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1.
Cell Tissue Res ; 395(1): 21-38, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38015266

RESUMO

Nothobranchius furzeri is emerging as an exciting vertebrate organism in the field of biomedicine, developmental biology and ecotoxicology research. Its short generation time, compressed lifespan and accelerated ageing make it a versatile model for longitudinal studies with high traceability. Although in recent years the use of this model has increased enormously, there is still little information on the anatomy, morphology and histology of its main organs. In this paper, we present a description of the digestive system of N. furzeri, with emphasis on the intestine. We note that the general architecture of the intestinal tissue is shared with other vertebrates, and includes a folding mucosa, an outer muscle layer and a myenteric plexus. By immunohistochemical analysis, we reveal that the mucosa harbours the same type of epithelial cells observed in mammals, including enterocytes, goblet cells and enteroendocrine cells, and that the myenteric neurons express neurotransmitters common to other species, such as serotonin, substance P and tyrosine hydroxylase. In addition, we detect the presence of a proliferative compartment at the base of the intestinal folds. The description of the normal intestinal morphology provided here constitutes a baseline information to contrast with tissue alterations in future lines of research assessing pathologies, ageing-related diseases or damage caused by toxic agents.


Assuntos
Envelhecimento , Intestinos , Animais , Mamíferos
3.
Front Pharmacol, v. 14, 1176805, mar. 2023
Artigo em Inglês | Sec. Est. Saúde SP, SESSP-IBPROD, Sec. Est. Saúde SP | ID: bud-4850
4.
EMBO J ; 41(22): e111952, 2022 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-36314651

RESUMO

Aging is a major risk factor to develop neurodegenerative diseases and is associated with decreased buffering capacity of the proteostasis network. We investigated the significance of the unfolded protein response (UPR), a major signaling pathway activated to cope with endoplasmic reticulum (ER) stress, in the functional deterioration of the mammalian brain during aging. We report that genetic disruption of the ER stress sensor IRE1 accelerated age-related cognitive decline. In mouse models, overexpressing an active form of the UPR transcription factor XBP1 restored synaptic and cognitive function, in addition to reducing cell senescence. Proteomic profiling of hippocampal tissue showed that XBP1 expression significantly restore changes associated with aging, including factors involved in synaptic function and pathways linked to neurodegenerative diseases. The genes modified by XBP1 in the aged hippocampus where also altered. Collectively, our results demonstrate that strategies to manipulate the UPR in mammals may help sustain healthy brain aging.


Assuntos
Envelhecimento , Encéfalo , Proteínas Serina-Treonina Quinases , Resposta a Proteínas não Dobradas , Proteína 1 de Ligação a X-Box , Animais , Camundongos , Envelhecimento/genética , Encéfalo/metabolismo , Estresse do Retículo Endoplasmático/genética , Proteínas Serina-Treonina Quinases/genética , Proteômica , Transdução de Sinais/fisiologia , Proteína 1 de Ligação a X-Box/genética , Proteína 1 de Ligação a X-Box/metabolismo
5.
EMBO J ; 41(2): e105531, 2022 12 17.
Artigo em Inglês | MEDLINE | ID: mdl-34904718

RESUMO

Recessive gene mutations underlie many developmental disorders and often lead to disabling neurological problems. Here, we report identification of a homozygous c.170G>A (p.Cys57Tyr or C57Y) mutation in the gene coding for protein disulfide isomerase A3 (PDIA3, also known as ERp57), an enzyme that catalyzes formation of disulfide bonds in the endoplasmic reticulum, to be associated with syndromic intellectual disability. Experiments in zebrafish embryos show that PDIA3C57Y expression is pathogenic and causes developmental defects such as axonal disorganization as well as skeletal abnormalities. Expression of PDIA3C57Y in the mouse hippocampus results in impaired synaptic plasticity and memory consolidation. Proteomic and functional analyses reveal that PDIA3C57Y expression leads to dysregulation of cell adhesion and actin cytoskeleton dynamics, associated with altered integrin biogenesis and reduced neuritogenesis. Biochemical studies show that PDIA3C57Y has decreased catalytic activity and forms disulfide-crosslinked aggregates that abnormally interact with chaperones in the endoplasmic reticulum. Thus, rare disease gene variant can provide insight into how perturbations of neuronal proteostasis can affect the function of the nervous system.


