ATPase and cathepsin, thus enhancing lysosomal activity. This led to an increase in autophagic flux, facilitating more pronounced degradation of Aβ. Moreover, HLH-30 reduced the level of ROS induction by Aβ and enhanced the antioxidant stress capacity of the worms through the gsto-1 gene. Additionally, we identified two HLH-30/TFEB activators, saikosaponin B2 and hypericin, that improved autophagic flux, thereby enhancing protein homeostasis in C. elegans. Overall, our findings suggested that HLH-30/TFEB plays a key role in modulating autophagy and can be considered a promising drug target for AD treatments.

Keywords:  Caenorhabditis elegans; HLH-30; HLH-30/TFEB activators; amyloid beta; autophagy

DOI:  https://doi.org/10.3389/fphar.2024.1433030
Front Mol Neurosci. 2024 ;17 1417961

Epigenetics in the formation of pathological aggregates in amyotrophic lateral sclerosis.

Veronica Noches, Danae Campos-Melo, Cristian A Droppelmann, Michael J Strong.

  The progressive degeneration of motor neurons in amyotrophic lateral sclerosis (ALS) is accompanied by the formation of a broad array of cytoplasmic and nuclear neuronal inclusions (protein aggregates) largely containing RNA-binding proteins such as TAR DNA-binding protein 43 (TDP-43) or fused in sarcoma/translocated in liposarcoma (FUS/TLS). This process is driven by a liquid-to-solid phase separation generally from proteins in membrane-less organelles giving rise to pathological biomolecular condensates. The formation of these protein aggregates suggests a fundamental alteration in the mRNA expression or the levels of the proteins involved. Considering the role of the epigenome in gene expression, alterations in DNA methylation, histone modifications, chromatin remodeling, non-coding RNAs, and RNA modifications become highly relevant to understanding how this pathological process takes effect. In this review, we explore the evidence that links epigenetic mechanisms with the formation of protein aggregates in ALS. We propose that a greater understanding of the role of the epigenome and how this inter-relates with the formation of pathological LLPS in ALS will provide an attractive therapeutic target.

Keywords:  ALS; DNA methylation; RNA modifications; chromatin remodeling enzymes; epigenetics; histone modifications; non-coding RNAs; protein aggregates

DOI:  https://doi.org/10.3389/fnmol.2024.1417961
Proc Natl Acad Sci U S A. 2024 Sep 24. 121(39): e2402162121

Spatiotemporal formation of a single liquid-like condensate and amyloid fibrils of α-synuclein by optical trapping at solution surface.

Keisuke Yuzu, Ching-Yang Lin, Po-Wei Yi, Chih-Hao Huang, Hiroshi Masuhara, Eri Chatani.

  Liquid-like protein condensates have recently attracted much attention due to their critical roles in biological phenomena. They typically show high fluidity and reversibility for exhibiting biological functions, while occasionally serving as sites for the formation of amyloid fibrils. To comprehend the properties of protein condensates that underlie biological function and pathogenesis, it is crucial to study them at the single-condensate level; however, this is currently challenging due to a lack of applicable methods. Here, we demonstrate that optical trapping is capable of inducing the formation of a single liquid-like condensate of α-synuclein in a spatiotemporally controlled manner. The irradiation of tightly focused near-infrared laser at an air/solution interface formed a condensate under conditions coexisting with polyethylene glycol. The fluorescent dye-labeled imaging showed that the optically induced condensate has a gradient of protein concentration from the center to the edge, suggesting that it is fabricated through optical pumping-up of the α-synuclein clusters and the expansion along the interface. Furthermore, Raman spectroscopy and thioflavin T fluorescence analysis revealed that continuous laser irradiation induces structural transition of protein molecules inside the condensate to β-sheet rich structure, ultimately leading to the condensate deformation and furthermore, the formation of amyloid fibrils. These observations indicate that optical trapping is a powerful technique for examining the microscopic mechanisms of condensate appearance and growth, and furthermore, subsequent aging leading to amyloid fibril formation.

Keywords:  amyloid fibrils; optical trapping; protein condensate; α-synuclein

DOI:  https://doi.org/10.1073/pnas.2402162121
bioRxiv. 2024 Sep 05. pii: 2024.09.03.611033. [Epub ahead of print]

Gigaxonin, mutated in Giant Axonal Neuropathy, interacts with TDP-43 and other RNA binding proteins.

