Objective To investigate the effect of ischemic preconditioning on chaperone hsp70 expression and protein aggregation in the CA1 neurons of rats, and to further explore its potential neuroprotective mechanism. Methods...Objective To investigate the effect of ischemic preconditioning on chaperone hsp70 expression and protein aggregation in the CA1 neurons of rats, and to further explore its potential neuroprotective mechanism. Methods Two-vesseloccluded transient global ischemia rat model was used. The rats were divided into sublethal 3-min ischemia group, lethal 10- min ischemia group and ischemic preconditioning group. Neuronal death in the CA1 region was observed by hematoxylineosin staining, and number of live neurons was assessed by cell counting under a light microscope. Immunochemistry and laser scanning confocal microscopy were used to observe the distribution of chaperone hsp70 in the CA1 neurons. Differential centrifuge was used to isolate cytosol, nucleus and protein aggregates fractions. Western blot was used to analyze the quantitative alterations of protein aggregates and inducible chaperone hsp70 in cellular fractions and in protein aggregates under different ischemic conditions. Results Histological examination showed that ischemic preconditioning significantly reduced delayed neuronal death in the hippocampus CA1 region （P 〈 0.01 vs 10-min ischemia group）. Sublethal ischemic preconditioning induced chaperone hsp70 expression in the CA1 neurons after 24 h reperfusion following 10-min ischemia. Induced-hsp70 combined with the abnormal proteins produced during the secondary lethal 10-min ischemia and inhibited the formation of cytotoxic protein aggregates（P〈0.01 vs 10-min ischemia group）.Conelusion Ischemic preconditioning induced chaperone hsp70 expression and inhibited protein aggregates formation in the CA1 neurons when suffered secondary lethal ischemia, which may protect neurons from death.展开更多
Pyropia haitanensis polysaccharide(LP)have been found for having many excellent functions such as anti-aging.Using Caenorhabditis elegans models,we evaluated the anti-aging activity of LP by observing the lifespan,rep...Pyropia haitanensis polysaccharide(LP)have been found for having many excellent functions such as anti-aging.Using Caenorhabditis elegans models,we evaluated the anti-aging activity of LP by observing the lifespan,reproduction,pharyngeal pumping,stress response,quantitative fluorescence of polyglutamic acid,and nuclear localization of DAF-16 of worms.The results reveal that LP could extend the adult lifespan of wild-type and polyQ nematodes,indicating a connection of its anti-aging benefit with the toxicity-suppressing effect.The number of polyglutamic acid aggregates in high concentration groups decreased by 24.39%(P<0.05)to the control.The high-dose group strongly induced DAF-16 nuclear translocation over intermediate and cytosolic localizations compared with the control(P<0.001).Therefore,we believe that LP could extend the lifespan and reduce the protein aggregation in C.elegans through nuclear DAF-16∷GFP expression.展开更多
Mutations in the fused in sarcoma/translocated in liposarcoma(FUS/TLS)gene have been associated with amyotrophic lateral sclerosis(ALS).FUS-positive neuropathology is reported in a range of neurodegenerative diseases,...Mutations in the fused in sarcoma/translocated in liposarcoma(FUS/TLS)gene have been associated with amyotrophic lateral sclerosis(ALS).FUS-positive neuropathology is reported in a range of neurodegenerative diseases,including ALS and fronto-temporal lobar degeneration with ubiquitin-positive pathology(FTLDU).To examine protein aggregation and cytotoxicity,we expressed human FUS protein in yeast.Expression of either wild type or ALS-associated R524S or P525L mutant FUS in yeast cells led to formation of aggregates and cytotoxicity,with the two ALS mutants showing increased cytotoxicity.Therefore,yeast cells expressing human FUS protein recapitulate key features of FUSpositive neurodegenerative diseases.Interestingly,a significant fraction of FUS expressing yeast cells stained by propidium iodide were without detectable protein aggregates,suggesting that membrane impairment and cellular damage caused by FUS expression may occur before protein aggregates become microscopically detectable and that aggregate formation might protect cells from FUS-mediated cytotoxicity.The N-terminus of FUS,containing the QGSY and G rich regions,is sufficient for the formation of aggregates but not cytotoxicity.The C-terminal domain,which contains a cluster of mutations,did not show aggregation or cytotoxicity.Similar to TDP-43 when expressed in yeast,FUS protein has the intrinsic property of forming aggregates in the absence of other human proteins.On the other hand,the aggregates formed by FUS are thioflavin T-positive and resistant to 0.5%sarkosyl,unlike TDP-43 when expressed in yeast cells.Furthermore,TDP-43 and FUS display distinct domain requirements in aggregate formation and cytotoxicity.展开更多
Freeze-thaw cycles impact the amount of aggregation observed in antibodies and Fc-fusion proteins. Various formulation strategies are used to mitigate the amount of aggregation that occurs upon putting a protein solut...Freeze-thaw cycles impact the amount of aggregation observed in antibodies and Fc-fusion proteins. Various formulation strategies are used to mitigate the amount of aggregation that occurs upon putting a protein solution through a freeze-thaw cycle. Additionally, low pH solutions cause native antibodies to unfold, which are prone to aggregate upon pH neutralization, There is great interest in the mechanism that causes therapeutic proteins to aggregate since aggregate species can cause unwanted immunogenicity in patients, Herein, an increase in aggregation is reported when the pH is adjusted from pH 3 up to a pH ranging from pH 4 to pH 7 during the thaw process of a frozen antibody solution, Raising the pH during the thaw process caused a significant increase in the percent aggregation observed. Two antibodies and one Fc-fusion protein were evaluated during the pH jump thaw process and similar effects were observed. The results provide a new tool to study the kinetics of therapeutic protein ag- gregation upon pH increase,展开更多
The generation of toxic non-native protein conformers has emerged as a unifying thread among disorders such as Alzheimer＇s disease, Parkinson＇s disease, and amyotrophic lateral sclerosis. Atomic-level detail regardi...The generation of toxic non-native protein conformers has emerged as a unifying thread among disorders such as Alzheimer＇s disease, Parkinson＇s disease, and amyotrophic lateral sclerosis. Atomic-level detail regarding dynamical changes that facilitate protein aggre- gation, as well as the structural features of large-scale ordered aggregates and soluble non-native oligomers, would contribute signifi- cantly to current understanding of these complex phenomena and offer potential strategies for inhibiting formation of cytotoxic species. However, experimental limitations often preclude the acquisition of high-resolution structural and mechanistic information for aggregating systems. Computational methods, particularly those combine both aU-atom and coarse-grained simulations to cover a wide range of time and length scales, have thus emerged as crucial tools for investigating protein aggregation. Here we review the current state of computational methodology for the study of protein self-assembly, with a focus on the application of these methods toward understanding of protein aggregates in human neurodegenerative disorders.展开更多
