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1.
Adv Exp Med Biol ; 1185: 305-310, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31884629

RESUMO

Activating transcription factor 6 (ATF6) is a key regulator of the unfolded protein response (UPR). In response to endoplasmic reticulum (ER) stress, ATF6 is transported from the ER to the Golgi apparatus where it is cleaved by intramembrane proteolysis, releasing its cytosolic fragment. The cleaved ATF6 fragment, which is a basic leucine zipper (bZip) transcription factor, translocates to the nucleus and upregulates the expression of ER protein-folding chaperones and enzymes. Mutations in ATF6 cause heritable forms of cone photoreceptor dysfunction diseases. These mutations include missense, nonsense, splice site, and deletion or duplication changes found across the entire ATF6. To date, there are 11 ATF6 mutations reported, and we classified them into three classes based on their functional defects that interrupt distinct steps in the ATF6 signaling pathway.


Assuntos
Fator 6 Ativador da Transcrição/metabolismo , Células Fotorreceptoras Retinianas Cones/patologia , Transdução de Sinais , Estresse do Retículo Endoplasmático , Complexo de Golgi , Humanos , Mutação , Dobramento de Proteína
2.
Biomed Pharmacother ; 118: 109407, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31545290

RESUMO

The purpose of this study was to observe the functions of preconditioning with endoplasmic reticulum stress (ERS) whether alleviated heart ischemia/reperfusion injury (HI/RI) via modulating IRE1/ATF6/RACK1/PERK and PGC-1α expressions in diabetes mellitus (DM) or not. Diabetic rats were pretreated with 0.6 mg/kg tunicamycin (TM, 0.6 mg/kg tunicamycin was administered via intraperitoneal injection 30 minutes prior to the I/R procedures), and then subjected to 45 minutes of ischemia and 3 hours of reperfusion. Blood and myocardial tissues were collected, myocardial pathological injuries were investigated, serum creatine kinase-MB (CK-MB) and cardiac troponin T (cTnT) levels were measured, left ventricular systolic pressure (LVSP), left ventricular end diastolic pressure (LVEDP), maximum rate of left ventricular pressure rise (+dp/dtmax) and maximum rate of left ventricular pressure drop (-dp/dtmax) were evaluated, reactive oxygen species (ROS) and caspase-3 levels were observed, ΔΨm level and ROS expression were measured, and activated transcript factor 6 (ATF6), receptor for activated C kinase 1 (RACK1), PRK-like ER kinase (PERK), glucose regulated protein 78 (GRP78) and peroxisome proliferator-activated receptor γ co-activator 1-α (PGC-1α) expressions were assessed. The TM ameliorated the pathological damages, reduced myocardial oxidative stress damages, restrained apoptosis, and upregulated the expressions of ATF6, RACK1, PERK, GRP78 and PGC-1α compared with those of the ischemia/reperfusion (I/R) group in DM. This study suggested the preconditioning with endoplasmic reticulum stress (TM) strategy that could enhance protection against HI/RI in DM in clinical myocardial diseases.


Assuntos
Fator 6 Ativador da Transcrição/metabolismo , Diabetes Mellitus Experimental/patologia , Estresse do Retículo Endoplasmático , Proteínas de Membrana/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Quinase C Ativada/metabolismo , eIF-2 Quinase/metabolismo , Animais , Creatina Quinase/sangue , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/complicações , Proteínas de Choque Térmico/metabolismo , Masculino , Potencial da Membrana Mitocondrial , Traumatismo por Reperfusão Miocárdica/sangue , Traumatismo por Reperfusão Miocárdica/complicações , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Troponina T/metabolismo
3.
EMBO J ; 38(15): e100990, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31368601

RESUMO

Activation of the ATF6α signaling pathway is initiated by trafficking of ATF6α from the ER to the Golgi apparatus. Its subsequent proteolysis releases a transcription factor that translocates to the nucleus causing downstream gene activation. How ER retention, Golgi trafficking, and proteolysis of ATF6α are regulated and whether additional protein partners are required for its localization and processing remain unresolved. Here, we show that ER-resident oxidoreductase ERp18 associates with ATF6α following ER stress and plays a key role in both trafficking and activation of ATF6α. We find that ERp18 depletion attenuates the ATF6α stress response. Paradoxically, ER stress accelerates trafficking of ATF6α to the Golgi in ERp18-depleted cells. However, the translocated ATF6α becomes aberrantly processed preventing release of the soluble transcription factor. Hence, we demonstrate that ERp18 monitors ATF6α ER quality control to ensure optimal processing following trafficking to the Golgi.


Assuntos
Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Proteína Dissulfeto Redutase (Glutationa)/metabolismo , Ativação Transcricional , Linhagem Celular , Núcleo Celular/metabolismo , Retículo Endoplasmático/metabolismo , Deleção de Genes , Complexo de Golgi/metabolismo , Células HEK293 , Humanos , Transdução de Sinais , Resposta a Proteínas não Dobradas
4.
Oncogene ; 38(34): 6184-6195, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31312025

RESUMO

Missense mutations in the TP53 gene are frequent in human cancers, giving rise to mutant p53 proteins that can acquire oncogenic properties. Gain of function mutant p53 proteins can enhance tumour aggressiveness by promoting cell invasion, metastasis and chemoresistance. Accumulating evidences indicate that mutant p53 proteins can also modulate cell homeostatic processes, suggesting that missense p53 mutation may increase resistance of tumour cells to intrinsic and extrinsic cancer-related stress conditions, thus offering a selective advantage. Here we provide evidence that mutant p53 proteins can modulate the Unfolded Protein Response (UPR) to increase cell survival upon Endoplasmic Reticulum (ER) stress, a condition to which cancer cells are exposed during tumour formation and progression, as well as during therapy. Mechanistically, this action of mutant p53 is due to enhanced activation of the pro-survival UPR effector ATF6, coordinated with inhibition of the pro-apoptotic UPR effectors JNK and CHOP. In a triple-negative breast cancer cell model with missense TP53 mutation, we found that ATF6 activity is necessary for viability and invasion phenotypes. Together, these findings suggest that ATF6 inhibitors might be combined with mutant p53-targeting drugs to specifically sensitise cancer cells to endogenous or chemotherapy-induced ER stress.