Assuntos
Deficiências do Desenvolvimento/genética , Retículo Endoplasmático/metabolismo , Isomerases de Dissulfetos de Proteínas/genética , Proteostase , Adolescente , Adulto , Animais , Axônios/metabolismo , Axônios/patologia , Adesão Celular , Células Cultivadas , Criança , Citoesqueleto/metabolismo , Deficiências do Desenvolvimento/metabolismo , Deficiências do Desenvolvimento/patologia , Feminino , Hipocampo/metabolismo , Hipocampo/patologia , Humanos , Integrinas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mutação de Sentido Incorreto , Crescimento Neuronal , Plasticidade Neuronal , Linhagem , Isomerases de Dissulfetos de Proteínas/metabolismo , Peixe-Zebra
6.
Prog Mol Subcell Biol ; 59: 239-278, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34050870

RESUMO

Endoplasmic reticulum (ER) stress is a prominent cellular alteration of diseases impacting the nervous system that are associated to the accumulation of misfolded and aggregated protein species during aging. The unfolded protein response (UPR) is the main pathway mediating adaptation to ER stress, but it can also trigger deleterious cascades of inflammation and cell death leading to cell dysfunction and neurodegeneration. Genetic and pharmacological studies in experimental models shed light into molecular pathways possibly contributing to ER stress and the UPR activation in human neuropathies. Most of experimental models are, however, based on the overexpression of mutant proteins causing familial forms of these diseases or the administration of neurotoxins that induce pathology in young animals. Whether the mechanisms uncovered in these models are relevant for the etiology of the vast majority of age-related sporadic forms of neurodegenerative diseases is an open question. Here, we provide a systematic analysis of the current evidence linking ER stress to human pathology and the main mechanisms elucidated in experimental models. Furthermore, we highlight the recent association of metabolic syndrome to increased risk to undergo neurodegeneration, where ER stress arises as a common denominator in the pathogenic crosstalk between peripheral organs and the nervous system.


Assuntos
Retículo Endoplasmático , Proteostase , Animais , Retículo Endoplasmático/genética , Retículo Endoplasmático/metabolismo , Estresse do Retículo Endoplasmático/genética , Humanos , Doenças Neurodegenerativas/genética , Proteostase/genética , Resposta a Proteínas não Dobradas/genética
7.
Acta Neuropathol Commun ; 9(1): 21, 2021 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-33541434

RESUMO

Amyotrophic lateral sclerosis (ALS) is a progressive fatal neurodegenerative disease that affects motoneurons. Mutations in superoxide dismutase 1 (SOD1) have been described as a causative genetic factor for ALS. Mice overexpressing ALS-linked mutant SOD1 develop ALS symptoms accompanied by histopathological alterations and protein aggregation. The protein disulfide isomerase family member ERp57 is one of the main up-regulated proteins in tissue of ALS patients and mutant SOD1 mice, whereas point mutations in ERp57 were described as possible risk factors to develop the disease. ERp57 catalyzes disulfide bond formation and isomerization in the endoplasmic reticulum (ER), constituting a central component of protein quality control mechanisms. However, the actual contribution of ERp57 to ALS pathogenesis remained to be defined. Here, we studied the consequences of overexpressing ERp57 in experimental ALS using mutant SOD1 mice. Double transgenic SOD1G93A/ERp57WT animals presented delayed deterioration of electrophysiological activity and maintained muscle innervation compared to single transgenic SOD1G93A littermates at early-symptomatic stage, along with improved motor performance without affecting survival. The overexpression of ERp57 reduced mutant SOD1 aggregation, but only at disease end-stage, dissociating its role as an anti-aggregation factor from the protection of neuromuscular junctions. Instead, proteomic analysis revealed that the neuroprotective effects of ERp57 overexpression correlated with increased levels of synaptic and actin cytoskeleton proteins in the spinal cord. Taken together, our results suggest that ERp57 operates as a disease modifier at early stages by maintaining motoneuron connectivity.