To investigate the relationship between protein aggregation and delayed neuronal death,we adopted rat models of 20 min ischemia.Brain ischemia was produced using the 2-vessel occlusion(2VO)model in rats Light microsco...To investigate the relationship between protein aggregation and delayed neuronal death,we adopted rat models of 20 min ischemia.Brain ischemia was produced using the 2-vessel occlusion(2VO)model in rats Light microscopy,transmission electronic microscopy and Western blot analysis were performed for morphological analysis of neurons,and protein detection.The results showed delayed neuronal death took place at 72 h after ischemia-reperfusion,protein aggregates formed at 4 h after reperfusion and reached the peak at 24 h after reper-fusion,and Western blot analysis was consistent with transmission electronic microscopy.We conclude that protein aggregation is one of the important factors leading to delayed neuronal death.展开更多
Protein-based therapeutics (PPTs) are drugs used to treat a variety of different conditions in the human body by alleviating enzymatic deficiencies, augmenting other proteins and drugs, modulating signal pathways, and...Protein-based therapeutics (PPTs) are drugs used to treat a variety of different conditions in the human body by alleviating enzymatic deficiencies, augmenting other proteins and drugs, modulating signal pathways, and more. However, many PPTs struggle from a short half-life due to degradation caused by irreversible protein aggregation in the bloodstream. Currently, the most researched strategies for improving the efficiency and longevity of PPTs are post-translational modifications (PTMs). The goal of our research was to determine which type of PTM increases longevity the most for each of three commonly-used therapeutic proteins by comparing the docking scores (DS) and binding free energies (BFE) from protein aggregation and reception simulations. DS and BFE values were used to create a quantitative index that outputs a relative number from −1 to 1 to show reduced performance, no change, or increased performance. Results showed that methylation was the most beneficial for insulin (p < 0.1) and human growth hormone (p < 0.0001), and both phosphorylation and methylation were somewhat optimal for erythropoietin (p < 0.1 and p < 0.0001, respectively). Acetylation consistently provided the worst benefits with the most negative indices, while methylation had the most positive indices throughout. However, PTM efficacy varied between PPTs, supporting previous studies regarding how each PTM can confer different benefits based on the unique structures of recipient proteins.展开更多
Mitochondria are crucial sites for protein quality control within cells.When mitochondrial stress is triggered by protein misfolding,it can accelerate abnormal protein aggregation,potentially inducing various diseases...Mitochondria are crucial sites for protein quality control within cells.When mitochondrial stress is triggered by protein misfolding,it can accelerate abnormal protein aggregation,potentially inducing various diseases.This study developed a cascade-responsive sensor,named AggHX,to monitor the microenvironment of protein aggregation induced by zinc(II)ions and the accompanying mitochondrial dysfunction.The AggHX consists of two key components:(1)A Zn^(2+) þrecognition group for triggering a fluorescent enhance response,and(2)a near-infrared BODIPY scaffold that detects viscosity changes in cell aggregation via HaloTag.This sensor's mechanism of action is elucidated through photochemical and biochemical characterizations.To further investigate the relationship between protein aggregation and mitochondrial homeostasis,we employ fluorescence lifetime imaging microscopy to assess viscosity changes in protein aggregates under intracellular Zn2þstress.This research provides insights into the dynamic behavior and spatial distribution of protein aggregates and mitochondria,contributing to a deeper understanding of their physiological roles in cellular processes and potential implications in disease pathology.展开更多
Neuroinflammation is a key process in the pathogenesis of various neurodegenerative diseases,such as multiple sclerosis(MS),Alzheimer's disease,and traumatic brain injury.Even for disorders historically unrelated ...Neuroinflammation is a key process in the pathogenesis of various neurodegenerative diseases,such as multiple sclerosis(MS),Alzheimer's disease,and traumatic brain injury.Even for disorders historically unrelated to neuroinflammation,such as Alzheimer's disease,it is now shown to precede pathological protein aggregations.展开更多
The modulation of protein aggregation is involved not only in biochemical engineering processes,but also in in vivo biological events such as Alzheimer's disease(AD)that features amyloid-βprotein(Aβ)deposits.Ins...The modulation of protein aggregation is involved not only in biochemical engineering processes,but also in in vivo biological events such as Alzheimer's disease(AD)that features amyloid-βprotein(Aβ)deposits.Inspired by the different pharmacological efficacy of enantiomers,taking heptapeptide LVFFARK(LK7)as an example,herein the chiral influence of peptide inhibitors on Aβfibrillogenesis and cytotoxicity was investigated by extensive biophysical and biological analyses.It was intriguing to find that although both LLK7 and D-LK7 could inhibit Aβaggregation in a concentration-dependent manner,it was the D-enan-tiomer that exhibited chirality preference and selectivity for modulation of Aβself-assembly.As com-pared with L-LK7 at the same conditions,D-LK7 showed significantly enhanced potency on suppressing cross-βsheet formation,fibrillar Aβaggregates deposition,Aβconformational transition,and Aβ-triggered neurotoxicity on cultured cells.For instance,L.LK7 and D-LK7 rescued cells by increasing cell via-bility from 60%to 62%and 84%at 100μmolL^(-1),respectively.The chiral discrimination of L-LK7 and D-LK7 was further validated by the different elimination efficiency on amyloid accumulation in AD model nematodes.It is considered that the higher binding affinity of D-LK7 to Aβmonomers than that of L LK7 resulted in the stronger inhibition effect.This work provided new insights into understanding chiral-ity in the interaction with Aβand the consequent inhibitory effect,and would contribute to the design of anti-amyloid agents.展开更多