Assuntos
Fator 6 Ativador da Transcrição/genética , Estresse do Retículo Endoplasmático/genética , Neoplasias/genética , Proteína Supressora de Tumor p53/genética , Resposta a Proteínas não Dobradas/genética , Fator 6 Ativador da Transcrição/metabolismo , Animais , Células Cultivadas , Progressão da Doença , Retículo Endoplasmático/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Camundongos , Camundongos Transgênicos , Mutação/fisiologia , Invasividade Neoplásica , Neoplasias/metabolismo , Neoplasias/patologia , Regulação para Cima
5.
Redox Biol ; 26: 101232, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31181458

RESUMO

There is more skeletal muscle tissue in the body than any other tissue and, as it is the organ of the majority of metabolic activity, muscle defect can affect the health of the entire body. Endoplasmic reticulum (ER) stress due to defects in protein folding/degradation balance, altered calcium and lipid levels and alterations in ER-mitochondria contacts has recently been recognised as the pathogenic cause of many different myopathies. In addition, a maladaptive ER stress response triggered by ER stress and mediated by three ER stress sensors (PERK, IRE1 and ATF6) is involved in a failure to relieve muscle tissue from this stress. Targeting ER stress and the ER stress response pathway offers a broad range of opportunities for treating myopathies but, as the inhibition of the three ER stress sensors may not be safe because it could lead to unexpected effects; it therefore calls for careful analysis of the changes in downstream signal transduction in the different myopathies so these sub-pathways can be pharmacologically targeted. This review summarises the known inhibitors of the ER stress response and the successful results obtained using some of them in mouse models of muscle diseases caused by ER stress/ER stress response.


Assuntos
Retículo Endoplasmático/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Doenças Musculares/tratamento farmacológico , Fenilbutiratos/farmacologia , Sulfonamidas/farmacologia , Tiofenos/farmacologia , Fator 6 Ativador da Transcrição/antagonistas & inibidores , Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Adenina/análogos & derivados , Adenina/farmacologia , Aldeídos/farmacologia , Animais , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/patologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Endorribonucleases/antagonistas & inibidores , Endorribonucleases/genética , Endorribonucleases/metabolismo , Humanos , Indóis/farmacologia , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Terapia de Alvo Molecular/métodos , Fibras Musculares Esqueléticas/efeitos dos fármacos , Fibras Musculares Esqueléticas/metabolismo , Fibras Musculares Esqueléticas/patologia , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Doenças Musculares/genética , Doenças Musculares/metabolismo , Doenças Musculares/patologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transdução de Sinais , eIF-2 Quinase/antagonistas & inibidores , eIF-2 Quinase/genética , eIF-2 Quinase/metabolismo
6.
Elife ; 82019 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-31149896

RESUMO

The unfolded protein response (UPR) detects and restores deficits in the endoplasmic reticulum (ER) protein folding capacity. Ceapins specifically inhibit the UPR sensor ATF6α, an ER-tethered transcription factor, by retaining it at the ER through an unknown mechanism. Our genome-wide CRISPR interference (CRISPRi) screen reveals that Ceapins function is completely dependent on the ABCD3 peroxisomal transporter. Proteomics studies establish that ABCD3 physically associates with ER-resident ATF6α in cells and in vitro in a Ceapin-dependent manner. Ceapins induce the neomorphic association of ER and peroxisomes by directly tethering the cytosolic domain of ATF6α to ABCD3's transmembrane regions without inhibiting or depending on ABCD3 transporter activity. Thus, our studies reveal that Ceapins function by chemical-induced misdirection which explains their remarkable specificity and opens up new mechanistic routes for drug development and synthetic biology.


Assuntos
Fator 6 Ativador da Transcrição/antagonistas & inibidores , Organelas/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Resposta a Proteínas não Dobradas , Transportadores de Cassetes de Ligação de ATP/metabolismo , Fator 6 Ativador da Transcrição/metabolismo , Sistemas CRISPR-Cas/genética , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Células HEK293 , Células Hep G2 , Humanos , Organelas/efeitos dos fármacos , Peroxissomos/efeitos dos fármacos , Peroxissomos/metabolismo , Fenótipo , Ligação Proteica/efeitos dos fármacos , Resposta a Proteínas não Dobradas/efeitos dos fármacos
7.
J Dairy Sci ; 102(8): 7359-7370, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31155263