Assuntos
Esclerose Lateral Amiotrófica/enzimologia , Esclerose Lateral Amiotrófica/prevenção & controle , Isomerases de Dissulfetos de Proteínas/genética , Isomerases de Dissulfetos de Proteínas/metabolismo , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/patologia , Animais , Modelos Animais de Doenças , Eletromiografia , Camundongos , Camundongos Transgênicos , Neurônios Motores/metabolismo , Denervação Muscular , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Junção Neuromuscular/metabolismo , Proteômica , Medula Espinal/patologia , Superóxido Dismutase-1/genética , Superóxido Dismutase-1/metabolismo
9.
Neurobiol Dis ; 130: 104497, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31176720

RESUMO

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by motor neuron death. A 20% of familial ALS cases are associated with mutations in the gene coding for superoxide dismutase 1 (SOD1). The accumulation of abnormal aggregates of different proteins is a common feature in motor neurons of patients and transgenic ALS mice models, which are thought to contribute to disease pathogenesis. Developmental morphogens, such as the Wnt family, regulate numerous features of neuronal physiology in the adult brain and have been implicated in neurodegeneration. ß-catenin is a central mediator of both, Wnt signaling activity and cell-cell interactions. We previously reported that the expression of mutant SOD1 in the NSC34 motor neuron cell line decreases basal Wnt pathway activity, which correlates with cytosolic ß-catenin accumulation and impaired neuronal differentiation. In this work, we aimed a deeper characterization of ß-catenin distribution in models of ALS motor neurons. We observed extensive accumulation of ß-catenin supramolecular structures in motor neuron somas of pre-symptomatic mutant SOD1 mice. In cell-cell appositional zones of NSC34 cells expressing mutant SOD1, ß-catenin displays a reduced co-distribution with E-cadherin accompanied by an increased association with the gap junction protein Connexin-43; these findings correlate with impaired intercellular adhesion and exacerbated cell coupling. Remarkably, pharmacological inhibition of the glycogen synthase kinase-3ß (GSK3ß) in both NSC34 cell lines reverted both, ß-catenin aggregation and the adverse effects of mutant SOD1 expression on neuronal differentiation. Our findings suggest that early defects in ß-catenin distribution could be an underlying factor affecting the onset of neurodegeneration in familial ALS.


Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Neurônios Motores/metabolismo , beta Catenina/metabolismo , Animais , Diferenciação Celular/fisiologia , Células Cultivadas , Modelos Animais de Doenças , Humanos , Camundongos
11.
Sci Rep ; 7(1): 14266, 2017 10 27.
Artigo em Inglês | MEDLINE | ID: mdl-29079747

RESUMO

Tar DNA binding protein 43 (TDP-43) is the principal component of ubiquitinated protein inclusions present in nervous tissue of most cases of both amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Previous studies described a TDP-43A315T transgenic mouse model that develops progressive motor dysfunction in the absence of protein aggregation or significant motoneuron loss, questioning its validity to study ALS. Here we have further characterized the course of the disease in TDP-43A315T mice using a battery of tests and biochemical approaches. We confirmed that TDP-43 mutant mice develop impaired motor performance, accompanied by progressive body weight loss. Significant differences were observed in life span between genders, where females survived longer than males. Histopathological analysis of the spinal cord demonstrated a significant motoneurons loss, accompanied by axonal degeneration, astrogliosis and microglial activation. Importantly, histopathological alterations observed in TDP-43 mutant mice were similar to some characteristic changes observed in mutant SOD1 mice. Unexpectedly, we identified the presence of different species of disulfide-dependent TDP-43 aggregates in cortex and spinal cord tissue. Overall, this study indicates that TDP-43A315T transgenic mice develop key features resembling key aspects of ALS, highlighting its relevance to study disease pathogenesis.


Assuntos
Esclerose Lateral Amiotrófica/patologia , Proteínas de Ligação a DNA/química , Dissulfetos/química , Demência Frontotemporal/patologia , Neurônios Motores/patologia , Multimerização Proteica , Medula Espinal/patologia , Esclerose Lateral Amiotrófica/metabolismo , Animais , Contagem de Células , Proteínas de Ligação a DNA/genética , Modelos Animais de Doenças , Feminino , Demência Frontotemporal/metabolismo , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Córtex Pré-Frontal/metabolismo , Agregados Proteicos , Estrutura Quaternária de Proteína , Medula Espinal/metabolismo
12.
Hum Mol Genet ; 26(R2): R91-R104, 2017 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-28977445

RESUMO

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease affecting motoneurons in the brain and spinal cord leading to paralysis and death. Although the etiology of ALS remains poorly understood, abnormal protein aggregation and altered proteostasis are common features of sporadic and familial ALS forms. The proteostasis network is decomposed into different modules highly conserved across species and comprehends a collection of mechanisms related to protein synthesis, folding, trafficking, secretion and degradation that is distributed in different compartments inside the cell. Functional studies in various ALS models are revealing a complex scenario where distinct and even opposite effects in disease progression are observed depending on the targeted component of the proteostasis network. Importantly, alteration of the folding capacity of the endoplasmic reticulum (ER) is becoming a common pathological alteration in ALS, representing one of the earliest defects observed in disease models, contributing to denervation and motoneuron dysfunction. Strategies to target-specific components of the proteostasis network using small molecules and gene therapy are under development, and promise interesting avenues for future interventions to delay or stop ALS progression.


