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1.
DNA Cell Biol ; 39(3): 428-440, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31977244

RESUMO

In this study, we analyzed the gene structure, chemical characterizations, chromosome locations, evolutionary relationship, and expression profile of hsp90 genes with online database. In addition, the expression levels of hsp90s were also investigated under heat stress by quantitative real-time (qRT)-PCR. A total of eight hsp90 genes were identified from the rainbow trout genome. They were all distributed on chromosomes 2, 4, 8, and 13. The molecular weight ranged from 78.93 to 91.39 kDa, and the isoelectric point ranged from 4.84 to 4.96. The eight hsp90 genes were clustered into six subfamilies (A, B, C, D, E, and F). Genetic structure and conserved domain analysis revealed that all eight hsp90 genes had only one exon, and motif 1-motif 10 was shared by most genes. According to RNA-seq analysis of rainbow trout liver and head kidney, a total of seven out of eight genes were significantly upregulated under heat stress, and qRT-PCR was carried out on these seven genes; the expression levels of these genes were significantly upregulated under heat stress. The significantly regulated expressions of hsp90 genes under heat stress indicated that hsp90 genes are involved in heat stress response in rainbow trout. This study provides a theoretical basis for further study on the role of hsp90 in the heat stress tolerance of rainbow trout.


Assuntos
Proteínas de Peixes/genética , Proteínas de Choque Térmico HSP90/genética , Resposta ao Choque Térmico , Truta/genética , Animais , Proteínas de Peixes/química , Proteínas de Peixes/metabolismo , Proteínas de Choque Térmico HSP90/química , Proteínas de Choque Térmico HSP90/metabolismo , Domínios Proteicos , Truta/metabolismo , Regulação para Cima
2.
Gene ; 727: 144232, 2020 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-31715300

RESUMO

Colorectal cancer (CRC) is a global disease with high incidence and mortality rate. Hsp90 inhibitors induce cell death in various cancers, including CRC. However, the underlying mechanisms need to be clarified further. In this study, Caco-2 cells were treated with 0.25 µM SNX-2112, an Hsp90 inhibitor, for 48 h; subsequently, whole-transcriptome sequencing was performed. At the mRNA level in SNX-2112-treated Caco-2 cells, 1588 genes were upregulated, and 433 genes were downregulated. Six genes were found to be associated with necroptosis and apoptosis, and these 6 upregulated genes were validated by RT-qPCR. Hundred and six miRNAs were upregulated, and 48 miRNAs were downregulated in SNX-2112-treated Caco-2 cells. Eleven downregulated miRNAs were found to interact with the 6 upregulated genes. Moreover, 676 circRNAs were upregulated, and 291 circRNAs were downregulated in SNX-2112-treated Caco-2 cells. Among them, 126 circRNAs were found to be the target of the 11 downregulated miRNAs. The circRNA-miRNA-mRNA regulatory network of Hsp90 inhibitor-induced cell death in colorectal cancer was constructed. This regulatory network extends the underlying mechanism of Hsp90 and improves our understanding of Hsp90 inhibitors as potential targeted therapeutic agents.


Assuntos
Neoplasias Colorretais/genética , Redes Reguladoras de Genes/genética , Células CACO-2 , Neoplasias do Colo/genética , Neoplasias Colorretais/metabolismo , Regulação para Baixo , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Ontologia Genética , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Proteínas de Choque Térmico HSP90/genética , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Humanos , MicroRNAs/genética , Análise de Sequência com Séries de Oligonucleotídeos , RNA/genética , RNA/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Regulação para Cima
3.
Gene ; 729: 144319, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31884108

RESUMO

In previous study, we have found that microRNA-23a is down regulated in atherosclerotic tissues. Here we demonstrate that miR-23a directly binds to 3'UTR of HSP90 mRNA to suppress the expression of HSP90. To investigate the potential roles of miR-23a in macrophage, THP-1 macrophages were transfected with miR-23a mimics or inhibitors. Our results showed inflammatory factors IL-6 and MCP-1 concentrations in cell culture medium of macrophage and foam cell transfected with miR-23a mimics were decreased. Furthermore, we find that apoptosis of macrophage and foam cells transfected with miR-23a mimics were inhibited. Over expression of miR-23a in foam cells could reduced lipid intake and accumulation in foam cells. Meanwhile, we found that in inflammatory macrophages and foam cells transfected with miR-23a mimcs, HSP90 and NF-κB proteins are significantly decreased. Our results have suggested a promising and potential therapeutic target for atherosclerosis.


Assuntos
Aterosclerose/genética , Aterosclerose/patologia , Células Espumosas/patologia , Proteínas de Choque Térmico HSP90/genética , Macrófagos/patologia , MicroRNAs/genética , Regiões 3' não Traduzidas , Apoptose/genética , Aterosclerose/metabolismo , Proliferação de Células/genética , Células Espumosas/metabolismo , Humanos , Inflamação/genética , Macrófagos/metabolismo , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Células THP-1
4.
Neurochem Res ; 44(11): 2643-2657, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31606837