RESUMO

Disruption of endoplasmic reticulum (ER) homeostasis, often termed ER stress, is intrinsically linked with perturbation of lipid metabolism in humans and mice. Whether ER homeostasis is affected in cows experiencing fatty liver is unknown. The aim of this study was to investigate the potential role of ER stress in hepatic lipid accumulation in calf hepatocytes and ER stress status in dairy cows with severe fatty liver. In vitro experiments were conducted in which hepatocytes were isolated from calves and treated with different concentrations of fatty acids, tauroursodeoxycholic acid (TUDCA; a canonical inhibitor of ER stress), or both. The increase in phosphorylation level of protein kinase RNA-like ER kinase (PERK) and inositol requiring protein-1α (IRE1α) proteins, and the cleavage of activating transcription factor-6 (ATF6) protein in response to increasing doses of fatty acids (which were reversed by TUDCA treatment) in primary hepatocytes underscored a mechanistic link between fatty acids and ER stress. In addition, fatty acid treatment increased the abundance of sterol regulatory element-binding protein 1c, acetyl-CoA carboxylase-α, fatty acid synthase, and diacylglycerol acyltransferase 1, and lipid accumulation in calf primary hepatocytes, whereas inhibition of ER stress by incubating with TUDCA significantly weakened these effects. Overall, results in vitro indicate that inhibition of ER stress in calf hepatocytes alleviates fatty acid-induced lipid accumulation by downregulating the expression of lipogenic genes. In vivo experiments, liver and blood samples were collected from cows diagnosed as healthy (n = 15) or with severe fatty liver (n = 15). The phosphorylation level of PERK and IRE1α, the cleavage of ATF6 protein, and the abundance of several unfolded protein response genes (78 kDa glucose-regulated protein, AMP-dependent transcription factor 4, and spliced X-box binding protein 1) were greater in liver of cows with severe fatty liver. The present in vivo study confirms the occurrence of ER stress in dairy cows with severe fatty liver. Considering the causative role of fatty acid-induced ER stress in hepatic lipid accumulation in calf hepatocytes, the existence of ER stress in the liver of severe fatty liver cows may presage its participation in fatty liver progression in dairy cows. However, the mechanistic relationship between ER stress and fatty liver in dairy cows remain to be determined.


Assuntos
Doenças dos Bovinos/fisiopatologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Ácidos Graxos/administração & dosagem , Fígado Gorduroso/veterinária , Metabolismo dos Lipídeos/efeitos dos fármacos , Fígado/metabolismo , Fator 6 Ativador da Transcrição/metabolismo , Animais , Bovinos , Células Cultivadas , Estresse do Retículo Endoplasmático/fisiologia , Endorribonucleases/metabolismo , Feminino , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Lipogênese/genética , Fígado/efeitos dos fármacos , Camundongos , Fosforilação , Ácido Tauroquenodesoxicólico/administração & dosagem , Resposta a Proteínas não Dobradas/genética , eIF-2 Quinase/metabolismo
8.
J Ethnopharmacol ; 238: 111857, 2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-30959142

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional Chinese medicine Astragalus membranaceus (Fisch.) Bunge (AM) has been utilized for the treatment of diabetes mellitus and its complications for centuries. Astragalus polysaccharides (APS), the main bioactive ingredient extracted from the root of AM, is prescribed widely in China and has definite cardioprotective effect during diabetic cardiomyopathy (DCM). Endoplasmic reticulum (ER) stress-induced apoptosis played a crucial role in the progression of DCM. However, the regulatory mechanisms of APS on ER stress pathway haven't been comprehensively studied so far. AIM OF THE STUDY: The aim of this study was to identify the effect of APS on cardiomyocyte apoptosis and to investigate the mechanisms for the anti-apoptotic effect of APS during DCM. MATERIALS AND METHODS: DCM rat model was induced by intraperitoneal streptozotocin (STZ) injection and treated with APS for 16 weeks. Cardiac function, pathological changes and apoptotic cells were assessed by echocardiography, hematoxylin-eosin (HE) staining and TUNEL assay, respectively. Expressions of key molecules in ER stress pathway were detected by Western blot analysis. Cardiomyocytes were exposed to high glucose (HG) and treated with APS for 24 h. Cell viability, apoptosis and protein expressions were assessed by MTT, flow cytometer and Western blot analysis, respectively. Moreover, lentivirus over-expressing (OE) C/EBP homologous protein (CHOP) was employed to further investigate the causative role of ER stress pathway in APS-mediated effect on cardiomyocyte apoptosis. RESULTS: In vivo, the results demonstrated that APS could improve heart function and attenuate myocardial apoptosis in DCM rat model. Further study demonstrated that APS could down-regulate the protein expressions of activating transcription factor 6 (ATF6) and protein kinase RNA-like ER kinase (PERK) related factors of ER stress pathway. In vitro, APS significantly inhibit HG stimulated H9C2 cell apoptosis and the expressions of ATF6 and PERK related proteins of ER stress pathway. However, after CHOP-OE lentivirus transfection, the protective effects of APS were diminished as increased apoptotic rate and higher expression of CHOP. CONCLUSIONS: APS could attenuate cardiomyocyte apoptosis via down-regulating the expression of ATF6 and PERK related factors of ER stress pathway in DCM rats and HG-stimulated H9C2 cells.


Assuntos
Astrágalo (Planta) , Cardiotônicos , Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 1 , Cardiomiopatias Diabéticas , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Polissacarídeos , Fator 6 Ativador da Transcrição/metabolismo , Animais , Apoptose/efeitos dos fármacos , Cardiotônicos/farmacologia , Cardiotônicos/uso terapêutico , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/metabolismo , Cardiomiopatias Diabéticas/tratamento farmacológico , Cardiomiopatias Diabéticas/metabolismo , Masculino , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/patologia , Polissacarídeos/farmacologia , Polissacarídeos/uso terapêutico , Ratos Sprague-Dawley , eIF-2 Quinase/metabolismo
9.
Arch Virol ; 164(5): 1323-1334, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30877450