Assuntos
Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/fisiopatologia , Proteostase/fisiologia , Animais , Autofagia/fisiologia , Humanos , Neurônios Motores/metabolismo , Agregação Patológica de Proteínas/fisiopatologia , Dobramento de Proteína , Transporte Proteico/fisiologia , Fatores de Risco , Medula Espinal/metabolismo , Ubiquitinação
13.
Front Mol Neurosci ; 10: 216, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28725179

RESUMO

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the progressive loss of motoneurons and paralysis. The mechanisms underlying neuronal degeneration in ALS are starting to be elucidated, highlighting disturbances in motoneuron proteostasis. Endoplasmic reticulum (ER) stress has emerged as an early pathogenic event underlying motoneuron vulnerability and denervation in ALS. Maintenance of ER proteostasis is controlled by a dynamic signaling network known as the unfolded protein response (UPR). Inositol-requiring enzyme 1 (IRE1) is an ER-located kinase and endoribonuclease that operates as a major ER stress transducer, mediating the establishment of adaptive and pro-apoptotic programs. Here we discuss current evidence supporting the role of ER stress in motoneuron demise in ALS and build the rational to target IRE1 to ameliorate neurodegeneration.

14.
Acta Neuropathol ; 134(3): 489-506, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28341998

RESUMO

Altered proteostasis is a salient feature of Alzheimer's disease (AD), highlighting the occurrence of endoplasmic reticulum (ER) stress and abnormal protein aggregation. ER stress triggers the activation of the unfolded protein response (UPR), a signaling pathway that enforces adaptive programs to sustain proteostasis or eliminate terminally damaged cells. IRE1 is an ER-located kinase and endoribonuclease that operates as a major stress transducer, mediating both adaptive and proapoptotic programs under ER stress. IRE1 signaling controls the expression of the transcription factor XBP1, in addition to degrade several RNAs. Importantly, a polymorphism in the XBP1 promoter was suggested as a risk factor to develop AD. Here, we demonstrate a positive correlation between the progression of AD histopathology and the activation of IRE1 in human brain tissue. To define the significance of the UPR to AD, we targeted IRE1 expression in a transgenic mouse model of AD. Despite initial expectations that IRE1 signaling may protect against AD, genetic ablation of the RNase domain of IRE1 in the nervous system significantly reduced amyloid deposition, the content of amyloid ß oligomers, and astrocyte activation. IRE1 deficiency fully restored the learning and memory capacity of AD mice, associated with improved synaptic function and improved long-term potentiation (LTP). At the molecular level, IRE1 deletion reduced the expression of amyloid precursor protein (APP) in cortical and hippocampal areas of AD mice. In vitro experiments demonstrated that inhibition of IRE1 downstream signaling reduces APP steady-state levels, associated with its retention at the ER followed by proteasome-mediated degradation. Our findings uncovered an unanticipated role of IRE1 in the pathogenesis of AD, offering a novel target for disease intervention.


Assuntos
Doença de Alzheimer/metabolismo , Hipocampo/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais/fisiologia , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Modelos Animais de Doenças , Progressão da Doença , Estresse do Retículo Endoplasmático/fisiologia , Hipocampo/patologia , Humanos , Potenciação de Longa Duração/fisiologia , Proteínas de Membrana/genética , Camundongos , Camundongos Transgênicos , Neurônios/metabolismo , Neurônios/patologia , Proteínas Serina-Treonina Quinases/genética , Memória Espacial/fisiologia , Resposta a Proteínas não Dobradas/fisiologia
15.
Rev Med Chil ; 144(6): 796-806, 2016 Jun.
Artigo em Espanhol | MEDLINE | ID: mdl-27598501

RESUMO

Creutzfeldt-Jakob disease has a higher incidence in Chile than in other countries. The post mortem pathological characterization of brain tissue is necessary to reach a definitive diagnosis. We report a 73 years old man with a history compatible with of a rapidly progressive dementia, in which the first electroencephalographic study showed a pattern consistent with non-convulsive status epilepticus. Besides discarding this diagnosis, it was necessary to rule out other causes of rapidly progressive dementia such as Hashimoto encephalopathy. Finally, the sustained clinical deterioration with no response to anticonvulsants and corticosteroids, the imaging studies, a serial electroencephalographic monitoring study and the detection of 14-3-3 protein in cerebrospinal fluid were the keys to achieve the diagnosis of the disease.