Late Embryogenesis Abundant (LEA) proteins, a group of hydrophilic proteins, have been linked to survival in plants and animals in periods of stress, putatively through safeguarding enzymatic function and prevention o...Late Embryogenesis Abundant (LEA) proteins, a group of hydrophilic proteins, have been linked to survival in plants and animals in periods of stress, putatively through safeguarding enzymatic function and prevention of aggregation in times of dehydration/heat. Yet despite decades of effort, the molecular-level mechanisms defining this protective function remain unknown. In this paper, we summarize and review research discoveries of the classification of the LEA protein groups based on their amino acid sequence similarity and on the presence of distinctive conserved motifs. Moreover, we focus on high correlation between their accumulation and water deficit, reinforcing their functional relevance under abiotic stresses. We also discuss the biochemical properties of LEA proteins arising from their hydrophilic nature and by amino acid composition. Although significant similarities have not been found between the members of the different groups, a unifying and outstanding feature of most of them is their high hydrophilicity and high content of glycine. Therefore, we have highlighted the biotechnological applications of LEA genes, and the effects of over-expressing LEA genes from all LEA groups from different species of origin into different plant hosts. Apart from agronomical purposes, LEA proteins could be useful for other biotechnological applications in relation to their capacity to prevent aggregation of proteins.展开更多
Parkinson's disease,the second most prevalent neurodegenerative disorder worldwide,is characterized by a progressive loss of dopaminergic neurons in substantia nigra pars compacta,causing motor symptoms.This disor...Parkinson's disease,the second most prevalent neurodegenerative disorder worldwide,is characterized by a progressive loss of dopaminergic neurons in substantia nigra pars compacta,causing motor symptoms.This disorder's main hallmark is the formation of intraneuronal protein inclusions,named Lewy bodies and neurites.The major component of these arrangements is α-synuclein,an intrinsically disordered and soluble protein that,in pathological conditions,can form toxic and cell-to-cell transmissible amyloid structures.Preventing α-synuclein aggregation has attracted significant effort in the search for a disease-modifying therapy for Parkinson's disease.Small molecules like Synu Clean-D,epigallocatechin gallate,trodusquemine,or anle138 b exemplify this therapeutic potential.Here,we describe a subset of compounds containing a single aromatic ring,like dopamine,ZPDm,gallic acid,or entacapone,which act as molecular chaperones against α-synuclein aggregation.The simplicity of their structures contrasts with the complexity of the aggregation process,yet the block efficiently α-synuclein assembly into amyloid fibrils,in many cases,redirecting the reaction towards the formation of non-toxic off-pathway oligomers.Moreover,some of these compounds can disentangle mature α-synuclein amyloid fibrils.Their simple structures allow structure-activity relationship analysis to elucidate the role of different functional groups in the inhibition of α-synuclein aggregation and fibril dismantling,making them informative lead scaffolds for the rational development of efficient drugs.展开更多
Amyloids have traditionally been considered pathologic protein aggregates which contribute to neurodegeneration.New evidence however increasingly suggests that non-pathological amyloids are formed in animals during no...Amyloids have traditionally been considered pathologic protein aggregates which contribute to neurodegeneration.New evidence however increasingly suggests that non-pathological amyloids are formed in animals during normal development.Amyloid-like aggregate formation was originally thought to be a conserved feature of animal gametogenesis.This hypothesis was based on findings which suggest that regulated amyloid formations govern yeast meiosis by way of meiosis-specific RNA binding proteins.Additional support came from studies which demonstrate that DAZL,a mammalian gametogenesis-specific RNA binding protein,also forms SDS-resistant aggregates in vivo.Here,we report evidence of aggregated BOULE formations,another DAZ family protein,during sperm development.Data suggest that in mouse testis,BOULE forms SDS-resistant amyloid-like aggregates.BOULE aggregate formation correlates with dynamic developmental expression during spermatogenesis but disappeared in Boule knockout testis.We also mapped essential small region in vitro BOULE aggregations,immediately downstream DAZ repeats,and found that aggregations positively correlated with temperature.We also performed enhanced UV cross-linking immunoprecipitation on BOULE aggregates from mouse testes and found that aggregates bind with a large number of spermatogenesis-related mRNAs.These findings provide insight into the amyloidogenic properties of gametogenesis-specific RNA binding proteins as a conserved feature in mammalian reproduction.Further investigation is warranted to understand the functional significance of BOULE amyloid-like formation during mouse spermatogenesis.展开更多
Proteasome activity reduction is an important pathological phenomenon, resulting in proteins aggregation and neuronal death in the injured neurons induced by transient ischemia. Our previous report showed that the tra...Proteasome activity reduction is an important pathological phenomenon, resulting in proteins aggregation and neuronal death in the injured neurons induced by transient ischemia. Our previous report showed that the trap of proteasome in the protein aggregates was a reason to lead to the reduction of proteasome activity. However, the patterns of proteasome entered into protein aggregates are not clear. In this study, we used a global ischemia model, Hematoxylin-Eosin staining, differential centrifuge, proteasome activity assay, sucrose gradient density centrifuge, and Western blot analysis to investigate this problem. Our results show that there are two aggregation patterns of proteasome after transient ischemia and reperfusion. One is that 26S proteasome is trapped by protein aggregates as a whole unit, and the other is that 19S or 20S is trapped in the protein aggregates, respectively, after 26S disassociates.展开更多
Protein aggregates,mitochondrial import stress and neurodegenerative disorders:A salient hallmark of several neurodegenerative diseases,including Parkinson’s disease,is the abundance of protein aggregates(Goiran et a...Protein aggregates,mitochondrial import stress and neurodegenerative disorders:A salient hallmark of several neurodegenerative diseases,including Parkinson’s disease,is the abundance of protein aggregates(Goiran et al.,2022).This molecular event is believed to lead to activation of stress pathways ultimately resulting in cellular dysfunction(Eldeeb et al.,2022).Accordingly,many lines of research investigations focused on dampening the formation of protein aggregates or augmenting the clearance of protein aggregates as a potential therapeutic strategy to counteract the progression of neurodegenerative diseases,albeit with little success(Costa-Mattioli and Walter,2020).Cell stress cues such as the accumulation of protein aggregates lead to the activation of stress response pathways that aid cells in responding to the damage.Despite the notion that the transient activation of these pathways helps cells cope with stressors,persistent activation can induce unwanted apoptosis of cells and reduce overall tissue strength as well as lead to an accumulation of aggregation-prone proteins(Hetz and Papa,2018).Mutations in proteins involved in stress signaling termination can cause conditions like ataxia and early-onset dementia(Conroy et al.,2014).Therefore,it is crucial for stress response signaling to be turned off once conditions have improved.Nevertheless,the mechanisms by which cells silence these signals are still elusive.展开更多