Cassandra L Phillips, Maryam Faridounnia, Rachel A Battaglia, Baggio A Evangelista, Todd J Cohen, Puneet Opal, Thomas W Bouldin, Diane Armao, Natasha T Snider.

Giant Axonal Neuropathy (GAN) is a neurodegenerative disease caused by loss-of-function mutations in the KLHL16 gene, encoding the cytoskeleton regulator gigaxonin. In the absence of functional gigaxonin, intermediate filament (IF) proteins accumulate in neurons and other cell types due to impaired turnover and transport. GAN neurons exhibit distended, swollen axons and distal axonal degeneration, but the mechanisms behind this selective neuronal vulnerability are unknown. Our objective was to identify novel gigaxonin interactors pertinent to GAN neurons. Unbiased proteomics revealed a statistically significant predominance of RNA-binding proteins (RBPs) within the soluble gigaxonin interactome and among differentially-expressed proteins in iPSC-neuron progenitors from a patient with classic GAN. Among the identified RBPs was TAR DNA-binding protein 43 (TDP-43), which associated with the gigaxonin protein and its mRNA transcript. TDP-43 co-localized within large axonal neurofilament IFs aggregates in iPSC-motor neurons derived from a GAN patient with the 'axonal CMT-plus' disease phenotype. Our results implicate RBP dysfunction as a potential underappreciated contributor to GAN-related neurodegeneration.
Summary: This work reveals that the neurodegeneration-associated protein and cytoskeleton regulator gigaxonin and its mRNA associate with numerous RNA binding proteins. These findings shift understanding of normal gigaxonin function and provide insights into how disease-causing mutations in the gigaxonin-encoding gene ( KLHL16 ) may ignite a pathogenic cascade in neurons.

DOI: https://doi.org/10.1101/2024.09.03.611033
Eye (Lond). 2024 Sep 17.

Hallmarks of aging in age-related macular degeneration and age-related neurological disorders: novel insights into common mechanisms and clinical relevance.

Stela Vujosevic, Celeste Limoli, Igor Kozak.

Age-related macular degeneration (AMD) and age-related neurological diseases (ANDs), such as Alzheimer's and Parkinson's Diseases, are increasingly prevalent conditions that significantly contribute to global morbidity, disability, and mortality. The retina, as an accessible part of the central nervous system (CNS), provides a unique window to study brain aging and neurodegeneration. By examining the associations between AMD and ANDs, this review aims to highlight novel insights into fundamental mechanisms of aging and their role in neurodegenerative disease progression. This review integrates knowledge from the emerging field of aging research, which identifies common denominators of biological aging, specifically loss of proteostasis, impaired macroautophagy, mitochondrial dysfunction, and inflammation. Finally, we emphasize the clinical relevance of these pathways and the potential for cross-disease therapies that target common aging hallmarks. Identifying these shared pathways could open avenues to develop therapeutic strategies targeting mechanisms common to multiple degenerative diseases, potentially attenuating disease progression and promoting the healthspan.

DOI: https://doi.org/10.1038/s41433-024-03341-5
Cell Rep Med. 2024 Sep 17. pii: S2666-3791(24)00465-8. [Epub ahead of print]5(9): 101735

Integrating amyloid and tau imaging with proteomics and genomics in Alzheimer's disease.

Gabriele Vilkaite, Jacob Vogel, Niklas Mattsson-Carlgren.

Alzheimer's disease (AD) is the most common neurodegenerative disease and is characterized by the aggregation of β-amyloid (Aβ) and tau in the brain. Breakthroughs in disease-modifying treatments targeting Aβ bring new hope for the management of AD. But to effectively modify and someday even prevent AD, a better understanding is needed of the biological mechanisms that underlie and link Aβ and tau in AD. Developments of high-throughput omics, including genomics, proteomics, and transcriptomics, together with molecular imaging of Aβ and tau with positron emission tomography (PET), allow us to discover and understand the biological pathways that regulate the aggregation and spread of Aβ and tau in living humans. The field of integrated omics and PET studies of Aβ and tau in AD is growing rapidly. We here provide an update of this field, both in terms of biological insights and in terms of future clinical implications of integrated omics-molecular imaging studies.

DOI: https://doi.org/10.1016/j.xcrm.2024.101735