Assuntos
Síndrome de Creutzfeldt-Jakob/diagnóstico , Proteínas 14-3-3/líquido cefalorraquidiano , Idoso , Autopsia , Eletroencefalografia , Evolução Fatal , Humanos , Imageamento por Ressonância Magnética , Masculino
16.
Rev. méd. Chile ; 144(6): 796-806, jun. 2016. ilus, tab
Artigo em Espanhol | LILACS | ID: lil-793988

RESUMO

Creutzfeldt-Jakob disease has a higher incidence in Chile than in other countries. The post mortem pathological characterization of brain tissue is necessary to reach a definitive diagnosis. We report a 73 years old man with a history compatible with of a rapidly progressive dementia, in which the first electroencephalographic study showed a pattern consistent with non-convulsive status epilepticus. Besides discarding this diagnosis, it was necessary to rule out other causes of rapidly progressive dementia such as Hashimoto encephalopathy. Finally, the sustained clinical deterioration with no response to anticonvulsants and corticosteroids, the imaging studies, a serial electroencephalographic monitoring study and the detection of 14-3-3 protein in cerebrospinal fluid were the keys to achieve the diagnosis of the disease.


Assuntos
Humanos , Masculino , Idoso , Síndrome de Creutzfeldt-Jakob/diagnóstico , Autopsia , Imageamento por Ressonância Magnética , Evolução Fatal , Proteínas 14-3-3/líquido cefalorraquidiano , Eletroencefalografia
17.
Prion ; 10(1): 50-6, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26864548

RESUMO

Disturbance of endoplasmic reticulum (ER) proteostasis is observed in Prion-related disorders (PrDs). The protein disulfide isomerase ERp57 is a stress-responsive ER chaperone up-regulated in the brain of Creutzfeldt-Jakob disease patients. However, the actual role of ERp57 in prion protein (PrP) biogenesis and the ER stress response remained poorly defined. We have recently addressed this question using gain- and loss-of-function approaches in vitro and animal models, observing that ERp57 regulates steady-state levels of PrP. Our results revealed that ERp57 modulates the biosynthesis and maturation of PrP but, surprisingly, does not contribute to the global cellular reaction against ER stress in neurons. Here we discuss the relevance of ERp57 as a possible therapeutic target in PrDs and other protein misfolding disorders.


Assuntos
Doenças Priônicas/metabolismo , Proteínas Priônicas/biossíntese , Proteínas Priônicas/metabolismo , Isomerases de Dissulfetos de Proteínas/metabolismo , Estresse do Retículo Endoplasmático , Humanos , Modelos Biológicos , Proteínas Priônicas/análise , Proteínas Priônicas/química , Isomerases de Dissulfetos de Proteínas/química , Dobramento de Proteína
18.
PLoS One ; 10(9): e0136620, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26361352

RESUMO

ERp57 (also known as grp58 and PDIA3) is a protein disulfide isomerase that catalyzes disulfide bonds formation of glycoproteins as part of the calnexin and calreticulin cycle. ERp57 is markedly upregulated in most common neurodegenerative diseases downstream of the endoplasmic reticulum (ER) stress response. Despite accumulating correlative evidence supporting a neuroprotective role of ERp57, the contribution of this foldase to the physiology of the nervous system remains unknown. Here we developed a transgenic mouse model that overexpresses ERp57 in the nervous system under the control of the prion promoter. We analyzed the susceptibility of ERp57 transgenic mice to undergo neurodegeneration. Unexpectedly, ERp57 overexpression did not affect dopaminergic neuron loss and striatal denervation after injection of a Parkinson's disease-inducing neurotoxin. In sharp contrast, ERp57 transgenic animals presented enhanced locomotor recovery after mechanical injury to the sciatic nerve. These protective effects were associated with enhanced myelin removal, macrophage infiltration and axonal regeneration. Our results suggest that ERp57 specifically contributes to peripheral nerve regeneration, whereas its activity is dispensable for the survival of a specific neuronal population of the central nervous system. These results demonstrate for the first time a functional role of a component of the ER proteostasis network in peripheral nerve regeneration.