Autophagy is well-known for delivering cargo materials to lysosomes for proteolytic digestion.Recently,autophagy has emerged as a key mechanism in unconventional protein secretion(UPS).This perspective introduces unco...Autophagy is well-known for delivering cargo materials to lysosomes for proteolytic digestion.Recently,autophagy has emerged as a key mechanism in unconventional protein secretion(UPS).This perspective introduces unconventional secretion pathways,focusing on secretory autophagy and its role in secreting protein aggregates associated with neurodegenerative disorders.We also explore additional neuronal functions of secretory autophagy beyond the release of protein aggregates.We propose autophagosomes as transport organelles that deliver cargo material directly from the endoplasmatic reticulum(ER)to the plasma membrane rather than solely to lysosomes.展开更多
Alzheimer＇s and Parkinson＇s diseases are the most common neurodegenerative diseases. They are characterized by protein aggregates and so can be considered as prion-like disease. The major components of these deposit...Alzheimer＇s and Parkinson＇s diseases are the most common neurodegenerative diseases. They are characterized by protein aggregates and so can be considered as prion-like disease. The major components of these deposits are amyloid peptide and tau for Alzheimer＇s disease, α-synuclein and synphilin-1 for Parkinson＇s disease. Drugs currently proposed to treat these pathologies do not prevent neurodegenerative processes and are mainly symptomatic therapies. Molecules inducing inhibition of aggregation or disaggregation of these proteins could have beneficial effects, especially if they have other beneficial effects for these diseases. Thus, several natural polyphenols, which have antioxidative, anti-inflammatory and neuroprotective properties, have been largely studied, for their effects on protein aggregates found in these diseases, notably in vitro. In this article, we propose to review the significant papers concerning the role of polyphenols on aggregation and disaggregation of amyloid peptide, tau, α-synuclein, synphilin-1, suggesting that these compounds could be useful in the treatments in Alzheimer＇s and Parkinson＇s diseases.展开更多
In his beautiful book,Consilience:The Unity of Knowledge,the eminent biologist Edward O Wilson,advocates the need for integration and reconciliation across the sciences.He defines consilience as“literally a‘jumping ...In his beautiful book,Consilience:The Unity of Knowledge,the eminent biologist Edward O Wilson,advocates the need for integration and reconciliation across the sciences.He defines consilience as“literally a‘jumping together’of knowledge with a linking of facts…to create a common groundwork of explanation”.It is the premise of this paper that as much as basic biomedical research is in need of data generation using the latest available techniques–unifying available knowledge is just as critical.This involves the necessity to resolve contradictory findings,reduce silos,and acknowledge complexity.We take the cornea and the lens as case studies of our premise.Specifically,in this perspective,we discuss the conflicting and fragmented information on protein aggregation,oxidative damage,and fibrosis.These are fields of study that are integrally tied to anterior segment research.Our goal is to highlight the vital need for Wilson’s consilience and unity of knowledge which in turn should lead to enhanced rigor and reproducibility,and most importantly,to greater understanding and not simply knowing.展开更多
Amyotrophic Lateral Sclerosis(ALS)is a complex neurodegenerative disorder characterized by progressive axonopathy,jointly leading to the dying back of the motor neuron,disrupting both nerve signaling and motor control...Amyotrophic Lateral Sclerosis(ALS)is a complex neurodegenerative disorder characterized by progressive axonopathy,jointly leading to the dying back of the motor neuron,disrupting both nerve signaling and motor control.In this review,we highlight the roles of axonopathy in ALS progression,driven by the interplay of multiple factors including defective trafficking machinery,protein aggregation,and mitochondrial dysfunction.Dysfunctional intracellular transport,caused by disruptions in microtubules,molecular motors,and adaptors,has been identified as a key contributor to disease progression.Aberrant protein aggregation involving TDP-43,FUS,SOD1,and dipeptide repeat proteins further amplifies neuronal toxicity.Mitochondrial defects lead to ATP depletion,oxidative stress,and Ca2+imbalance,which are regarded as key factors underlying the loss of neuromuscular junctions and axonopathy.Mitigating these defects through interventions including neurotrophic treatments offers therapeutic potential.Collaborative research efforts aim to unravel ALS complexities,opening avenues for holistic interventions that target diverse pathological mechanisms.展开更多
The epididymal lumen represents a unique extracellular environment because of the active sperm maturation process that takes place within its confines. Although much focus has been placed on the interaction of epididy...The epididymal lumen represents a unique extracellular environment because of the active sperm maturation process that takes place within its confines. Although much focus has been placed on the interaction of epididymal secretory proteins with spermatozoa in the lumen, very little is known regarding how the complex epididymal milieu as a whole is maintained, including mechanisms to prevent or control proteins that may not stay in their native folded state following secretion. Because some misfolded proteins can form cytotoxic aggregate structures known as amyloid, it is likely that control/surveillance mechanisms exist within the epididymis to protect against this process and allow sperm maturation to occur. To study protein aggregation and to identify extracellular quality control mechanisms in the epididymis, we used the cystatin family of cysteine protease inhibitors, including cystatin-related epididymal spermatogenic and cystatin C as molecular models because both proteins have inherent properties to aggregate and form amyloid. In this chapter, we present a brief summary of protein aggregation by the amyloid pathway based on what is known from other organ systems and describe quality control mechanisms that exist intracellularly to control protein misfolding and aggregation. We then present a summary of our studies of cystatinrelated epididymal spermatogenic （CRES） oligomerization within the epididymal lumen, including studies suggesting that transglutaminase cross-linking may be one mechanism of extracellular quality control within the epididymis. （Asian J Androl 2007 July; 9： 500-507）展开更多
基金the grants from the Department of Science and Technology of Jilin Province, China (No. 20070721)the Bureau of Science and Technology of Changchun, Jilin Province, China (No. 2007129).