RESUMO

Porcine circovirus type 2 (PCV2) is the essential infectious agent causing porcine circovirus-associated disease (PCVD) in pigs and one of the important viruses that severely jeopardize the swine husbandry industry. PCV2 elicits the unfolded protein response (UPR) via activation of the PERK pathway, and its capsid protein (Cap) has also been found to induce UPR with subsequent activation of apoptosis. The open reading frame 5 (ORF5) protein is a recently discovered non-structural protein, and its function in PCV2 pathogenesis remains unknown. The aim of this study was to determine whether the PCV2 ORF5 protein could induce endoplasmic reticulum stress (ERS) and UPR in porcine alveolar macrophages (PAMs). pEGFP-tagged ORF5 protein was transiently overexpressed in PAMs. Transmission electron microscopy (TEM) was employed to examine changes in ER morphology, and quantitative real-time PCR and western blotting analysis were used to measure UPR-related cell signaling alterations. We found that the ORF5 protein triggers swelling and degranulation of the ER and upregulates the expression of ERS markers. Further experiments demonstrated that the PCV2 ORF5 protein induces ERS and UPR via the PERK (RNA-activated protein kinase-like endoplasmic reticulum kinase), ATF6 (activating transcription factor 6) and IRE1 (inositol requiring enzyme 1) signaling pathways. Together with previous studies, we provide new information on the ERS-UPR induced by the PCV2 ORF5 protein.


Assuntos
Circovirus/genética , Estresse do Retículo Endoplasmático/genética , Retículo Endoplasmático/ultraestrutura , Macrófagos Alveolares/patologia , Resposta a Proteínas não Dobradas/genética , Proteínas do Envelope Viral/genética , Proteínas não Estruturais Virais/genética , Fator 6 Ativador da Transcrição/metabolismo , Animais , Linhagem Celular , Infecções por Circoviridae/patologia , Infecções por Circoviridae/veterinária , Retículo Endoplasmático/virologia , Endorribonucleases/metabolismo , Macrófagos Alveolares/virologia , Microscopia Eletrônica de Transmissão , Suínos , Doenças dos Suínos , Proteínas do Envelope Viral/metabolismo , eIF-2 Quinase/metabolismo
10.
Pharmazie ; 74(2): 115-119, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30782262

RESUMO

The current study aimed to evaluate the role and underlying mechanism of cyclic adenosine phosphate (cAMP) on the functional recovery of spinal cord injury (SCI). Basso, Beattie and Bresnahan (BBB) scoring and inclined plane test indicated that cAMP treatment improved the functional recovery of SCI rats. Real time PCR and western blot analysis showed the mRNA and protein levels of IRE1, PERK, and ATF6 were increased in the SCI rats than those of sham control. However, higher levels of IRE1, PERK, and ATF6 were indicated after cAMP treatment. Meanwhile, more apoptotic cells were observed in the SCI rats, as evidenced by TUNEL staining and increased expression of GRP78, CHOP, and caspase12. In contrast, the expression of GRP78, CHOP, and caspase12 was decreased in SCI rats after cAMP treatment. In summary, we showed novel data that cAMP reduced cell apoptosis and functional recover after SCI mainly via activating UPR.


Assuntos
AMP Cíclico/farmacologia , Traumatismos da Medula Espinal/tratamento farmacológico , Traumatismos da Medula Espinal/metabolismo , Resposta a Proteínas não Dobradas/efeitos dos fármacos , Fator 6 Ativador da Transcrição/metabolismo , Animais , Apoptose/efeitos dos fármacos , Caspase 12/metabolismo , AMP Cíclico/farmacocinética , Proteínas de Choque Térmico/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Atividade Motora/efeitos dos fármacos , Proteínas Serina-Treonina Quinases/metabolismo , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica , Medula Espinal/metabolismo , Medula Espinal/patologia , Traumatismos da Medula Espinal/patologia , Vértebras Torácicas/patologia , Fator de Transcrição CHOP/metabolismo , eIF-2 Quinase/metabolismo
11.
Viruses ; 11(2)2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30717233

RESUMO

To understand the underlying mechanisms of endoplasmic reticulum (ER) stress caused by human rhinovirus (HRV) 16 and non-structural transmembrane protein 2B, the expressions of ER chaperone glucose-regulated protein 78 (GRP78) and three signal transduction pathways, including protein kinase RNA-like ER kinase (PERK), activating transcription factor 6 (ATF6) and inositol-requiring enzyme 1 (IRE1), were evaluated after HRV16 infection and 2B gene transfection. Our results showed that both HRV16 infection and 2B gene transfection increased the expression of ER chaperone GRP78, and induced phosphorylation of PERK and cleavage of ATF6 in a time-dependent manner. Our data also revealed that the HRV16 2B protein was localized to the ER membrane. However, both HRV16 infection and HRV16 2B gene transfection did not induce ER stress through the IRE1 pathway. Moreover, our results showed that apoptosis occurred in H1-HeLa cells infected with HRV16 or transfected with 2B gene accompanied with increased expression of CHOP and cleaved caspase-3. Taken together, non-structural protein 2B of HRV16 induced an ER stress response through the PERK and ATF6 pathways rather than the IRE1 pathway.