Assuntos
Axônios/fisiologia , Isomerases de Dissulfetos de Proteínas/genética , Isomerases de Dissulfetos de Proteínas/metabolismo , Regeneração , Animais , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Corpo Estriado/metabolismo , Denervação , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/patologia , Feminino , Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Modelos Animais , Atividade Motora/genética , Degeneração Neural/genética , Degeneração Neural/patologia , Fenômenos Fisiológicos do Sistema Nervoso , Oxidopamina/farmacologia , Traumatismos dos Nervos Periféricos/genética , Traumatismos dos Nervos Periféricos/fisiopatologia , Traumatismos dos Nervos Periféricos/reabilitação
19.
J Biol Chem ; 290(39): 23631-45, 2015 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-26170458

RESUMO

Although the accumulation of a misfolded and protease-resistant form of the prion protein (PrP) is a key event in prion pathogenesis, the cellular factors involved in its folding and quality control are poorly understood. PrP is a glycosylated and disulfide-bonded protein synthesized at the endoplasmic reticulum (ER). The ER foldase ERp57 (also known as Grp58) is highly expressed in the brain of sporadic and infectious forms of prion-related disorders. ERp57 is a disulfide isomerase involved in the folding of a subset of glycoproteins in the ER as part of the calnexin/calreticulin cycle. Here, we show that levels of ERp57 increase mainly in neurons of Creutzfeldt-Jacob patients. Using gain- and loss-of-function approaches in cell culture, we demonstrate that ERp57 expression controls the maturation and total levels of wild-type PrP and mutant forms associated with human disease. In addition, we found that PrP physically interacts with ERp57, and also with the closest family member PDIA1, but not ERp72. Furthermore, we generated a conditional knock-out mouse for ERp57 in the nervous system and detected a reduction in the steady-state levels of the mono- and nonglycosylated forms of PrP in the brain. In contrast, ERp57 transgenic mice showed increased levels of endogenous PrP. Unexpectedly, ERp57 expression did not affect the susceptibility of cells to ER stress in vitro and in vivo. This study identifies ERp57 as a new modulator of PrP levels and may help with understanding the consequences of ERp57 up-regulation observed in human disease.


Assuntos
Príons/metabolismo , Isomerases de Dissulfetos de Proteínas/metabolismo , Animais , Linhagem Celular , Síndrome de Creutzfeldt-Jakob/metabolismo , Humanos , Camundongos , Camundongos Knockout , Neurônios/metabolismo , Dobramento de Proteína
20.
PLoS One ; 10(4): e0125146, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25928076

RESUMO

Docosahexaenoic acid (C22:6, n-3, DHA) is a polyunsaturated fatty acid highly enriched in the brain. This fatty acid can be easily oxidized yielding hydroperoxides as primary products. Cu, Zn-Superoxide dismutase (SOD1) aggregation is a common hallmark of Amyotrophic Lateral Sclerosis (ALS) and the molecular mechanisms behind their formation are not completely understood. Here we investigated the effect of DHA and its hydroperoxides (DHAOOH) on human SOD1 oligomerization in vitro. DHA induced the formation of high-molecular-weight (HMW) SOD1 species (>700 kDa). Aggregation was dependent on free thiols and occurred primarily with the protein in its apo-form. SOD1 incubation with DHA was accompanied by changes in protein structure leading to exposure of protein hydrophobic patches and formation of non-amyloid aggregates. Site-directed mutagenesis studies demonstrated that Cys 6 and Cys 111 in wild-type and Cys 6 in ALS-linked G93A mutant are required for aggregation. In contrast, DHAOOH did not induce HMW species formation but promoted abnormal covalent dimerization of apo-SOD1 that was resistant to SDS and thiol reductants. Overall, our data demonstrate that DHA and DHAOOH induce distinct types of apo-SOD1 oligomerization leading to the formation of HMW and low-molecular-weight species, respectively.


Assuntos
Ácidos Docosa-Hexaenoicos/metabolismo , Peróxido de Hidrogênio/metabolismo , Superóxido Dismutase/química , Superóxido Dismutase/metabolismo , Cromatografia em Gel , Eletroforese em Gel de Poliacrilamida , Ácidos Graxos/metabolismo , Humanos , Multimerização Proteica , Compostos de Sulfidrila/metabolismo
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