文摘Objective To investigate the effect of ischemic preconditioning on chaperone hsp70 expression and protein aggregation in the CA1 neurons of rats, and to further explore its potential neuroprotective mechanism. Methods Two-vesseloccluded transient global ischemia rat model was used. The rats were divided into sublethal 3-min ischemia group, lethal 10- min ischemia group and ischemic preconditioning group. Neuronal death in the CA1 region was observed by hematoxylineosin staining, and number of live neurons was assessed by cell counting under a light microscope. Immunochemistry and laser scanning confocal microscopy were used to observe the distribution of chaperone hsp70 in the CA1 neurons. Differential centrifuge was used to isolate cytosol, nucleus and protein aggregates fractions. Western blot was used to analyze the quantitative alterations of protein aggregates and inducible chaperone hsp70 in cellular fractions and in protein aggregates under different ischemic conditions. Results Histological examination showed that ischemic preconditioning significantly reduced delayed neuronal death in the hippocampus CA1 region （P 〈 0.01 vs 10-min ischemia group）. Sublethal ischemic preconditioning induced chaperone hsp70 expression in the CA1 neurons after 24 h reperfusion following 10-min ischemia. Induced-hsp70 combined with the abnormal proteins produced during the secondary lethal 10-min ischemia and inhibited the formation of cytotoxic protein aggregates（P〈0.01 vs 10-min ischemia group）.Conelusion Ischemic preconditioning induced chaperone hsp70 expression and inhibited protein aggregates formation in the CA1 neurons when suffered secondary lethal ischemia, which may protect neurons from death.
基金Supported by the National Natural Science Foundation of China(Nos.31700307,41876165)the Science and Technology Project of Huzhou(No.2017ZD2017)the Key Research Program of Frontier Sciences,Chinese Academy of Sciences(No.QYZDB-SSW-DQC023)。
文摘Pyropia haitanensis polysaccharide(LP)have been found for having many excellent functions such as anti-aging.Using Caenorhabditis elegans models,we evaluated the anti-aging activity of LP by observing the lifespan,reproduction,pharyngeal pumping,stress response,quantitative fluorescence of polyglutamic acid,and nuclear localization of DAF-16 of worms.The results reveal that LP could extend the adult lifespan of wild-type and polyQ nematodes,indicating a connection of its anti-aging benefit with the toxicity-suppressing effect.The number of polyglutamic acid aggregates in high concentration groups decreased by 24.39%(P<0.05)to the control.The high-dose group strongly induced DAF-16 nuclear translocation over intermediate and cytosolic localizations compared with the control(P<0.001).Therefore,we believe that LP could extend the lifespan and reduce the protein aggregation in C.elegans through nuclear DAF-16∷GFP expression.
基金JYW is supported by NIH and James S.McDonnell Foundation.LL is supported by NIH(R01NS056086).
文摘Mutations in the fused in sarcoma/translocated in liposarcoma(FUS/TLS)gene have been associated with amyotrophic lateral sclerosis(ALS).FUS-positive neuropathology is reported in a range of neurodegenerative diseases,including ALS and fronto-temporal lobar degeneration with ubiquitin-positive pathology(FTLDU).To examine protein aggregation and cytotoxicity,we expressed human FUS protein in yeast.Expression of either wild type or ALS-associated R524S or P525L mutant FUS in yeast cells led to formation of aggregates and cytotoxicity,with the two ALS mutants showing increased cytotoxicity.Therefore,yeast cells expressing human FUS protein recapitulate key features of FUSpositive neurodegenerative diseases.Interestingly,a significant fraction of FUS expressing yeast cells stained by propidium iodide were without detectable protein aggregates,suggesting that membrane impairment and cellular damage caused by FUS expression may occur before protein aggregates become microscopically detectable and that aggregate formation might protect cells from FUS-mediated cytotoxicity.The N-terminus of FUS,containing the QGSY and G rich regions,is sufficient for the formation of aggregates but not cytotoxicity.The C-terminal domain,which contains a cluster of mutations,did not show aggregation or cytotoxicity.Similar to TDP-43 when expressed in yeast,FUS protein has the intrinsic property of forming aggregates in the absence of other human proteins.On the other hand,the aggregates formed by FUS are thioflavin T-positive and resistant to 0.5%sarkosyl,unlike TDP-43 when expressed in yeast cells.Furthermore,TDP-43 and FUS display distinct domain requirements in aggregate formation and cytotoxicity.