Assuntos
Fator 6 Ativador da Transcrição/metabolismo , Estresse do Retículo Endoplasmático , Endorribonucleases/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Rhinovirus/metabolismo , Proteínas não Estruturais Virais/metabolismo , eIF-2 Quinase/metabolismo , Fator 6 Ativador da Transcrição/genética , Apoptose , Retículo Endoplasmático/metabolismo , Endorribonucleases/genética , Células HeLa , Proteínas de Choque Térmico/genética , Interações entre Hospedeiro e Microrganismos , Humanos , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Transdução de Sinais , Fator de Transcrição CHOP/genética , Transfecção , Proteínas não Estruturais Virais/genética , eIF-2 Quinase/genética
12.
Kidney Int ; 95(3): 577-589, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30639234

RESUMO

Tubulointerstitial fibrosis is a strong predictor of progression in patients with chronic kidney disease, and is often accompanied by lipid accumulation in renal tubules. However, the molecular mechanisms modulating the relationship between lipotoxicity and tubulointerstitial fibrosis remain obscure. ATF6α, a transcription factor of the unfolded protein response, is reported to be an upstream regulator of fatty acid metabolism. Owing to their high energy demand, proximal tubular cells (PTCs) use fatty acids as their main energy source. We therefore hypothesized that ATF6α regulates PTC fatty acid metabolism, contributing to lipotoxicity-induced tubulointerstitial fibrosis. Overexpression of activated ATF6α transcriptionally downregulated peroxisome proliferator-activated receptor-α (PPARα), the master regulator of lipid metabolism, leading to reduced activity of fatty acid ß-oxidation and cytosolic accumulation of lipid droplets in a human PTC line (HK-2). ATF6α-induced lipid accumulation caused mitochondrial dysfunction, enhanced apoptosis, and increased expression of connective tissue growth factor (CTGF), as well as reduced cell viability. Atf6α-/- mice had sustained expression of PPARα and less tubular lipid accumulation following unilateral ischemia-reperfusion injury (uIRI), resulting in the amelioration of apoptosis; reduced expression of CTGF, α-smooth muscle actin, and collagen I; and less tubulointerstitial fibrosis. Administration of fenofibrate, a PPARα agonist, reduced lipid accumulation and tubulointerstitial fibrosis in the uIRI model. Taken together, these findings suggest that ATF6α deranges fatty acid metabolism in PTCs, which leads to lipotoxicity-mediated apoptosis and CTGF upregulation, both of which promote tubulointerstitial fibrosis.


Assuntos
Fator 6 Ativador da Transcrição/metabolismo , Túbulos Renais Proximais/patologia , PPAR alfa/metabolismo , Insuficiência Renal Crônica/patologia , Fator 6 Ativador da Transcrição/genética , Animais , Linhagem Celular , Modelos Animais de Doenças , Regulação para Baixo , Ácidos Graxos/metabolismo , Fibrose , Humanos , Túbulos Renais Proximais/citologia , Túbulos Renais Proximais/metabolismo , Metabolismo dos Lipídeos , Camundongos , Camundongos Knockout , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Insuficiência Renal Crônica/etiologia
13.
Nat Commun ; 10(1): 187, 2019 01 14.
Artigo em Inglês | MEDLINE | ID: mdl-30643122

RESUMO

Pharmacologic activation of stress-responsive signaling pathways provides a promising approach for ameliorating imbalances in proteostasis associated with diverse diseases. However, this approach has not been employed in vivo. Here we show, using a mouse model of myocardial ischemia/reperfusion, that selective pharmacologic activation of the ATF6 arm of the unfolded protein response (UPR) during reperfusion, a typical clinical intervention point after myocardial infarction, transcriptionally reprograms proteostasis, ameliorates damage and preserves heart function. These effects were lost upon cardiac myocyte-specific Atf6 deletion in the heart, demonstrating the critical role played by ATF6 in mediating pharmacologically activated proteostasis-based protection of the heart. Pharmacological activation of ATF6 is also protective in renal and cerebral ischemia/reperfusion models, demonstrating its widespread utility. Thus, pharmacologic activation of ATF6 represents a proteostasis-based therapeutic strategy for ameliorating ischemia/reperfusion damage, underscoring its unique translational potential for treating a wide range of pathologies caused by imbalanced proteostasis.


Assuntos
Fator 6 Ativador da Transcrição/metabolismo , Infarto Cerebral/prevenção & controle , Nefropatias/prevenção & controle , Infarto do Miocárdio/prevenção & controle , Substâncias Protetoras/farmacologia , Traumatismo por Reperfusão/tratamento farmacológico , Fator 6 Ativador da Transcrição/genética , Animais , Animais Recém-Nascidos , Células Cultivadas , Infarto Cerebral/etiologia , Infarto Cerebral/patologia , Modelos Animais de Doenças , Retículo Endoplasmático/metabolismo , Feminino , Ventrículos do Coração/patologia , Humanos , Rim/irrigação sanguínea , Rim/patologia , Nefropatias/etiologia , Nefropatias/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infarto do Miocárdio/etiologia , Infarto do Miocárdio/patologia , Miócitos Cardíacos , Cultura Primária de Células , Substâncias Protetoras/uso terapêutico , Proteostase/efeitos dos fármacos , Ratos , Traumatismo por Reperfusão/etiologia , Resultado do Tratamento , Resposta a Proteínas não Dobradas/efeitos dos fármacos
14.
EBioMedicine ; 40: 695-709, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30685387

RESUMO

BACKGROUND: Mutations in the SLC26A2 gene cause a spectrum of currently incurable human chondrodysplasias. However, genotype-phenotype relationships of SLC26A2-deficient chondrodysplasias are still perplexing and thus stunt therapeutic development. METHODS: To investigate the causative role of SLC26A2 deficiency in chondrodysplasias and confirm its skeleton-specific pathology, we generated and analyzed slc26a2-/- and Col2a1-Cre; slc26a2fl/fl mice. The therapeutic effect of NVP-BGJ398, an FGFR inhibitor, was tested with both explant cultures and timed pregnant females. FINDINGS: Two lethal forms of human SLC26A2-related chondrodysplasias, achondrogenesis type IB (ACG1B) and atelosteogenesis type II (AO2), are phenocopied by slc26a2-/- mice. Unexpectedly, slc26a2-/- chondrocytes are defective for collagen secretion, exhibiting intracellular retention and compromised extracellular deposition of ColII and ColIX. As a consequence, the ATF6 arm of the unfolded protein response (UPR) is preferentially triggered to overactivate FGFR3 signaling by inducing excessive FGFR3 in slc26a2-/- chondrocytes. Consistently, suppressing FGFR3 signaling by blocking either FGFR3 or phosphorylation of the downstream effector favors the recovery of slc26a2-/- cartilage cultures from impaired growth and unbalanced cell proliferation and apoptosis. Moreover, administration of an FGFR inhibitor to pregnant females shows therapeutic effects on pathological features in slc26a2-/- newborns. Finally, we confirm the skeleton-specific lethality and pathology of global SLC26A2 deletion through analyzing the Col2a1-Cre; slc26a2fl/fl mouse line. INTERPRETATION: Our study unveils a previously unrecognized pathogenic mechanism underlying ACG1B and AO2, and supports suppression of FGFR3 signaling as a promising therapeutic approach for SLC26A2-related chondrodysplasias. FUND: This work was supported by National Natural Science Foundation of China (81871743, 81730065 and 81772377).


Assuntos
Acondroplasia/genética , Acondroplasia/metabolismo , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/metabolismo , Transdução de Sinais , Transportadores de Sulfato/deficiência , Resposta a Proteínas não Dobradas , Acondroplasia/patologia , Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Animais , Cartilagem/metabolismo , Cartilagem/patologia , Diferenciação Celular/genética , Condrócitos/citologia , Condrócitos/metabolismo , Modelos Animais de Doenças , Suscetibilidade a Doenças , Regulação da Expressão Gênica no Desenvolvimento , Lâmina de Crescimento/embriologia , Lâmina de Crescimento/patologia , Humanos , Camundongos , Camundongos Knockout , Morfogênese/genética , Mutação , Osteocondrodisplasias/genética , Osteocondrodisplasias/metabolismo , Osteocondrodisplasias/patologia , Fenótipo , Resposta a Proteínas não Dobradas/genética
15.
FASEB J ; 33(2): 2422-2434, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30260700

RESUMO

The importance of proteostasis in preventing cellular senescence has been well recognized. However, the exact mechanism by which the loss of proteostasis or endoplasmic reticulum (ER) stress induces cellular senescence remains unclear. We report that ER stress mediates cellular senescence through the activating transcription factor (ATF)6α branch of the unfolded protein response (UPR). Cellular senescence was induced by the abrogation of neighbor of breast cancer (BRCA)1 gene (NBR1). NBR1 abrogation-induced senescence was p53 dependent and observed in both transformed and nontransformed human cell lines: MCF-7, Caki-1, and MRC-5. NBR1 bound to p38 MAPK, preferentially to an active form, and upon NBR1 abrogation, the activity of p38 increased. NADPH oxidase was activated in turn by p38, and the resulting oxidative stress triggered ER stress. It was found that ER stress mediated cellular senescence through the UPR sensor ATF6α. Knockdown of ATF6α prevented senescence, whereas ATF6α overexpression triggered it. The transcriptional activity of ATF6α was important. The ER stress-ATF6α axis also mediated cellular senescence induced by H-RasV12 overexpression and UV irradiation, suggesting a common role of this axis in senescence induction. In summary, we presented an evidence for the novel role of the ER stress-ATF6α axis in cellular senescence.-Kim, H. S., Kim, Y., Lim, M. J., Park, Y.-G., Park, S. I., Sohn, J. The p38-activated ER stress-ATF6α axis mediates cellular senescence.


Assuntos
Fator 6 Ativador da Transcrição/metabolismo , Senescência Celular , Estresse do Retículo Endoplasmático , Proteínas/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Fator 6 Ativador da Transcrição/genética , Humanos , Células MCF-7 , NADPH Oxidase 2/genética , NADPH Oxidase 2/metabolismo , NADPH Oxidase 4/genética , NADPH Oxidase 4/metabolismo , Proteínas/genética , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/genética
16.
Hum Pathol ; 83: 22-28, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30121368

RESUMO

Little is known about the association between the atypical adenomatous hyperplasia (AAH)-adenocarcinoma in situ sequence of the lung and endoplasmic reticulum-stress responders such as ATF6, XBP1, and GRP78. Using stored tissues, we examined (i) the percentage of a splicing form (active form) of XBP1 messenger RNA in normal lung tissue (NLT) and adenocarcinoma (ACA; using reverse-transcription polymerase chain reaction); (ii) ATF6 and GRP78 protein expressions in NLT and ACA (using Western blotting analysis); (iii) ATF6, XBP1, and GRP78 protein expressions in NLT, AAH, and ACA, including some adenocarcinoma in situ (using immunohistochemistry); and (iv) the incidence of nuclear translocation of the 3 proteins in these lesions. The percentage of the splicing form of XBP1 messenger RNA showed a borderline difference between NLT and ACA (P = .068). In the Western blotting analysis, the nuclear fractions of ATF6 (including the active form) and GRP78 proteins were higher in ACA than in NLT. In the immunohistochemistry, the values obtained for the incidence of the nuclear translocation of ATF6, XBP1, and GRP78 proteins were as follows, respectively: 13.3%, 2.2%, and 0.5% in low-grade AAH; 37.9%, 2.3%, and 2.2% in high-grade AAH; and 47.2%, 10.6%, and 4.4% in ACA. A significant difference was detected between low-grade AAH and ACA for ATF6. In terms of nuclear translocation, high-grade AAH seemed intermediate between low-grade AAH and ACA. These results support endoplasmic reticulum-stress responses, such as nuclear translocation of these 3 proteins (including their active forms), being in parallel with the progression of the adenoma-carcinoma sequence in the lung.