文摘Freeze-thaw cycles impact the amount of aggregation observed in antibodies and Fc-fusion proteins. Various formulation strategies are used to mitigate the amount of aggregation that occurs upon putting a protein solution through a freeze-thaw cycle. Additionally, low pH solutions cause native antibodies to unfold, which are prone to aggregate upon pH neutralization, There is great interest in the mechanism that causes therapeutic proteins to aggregate since aggregate species can cause unwanted immunogenicity in patients, Herein, an increase in aggregation is reported when the pH is adjusted from pH 3 up to a pH ranging from pH 4 to pH 7 during the thaw process of a frozen antibody solution, Raising the pH during the thaw process caused a significant increase in the percent aggregation observed. Two antibodies and one Fc-fusion protein were evaluated during the pH jump thaw process and similar effects were observed. The results provide a new tool to study the kinetics of therapeutic protein ag- gregation upon pH increase,
文摘The generation of toxic non-native protein conformers has emerged as a unifying thread among disorders such as Alzheimer＇s disease, Parkinson＇s disease, and amyotrophic lateral sclerosis. Atomic-level detail regarding dynamical changes that facilitate protein aggre- gation, as well as the structural features of large-scale ordered aggregates and soluble non-native oligomers, would contribute signifi- cantly to current understanding of these complex phenomena and offer potential strategies for inhibiting formation of cytotoxic species. However, experimental limitations often preclude the acquisition of high-resolution structural and mechanistic information for aggregating systems. Computational methods, particularly those combine both aU-atom and coarse-grained simulations to cover a wide range of time and length scales, have thus emerged as crucial tools for investigating protein aggregation. Here we review the current state of computational methodology for the study of protein self-assembly, with a focus on the application of these methods toward understanding of protein aggregates in human neurodegenerative disorders.
基金This project was supported by International Cooperation Fund of the Science and Technology Department of Jilin Province(No.20040707-1).
文摘To investigate the relationship between protein aggregation and delayed neuronal death,we adopted rat models of 20 min ischemia.Brain ischemia was produced using the 2-vessel occlusion(2VO)model in rats Light microscopy,transmission electronic microscopy and Western blot analysis were performed for morphological analysis of neurons,and protein detection.The results showed delayed neuronal death took place at 72 h after ischemia-reperfusion,protein aggregates formed at 4 h after reperfusion and reached the peak at 24 h after reper-fusion,and Western blot analysis was consistent with transmission electronic microscopy.We conclude that protein aggregation is one of the important factors leading to delayed neuronal death.
文摘Protein-based therapeutics (PPTs) are drugs used to treat a variety of different conditions in the human body by alleviating enzymatic deficiencies, augmenting other proteins and drugs, modulating signal pathways, and more. However, many PPTs struggle from a short half-life due to degradation caused by irreversible protein aggregation in the bloodstream. Currently, the most researched strategies for improving the efficiency and longevity of PPTs are post-translational modifications (PTMs). The goal of our research was to determine which type of PTM increases longevity the most for each of three commonly-used therapeutic proteins by comparing the docking scores (DS) and binding free energies (BFE) from protein aggregation and reception simulations. DS and BFE values were used to create a quantitative index that outputs a relative number from −1 to 1 to show reduced performance, no change, or increased performance. Results showed that methylation was the most beneficial for insulin (p < 0.1) and human growth hormone (p < 0.0001), and both phosphorylation and methylation were somewhat optimal for erythropoietin (p < 0.1 and p < 0.0001, respectively). Acetylation consistently provided the worst benefits with the most negative indices, while methylation had the most positive indices throughout. However, PTM efficacy varied between PPTs, supporting previous studies regarding how each PTM can confer different benefits based on the unique structures of recipient proteins.
基金support from the Qinglan Project of Jiangsu Province of China,National Natural Science Foundation of China(Grant No.22007048)the Natural Science Foundation of Jiangsu Basic Research Program(BK20221324).
文摘Mitochondria are crucial sites for protein quality control within cells.When mitochondrial stress is triggered by protein misfolding,it can accelerate abnormal protein aggregation,potentially inducing various diseases.This study developed a cascade-responsive sensor,named AggHX,to monitor the microenvironment of protein aggregation induced by zinc(II)ions and the accompanying mitochondrial dysfunction.The AggHX consists of two key components:(1)A Zn^(2+) þrecognition group for triggering a fluorescent enhance response,and(2)a near-infrared BODIPY scaffold that detects viscosity changes in cell aggregation via HaloTag.This sensor's mechanism of action is elucidated through photochemical and biochemical characterizations.To further investigate the relationship between protein aggregation and mitochondrial homeostasis,we employ fluorescence lifetime imaging microscopy to assess viscosity changes in protein aggregates under intracellular Zn2þstress.This research provides insights into the dynamic behavior and spatial distribution of protein aggregates and mitochondria,contributing to a deeper understanding of their physiological roles in cellular processes and potential implications in disease pathology.
基金supported by FWO(Fonds voor Wetenschappelijk Onderzoek),grant number G07562NFWO(to BB)。
文摘Neuroinflammation is a key process in the pathogenesis of various neurodegenerative diseases,such as multiple sclerosis(MS),Alzheimer's disease,and traumatic brain injury.Even for disorders historically unrelated to neuroinflammation,such as Alzheimer's disease,it is now shown to precede pathological protein aggregations.
基金supported by the National Natural Science Foundation of China(Nos.21621004 and 21978207)the Natural Science Foundation of Tianjin from Tianjin Municipal Science and Technology Commission(No.19JCZDJC36800).
文摘The modulation of protein aggregation is involved not only in biochemical engineering processes,but also in in vivo biological events such as Alzheimer's disease(AD)that features amyloid-βprotein(Aβ)deposits.Inspired by the different pharmacological efficacy of enantiomers,taking heptapeptide LVFFARK(LK7)as an example,herein the chiral influence of peptide inhibitors on Aβfibrillogenesis and cytotoxicity was investigated by extensive biophysical and biological analyses.It was intriguing to find that although both LLK7 and D-LK7 could inhibit Aβaggregation in a concentration-dependent manner,it was the D-enan-tiomer that exhibited chirality preference and selectivity for modulation of Aβself-assembly.As com-pared with L-LK7 at the same conditions,D-LK7 showed significantly enhanced potency on suppressing cross-βsheet formation,fibrillar Aβaggregates deposition,Aβconformational transition,and Aβ-triggered neurotoxicity on cultured cells.For instance,L.LK7 and D-LK7 rescued cells by increasing cell via-bility from 60%to 62%and 84%at 100μmolL^(-1),respectively.The chiral discrimination of L-LK7 and D-LK7 was further validated by the different elimination efficiency on amyloid accumulation in AD model nematodes.It is considered that the higher binding affinity of D-LK7 to Aβmonomers than that of L LK7 resulted in the stronger inhibition effect.This work provided new insights into understanding chiral-ity in the interaction with Aβand the consequent inhibitory effect,and would contribute to the design of anti-amyloid agents.