Assuntos
Adenocarcinoma de Pulmão/patologia , Estresse do Retículo Endoplasmático/fisiologia , Neoplasias Pulmonares/patologia , Lesões Pré-Cancerosas/patologia , Fator 6 Ativador da Transcrição/metabolismo , Adenocarcinoma de Pulmão/metabolismo , Idoso , Progressão da Doença , Feminino , Proteínas de Choque Térmico/metabolismo , Humanos , Hiperplasia/metabolismo , Hiperplasia/patologia , Neoplasias Pulmonares/metabolismo , Masculino , Pessoa de Meia-Idade , Lesões Pré-Cancerosas/metabolismo , Transporte Proteico/fisiologia , Proteína 1 de Ligação a X-Box/metabolismo
17.
Prostate ; 79(2): 140-150, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30280407

RESUMO

BACKGROUND: Prostate cancer (PCa) is diagnosed at the highest rate of all non-cutaneous male cancers in the United States. The androgen-dependent (AD) transcription factor, androgen receptor (AR), drives PCa-but inhibiting AR or androgen biosynthesis induces remission for only a short time. At which point, patients acquire more aggressive castration-resistant (CR) disease with re-activated AR-dependent signaling. To combat treatment resistance, down-regulating AR protein expression has been considered as a potential treatment strategy for CR-PCa. METHODS: AD- and CR-PCa cell lines were treated with the well-tolerated FDA-approved oral medicine, riluzole. Expression of full-length or wild-type AR (AR-FL) and constitutively active AR-splice variant 7 (AR-V7) was assessed by immunoblotting. AR-FL/AR-V7 activity was measured using qRT-PCR of AR-target genes. Cytoplasmic [Ca2+ ] levels were measured using a fluorescent Ca2+ indicator microplate assay. Markers of the endoplasmic reticulum stress (ERS) pathway and autophagy were assessed by immunoblotting. Direct interaction between AR and selective autophagy receptor p62 was demonstrated by co-immunoprecipitation. RESULTS: We demonstrate that riluzole downregulates AR-FL, mutant ARs, and AR-V7 proteins expression by protein degradation through ERS pathway and selective autophagy. Riluzole also significantly inhibited AR transcription activity by decreasing its target genes expression (PSA, TMPRSS2, and KLK2). CONCLUSIONS: We provide key mechanistic insights by which riluzole exerts its anti-tumorigenic effects and induces AR protein degradation via ERS pathways. Our findings support the potential utility of riluzole for treatment of PCa.


Assuntos
Androgênios/metabolismo , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Neoplasias da Próstata/tratamento farmacológico , Receptores Androgênicos/metabolismo , Riluzol/farmacologia , Fator 6 Ativador da Transcrição/metabolismo , Autofagia/efeitos dos fármacos , Linhagem Celular Tumoral , Interações de Medicamentos , Endorribonucleases/antagonistas & inibidores , Endorribonucleases/metabolismo , Humanos , Masculino , Neoplasias da Próstata/metabolismo , Neoplasias de Próstata Resistentes à Castração/metabolismo , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Receptores Androgênicos/biossíntese , Proteína Sequestossoma-1/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sulfonamidas/farmacologia , Tiofenos/farmacologia
18.
Biochim Biophys Acta Mol Basis Dis ; 1865(2): 485-493, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30529145

RESUMO

Increasing evidence shows that the endoplasmic reticulum (ER) stress is an early event that injures pancreatic acinar cells and contributes to the pathogenesis of acute pancreatitis. In the present work we sought to establish whether atrial natriuretic peptide (ANP) alleviated ER stress in rats with cerulein-induced pancreatitis. The major components of the unfolded protein response (UPR) and their downstream effectors were assessed by immunoblotting or fluorimetry and the ultrastructure of ER evaluated by electron transmission microscopy. Cross-talk with autophagy was evaluated by beclin-1 expression. ANP reduced binding immunoglobulin protein (Bip) expression (UPR major controller) which under non-stress conditions keeps inactive the stress sensor proteins: protein kinase-like ER kinase (PERK), inositol-requiring enzyme-1 (IRE1) and activating transcription factor 6 (ATF6). Although ANP did not change PERK expression it decreased p-eIF2α and enhanced downstream effector CHOP, suggesting that ANP stimulates ER-dependent apoptosis. In accordance, ANP also decreased Bcl2 expression and enhanced proapoptotic proteins Bax and Bak. The atrial peptide enhanced ATF6 expression and although it did not affect IRE1/sXBP1 signaling, it increased caspase-2 activity, also involved in ER-dependent apoptosis. Furthermore, ANP decreased beclin-1 expression. The ultrastructure of the RE revealed decreased swelling and conserved ribosomes in the presence of ANP. Present findings support that ANP alleviates ER stress in acute pancreatitis by modulating the three branches of the UPR and stimulates ER-dependent apoptosis. Gaining insights into the modulation of ER stress may help to develop specific therapeutic strategies for acute pancreatitis and/or medical interventions at risk of its developing like endoscopic retrograde cholangiopancreatography.