基金supported jointly by grants from the Ministry of Higher Education and Scientific Research,Tunisia and the Agence Espagnole de cooperation Internationale(AECI)Officina Tecnica de Cooperacion,Spain
文摘Late Embryogenesis Abundant (LEA) proteins, a group of hydrophilic proteins, have been linked to survival in plants and animals in periods of stress, putatively through safeguarding enzymatic function and prevention of aggregation in times of dehydration/heat. Yet despite decades of effort, the molecular-level mechanisms defining this protective function remain unknown. In this paper, we summarize and review research discoveries of the classification of the LEA protein groups based on their amino acid sequence similarity and on the presence of distinctive conserved motifs. Moreover, we focus on high correlation between their accumulation and water deficit, reinforcing their functional relevance under abiotic stresses. We also discuss the biochemical properties of LEA proteins arising from their hydrophilic nature and by amino acid composition. Although significant similarities have not been found between the members of the different groups, a unifying and outstanding feature of most of them is their high hydrophilicity and high content of glycine. Therefore, we have highlighted the biotechnological applications of LEA genes, and the effects of over-expressing LEA genes from all LEA groups from different species of origin into different plant hosts. Apart from agronomical purposes, LEA proteins could be useful for other biotechnological applications in relation to their capacity to prevent aggregation of proteins.
文摘Parkinson's disease,the second most prevalent neurodegenerative disorder worldwide,is characterized by a progressive loss of dopaminergic neurons in substantia nigra pars compacta,causing motor symptoms.This disorder's main hallmark is the formation of intraneuronal protein inclusions,named Lewy bodies and neurites.The major component of these arrangements is α-synuclein,an intrinsically disordered and soluble protein that,in pathological conditions,can form toxic and cell-to-cell transmissible amyloid structures.Preventing α-synuclein aggregation has attracted significant effort in the search for a disease-modifying therapy for Parkinson's disease.Small molecules like Synu Clean-D,epigallocatechin gallate,trodusquemine,or anle138 b exemplify this therapeutic potential.Here,we describe a subset of compounds containing a single aromatic ring,like dopamine,ZPDm,gallic acid,or entacapone,which act as molecular chaperones against α-synuclein aggregation.The simplicity of their structures contrasts with the complexity of the aggregation process,yet the block efficiently α-synuclein assembly into amyloid fibrils,in many cases,redirecting the reaction towards the formation of non-toxic off-pathway oligomers.Moreover,some of these compounds can disentangle mature α-synuclein amyloid fibrils.Their simple structures allow structure-activity relationship analysis to elucidate the role of different functional groups in the inhibition of α-synuclein aggregation and fibril dismantling,making them informative lead scaffolds for the rational development of efficient drugs.
基金supported by the National Natural Science Foundation of China(No.3197060323)SKLRM grant(SKLRM-2019B2)the Jiangsu ShuangChuang Talent Program,as well as the Jiangsu graduate student innovation fellowship to Y.S.
文摘Amyloids have traditionally been considered pathologic protein aggregates which contribute to neurodegeneration.New evidence however increasingly suggests that non-pathological amyloids are formed in animals during normal development.Amyloid-like aggregate formation was originally thought to be a conserved feature of animal gametogenesis.This hypothesis was based on findings which suggest that regulated amyloid formations govern yeast meiosis by way of meiosis-specific RNA binding proteins.Additional support came from studies which demonstrate that DAZL,a mammalian gametogenesis-specific RNA binding protein,also forms SDS-resistant aggregates in vivo.Here,we report evidence of aggregated BOULE formations,another DAZ family protein,during sperm development.Data suggest that in mouse testis,BOULE forms SDS-resistant amyloid-like aggregates.BOULE aggregate formation correlates with dynamic developmental expression during spermatogenesis but disappeared in Boule knockout testis.We also mapped essential small region in vitro BOULE aggregations,immediately downstream DAZ repeats,and found that aggregations positively correlated with temperature.We also performed enhanced UV cross-linking immunoprecipitation on BOULE aggregates from mouse testes and found that aggregates bind with a large number of spermatogenesis-related mRNAs.These findings provide insight into the amyloidogenic properties of gametogenesis-specific RNA binding proteins as a conserved feature in mammalian reproduction.Further investigation is warranted to understand the functional significance of BOULE amyloid-like formation during mouse spermatogenesis.
基金Supported by the National Natural Science Foundation of China(Nos.30973110 81072071)+2 种基金the International Cooperation Grant(No.20070721)the Outstanding Youth Grant from the Science and Technology Department of Jilin ProvinceChina (No.20080139)
文摘Proteasome activity reduction is an important pathological phenomenon, resulting in proteins aggregation and neuronal death in the injured neurons induced by transient ischemia. Our previous report showed that the trap of proteasome in the protein aggregates was a reason to lead to the reduction of proteasome activity. However, the patterns of proteasome entered into protein aggregates are not clear. In this study, we used a global ischemia model, Hematoxylin-Eosin staining, differential centrifuge, proteasome activity assay, sucrose gradient density centrifuge, and Western blot analysis to investigate this problem. Our results show that there are two aggregation patterns of proteasome after transient ischemia and reperfusion. One is that 26S proteasome is trapped by protein aggregates as a whole unit, and the other is that 19S or 20S is trapped in the protein aggregates, respectively, after 26S disassociates.