Assuntos
Fator Natriurético Atrial/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Pancreatite/patologia , Fator 6 Ativador da Transcrição/metabolismo , Doença Aguda , Animais , Apoptose/efeitos dos fármacos , Proteína Beclina-1/metabolismo , Caspase 12/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/ultraestrutura , Ativação Enzimática/efeitos dos fármacos , Fator de Iniciação 2 em Eucariotos/metabolismo , Proteínas de Choque Térmico/metabolismo , Proteínas de Membrana/metabolismo , Pâncreas/efeitos dos fármacos , Pâncreas/metabolismo , Pâncreas/patologia , Pâncreas/ultraestrutura , Pancreatite/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Resposta a Proteínas não Dobradas/efeitos dos fármacos , Proteína 1 de Ligação a X-Box/metabolismo , eIF-2 Quinase/metabolismo
19.
J Physiol Biochem ; 75(1): 73-81, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30426460

RESUMO

The protective effects of downregulated miR-199a-5p on ischemic and hypoxic cardiomyocytes were well recognized, but the underlying mechanism of inhibited miR-199a-5p is not yet clear. The present study explored the relationship between enhanced signal transducer and activator of transcription 3 (STAT3) signaling and lowered production of miR-199a-5p in hypoxic cardiomyocytes. This study firstly found the correlation between elevated interleukin (IL)-6 and IL-11, as well as subsequent STAT3 signaling activation and the downregulation of miR-199a-5p in hypoxic myocardial samples from children with congenital heart disease. Then, using model of hypoxic mice and the intervention of phosphorylated STAT3 (pSTAT3), it was observed that pSTAT3 affected the expression of miR-199a-5p and modulated the expression of its target genes, including endoplasmic reticulum stress (ERS)-related activating transcription factor 6 (ATF6) and 78 kDa glucose-regulated protein (GRP78). Further observation revealed that the pSTAT3 signal in cardiac tissue could affect the expression of pri-miR-199a-2, a precursor of miR-199a-5p. And the chromatin immunoprecipitation (ChIP) assay also confirmed that pSTAT3 could bind to the promoter region of miR-199a-2 gene, which is more significant under hypoxic conditions. In conclusion, the activation of STAT3 signaling in cardiomyocytes during chronic hypoxia leads to downregulation of miR-199a-5p, which promotes the expression of many downstream target genes. This is an important pathway in the adaptive protection mechanism of myocardium during hypoxia.


Assuntos
Fístula Arteriovenosa/genética , Cardiopatias Congênitas/genética , Hipóxia/genética , MicroRNAs/genética , Miócitos Cardíacos/metabolismo , Artéria Pulmonar/anormalidades , Veias Pulmonares/anormalidades , Fator de Transcrição STAT3/genética , Tetralogia de Fallot/genética , Fator 6 Ativador da Transcrição/genética , Fator 6 Ativador da Transcrição/metabolismo , Adolescente , Animais , Fístula Arteriovenosa/metabolismo , Fístula Arteriovenosa/patologia , Fístula Arteriovenosa/cirurgia , Ponte Cardiopulmonar , Criança , Pré-Escolar , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica , Cardiopatias Congênitas/metabolismo , Cardiopatias Congênitas/patologia , Cardiopatias Congênitas/cirurgia , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Hipóxia/metabolismo , Hipóxia/patologia , Interleucina-11/genética , Interleucina-11/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Masculino , Camundongos Endogâmicos BALB C , MicroRNAs/metabolismo , Miocárdio/metabolismo , Miocárdio/patologia , Miócitos Cardíacos/patologia , Regiões Promotoras Genéticas , Artéria Pulmonar/metabolismo , Artéria Pulmonar/patologia , Artéria Pulmonar/cirurgia , Veias Pulmonares/metabolismo , Veias Pulmonares/patologia , Veias Pulmonares/cirurgia , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais , Tetralogia de Fallot/metabolismo , Tetralogia de Fallot/patologia , Tetralogia de Fallot/cirurgia , Resposta a Proteínas não Dobradas
20.
J Endocrinol ; 240(2): 181-193, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30400033

RESUMO

The endoplasmic reticulum (ER) stress and inflammation relationship occurs at different levels and is essential for the adequate homeostatic function of cellular systems, becoming harmful when chronically engaged. Intense physical exercise enhances serum levels of interleukin 6 (IL-6). In response to a chronic exhaustive physical exercise protocol, our research group verified an increase of the IL-6 concentration and ER stress proteins in extensor digitorium longus (EDL) and soleus. Based on these results, we hypothesized that IL-6-knockout mice would demonstrate a lower modulation in the ER stress proteins compared to the wild-type mice. To clarify the relationship between exercise-induced IL-6 increased and ER stress, we studied the effects of an acute exhaustive physical exercise protocol on the levels of ER stress proteins in the skeletal muscles of IL-6-knockout (KO) mice. The WT group displayed a higher exhaustion time compared to the IL-6 KO group. After 1 h of the acute exercise protocol, the serum levels of IL-6 and IL-10 were enhanced in the WT group. Independent of the experimental group, the CHOP and cleaved caspase 12/total caspase 12 ratio in EDL as well as ATF6 and CHOP in soleus were sensitive to the acute exercise protocol. Compared to the WT group, the oscillation patterns over time of BiP in EDL and soleus as well as of peIF2-alpha/eIF2-alpha ratio in soleus were attenuated for the IL-6 KO group. In conclusion, IL-6 seems to be related with the ER stress homeostasis, once knockout mice presented attenuation of BiP in EDL and soleus as well as of pEiF2-alpha/EiF2-alpha ratio in soleus after the acute exhaustive physical exercise protocol.


Assuntos
Estresse do Retículo Endoplasmático/fisiologia , Interleucina-6/metabolismo , Músculo Esquelético/metabolismo , Condicionamento Físico Animal/fisiologia , Fator 6 Ativador da Transcrição/metabolismo , Animais , Western Blotting , Caspases/metabolismo , Estresse do Retículo Endoplasmático/genética , Interleucina-10/sangue , Interleucina-10/metabolismo , Interleucina-6/sangue , Interleucina-6/genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator de Transcrição CHOP/metabolismo
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