文摘Protein aggregates,mitochondrial import stress and neurodegenerative disorders:A salient hallmark of several neurodegenerative diseases,including Parkinson’s disease,is the abundance of protein aggregates(Goiran et al.,2022).This molecular event is believed to lead to activation of stress pathways ultimately resulting in cellular dysfunction(Eldeeb et al.,2022).Accordingly,many lines of research investigations focused on dampening the formation of protein aggregates or augmenting the clearance of protein aggregates as a potential therapeutic strategy to counteract the progression of neurodegenerative diseases,albeit with little success(Costa-Mattioli and Walter,2020).Cell stress cues such as the accumulation of protein aggregates lead to the activation of stress response pathways that aid cells in responding to the damage.Despite the notion that the transient activation of these pathways helps cells cope with stressors,persistent activation can induce unwanted apoptosis of cells and reduce overall tissue strength as well as lead to an accumulation of aggregation-prone proteins(Hetz and Papa,2018).Mutations in proteins involved in stress signaling termination can cause conditions like ataxia and early-onset dementia(Conroy et al.,2014).Therefore,it is crucial for stress response signaling to be turned off once conditions have improved.Nevertheless,the mechanisms by which cells silence these signals are still elusive.
基金supported by the Deutsche Forschungsgemeinschaft(DFG,German Research Foundation)grant LU 2347/3-1(to PL).
文摘Autophagy is well-known for delivering cargo materials to lysosomes for proteolytic digestion.Recently,autophagy has emerged as a key mechanism in unconventional protein secretion(UPS).This perspective introduces unconventional secretion pathways,focusing on secretory autophagy and its role in secreting protein aggregates associated with neurodegenerative disorders.We also explore additional neuronal functions of secretory autophagy beyond the release of protein aggregates.We propose autophagosomes as transport organelles that deliver cargo material directly from the endoplasmatic reticulum(ER)to the plasma membrane rather than solely to lysosomes.
文摘Alzheimer＇s and Parkinson＇s diseases are the most common neurodegenerative diseases. They are characterized by protein aggregates and so can be considered as prion-like disease. The major components of these deposits are amyloid peptide and tau for Alzheimer＇s disease, α-synuclein and synphilin-1 for Parkinson＇s disease. Drugs currently proposed to treat these pathologies do not prevent neurodegenerative processes and are mainly symptomatic therapies. Molecules inducing inhibition of aggregation or disaggregation of these proteins could have beneficial effects, especially if they have other beneficial effects for these diseases. Thus, several natural polyphenols, which have antioxidative, anti-inflammatory and neuroprotective properties, have been largely studied, for their effects on protein aggregates found in these diseases, notably in vitro. In this article, we propose to review the significant papers concerning the role of polyphenols on aggregation and disaggregation of amyloid peptide, tau, α-synuclein, synphilin-1, suggesting that these compounds could be useful in the treatments in Alzheimer＇s and Parkinson＇s diseases.
文摘In his beautiful book,Consilience:The Unity of Knowledge,the eminent biologist Edward O Wilson,advocates the need for integration and reconciliation across the sciences.He defines consilience as“literally a‘jumping together’of knowledge with a linking of facts…to create a common groundwork of explanation”.It is the premise of this paper that as much as basic biomedical research is in need of data generation using the latest available techniques–unifying available knowledge is just as critical.This involves the necessity to resolve contradictory findings,reduce silos,and acknowledge complexity.We take the cornea and the lens as case studies of our premise.Specifically,in this perspective,we discuss the conflicting and fragmented information on protein aggregation,oxidative damage,and fibrosis.These are fields of study that are integrally tied to anterior segment research.Our goal is to highlight the vital need for Wilson’s consilience and unity of knowledge which in turn should lead to enhanced rigor and reproducibility,and most importantly,to greater understanding and not simply knowing.
基金supported by the National Natural Science Foundation of China(32271001 and 31871036).
文摘Amyotrophic Lateral Sclerosis(ALS)is a complex neurodegenerative disorder characterized by progressive axonopathy,jointly leading to the dying back of the motor neuron,disrupting both nerve signaling and motor control.In this review,we highlight the roles of axonopathy in ALS progression,driven by the interplay of multiple factors including defective trafficking machinery,protein aggregation,and mitochondrial dysfunction.Dysfunctional intracellular transport,caused by disruptions in microtubules,molecular motors,and adaptors,has been identified as a key contributor to disease progression.Aberrant protein aggregation involving TDP-43,FUS,SOD1,and dipeptide repeat proteins further amplifies neuronal toxicity.Mitochondrial defects lead to ATP depletion,oxidative stress,and Ca2+imbalance,which are regarded as key factors underlying the loss of neuromuscular junctions and axonopathy.Mitigating these defects through interventions including neurotrophic treatments offers therapeutic potential.Collaborative research efforts aim to unravel ALS complexities,opening avenues for holistic interventions that target diverse pathological mechanisms.
文摘The epididymal lumen represents a unique extracellular environment because of the active sperm maturation process that takes place within its confines. Although much focus has been placed on the interaction of epididymal secretory proteins with spermatozoa in the lumen, very little is known regarding how the complex epididymal milieu as a whole is maintained, including mechanisms to prevent or control proteins that may not stay in their native folded state following secretion. Because some misfolded proteins can form cytotoxic aggregate structures known as amyloid, it is likely that control/surveillance mechanisms exist within the epididymis to protect against this process and allow sperm maturation to occur. To study protein aggregation and to identify extracellular quality control mechanisms in the epididymis, we used the cystatin family of cysteine protease inhibitors, including cystatin-related epididymal spermatogenic and cystatin C as molecular models because both proteins have inherent properties to aggregate and form amyloid. In this chapter, we present a brief summary of protein aggregation by the amyloid pathway based on what is known from other organ systems and describe quality control mechanisms that exist intracellularly to control protein misfolding and aggregation. We then present a summary of our studies of cystatinrelated epididymal spermatogenic （CRES） oligomerization within the epididymal lumen, including studies suggesting that transglutaminase cross-linking may be one mechanism of extracellular quality control within the epididymis. （Asian J Androl 2007 July; 9： 500-507）