RESUMO

Schwann cells (SCs) play an important role in producing myelin for rapid neurotransmission in the peripheral nervous system. Activation of the differentiation and myelination processes in SCs requires the expression of a series of transcriptional factors including Sox10, Oct6/Pou3f1, and Egr2/Krox20. However, functional interactions among several transcription factors are poorly defined and the important components of the regulatory network are still unknown. Until now, available evidence suggests that SCs require cAMP signaling to initiate the myelination program. Heat shock protein 90 (Hsp90) is known as a chaperone required to stabilize ErbB2 receptor. In recent years, it was reported that cAMP transactivated the ErbB2/ErbB3 signaling in SCs. However, the relationship between Hsp90 and cAMP-induced differentiation in SCs is undefined. Here we investigated the role of Hsp90 during cAMP-induced differentiation of SCs using Hsp90 inhibitor, geldanamycin and Hsp90 siRNA transfection. Our results showed that dibutyryl-cAMP (db-cAMP) treatment upregulated Hsp90 expression and led to nuclear translocation of Gab1/ERK, the downstream signaling pathway of the ErbB2 signaling mechanism in myelination. The expression of myelin-related genes and nuclear translocation of Gab1/ERK following db-cAMP treatment was inhibited by geldanamycin pretreatment and Hsp90 knockdown. These findings suggest that Hsp90 might play a role in cAMP-induced differentiation via stabilization of ErbB2 and nuclear translocation of Gab1/ERK in SCs.


Assuntos
Diferenciação Celular/fisiologia , Proteínas de Choque Térmico HSP90/fisiologia , Células de Schwann/fisiologia , Animais , Benzoquinonas/farmacologia , Bucladesina/farmacologia , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Expressão Gênica , Técnicas de Silenciamento de Genes , Proteínas de Choque Térmico HSP90/genética , Lactamas Macrocíclicas/farmacologia , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Ratos Sprague-Dawley , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Células de Schwann/citologia , Regulação para Cima
5.
Int J Mol Sci ; 20(20)2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31600883

RESUMO

Proper folding is crucial for proteins to achieve functional activity in the cell. However, it often occurs that proteins are improperly folded (misfolded) and form aggregates, which are the main hallmark of many diseases including cancers, neurodegenerative diseases and many others. Proteins that assist other proteins in proper folding into three-dimensional structures are chaperones and co-chaperones. The key role of chaperones/co-chaperones is to prevent protein aggregation, especially under stress. An imbalance between chaperone/co-chaperone levels has been documented in neurons, and suggested to contribute to protein misfolding. An essential protein and a major regulator of protein folding in all eukaryotic cells is the heat shock protein 90 (Hsp90). The function of Hsp90 is tightly regulated by many factors, including co-chaperones. In this review we summarize results regarding the role of Hsp90 and its co-chaperones in neurodegenerative disorders such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), and prionopathies.


Assuntos
Suscetibilidade a Doenças , Proteínas de Choque Térmico HSP90/metabolismo , Chaperonas Moleculares/metabolismo , Doenças Neurodegenerativas/etiologia , Doenças Neurodegenerativas/metabolismo , Animais , Biomarcadores , Regulação da Expressão Gênica , Proteínas de Choque Térmico HSP90/genética , Humanos , Chaperonas Moleculares/genética , Doenças Neurodegenerativas/diagnóstico , Doenças Neurodegenerativas/tratamento farmacológico , Transdução de Sinais
6.
Pol J Vet Sci ; 22(3): 565-572, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31560472

RESUMO

In broiler chickens, the relationship between dietary supplementation of vitamin C and hepatic, cardiac and renal heat shock proteins (HSP60, HSP70 and HSP90), heat shock factors (HSF-1 and HSF-3) and enzymatic antioxidants requires further investigation. The current study aimed to investigate this relationship at cellular and molecular levels in a 42 days experiment. Two hundred, one-day-old broiler chicks (Ross 308) were allocated into four equal groups. Chicks in the first and third groups were thermo-neutral (TN; 22°C for 24 hours/day) and fed basal diet without or with vitamin C (1g/kg basal diet), respectively. Chicks in the second and fourth groups were heat stressed (HS; 34°C for 8 hours/day) and fed basal diet without or with vitamin C, respectively. Performance parameters were recorded throughout the experiment. Levels of malondialdehyde (MDA), superoxide dismutase (SOD), glutathione S-transferase (GST), glutathione peroxidase (GPX), Catalase (CAT) and gene expression of heat shock proteins (HSP60, 70 and 90) and heat shock factors (HSF 1 and 3) were analyzed in liver, heart and kidney tissues of the studied groups. Heat stress induced a negative impact on performance parameters, significant reduction in activities of all examined antioxidant enzymes and a significant up-regulation in heat shock proteins and factors genes in all studied tissues. Dietary supplementation of vitamin C corrected these parameters towards the normal control values. Conclusively, dietary supplementation of the examined dose of vitamin C was efficient at ameliorating the detrimental effects of heat stress on liver, heart and kidney tissues of broilers chickens at cellular and molecular levels.


Assuntos
Antioxidantes/metabolismo , Ácido Ascórbico/uso terapêutico , Galinhas , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Choque Térmico/metabolismo , Animais , Chaperonina 60/genética , Chaperonina 60/metabolismo , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Fatores de Transcrição de Choque Térmico/genética , Fatores de Transcrição de Choque Térmico/metabolismo , Proteínas de Choque Térmico/genética , Resposta ao Choque Térmico , Rim/enzimologia , Peroxidação de Lipídeos , Fígado/enzimologia , Miocárdio/enzimologia , Doenças das Aves Domésticas/tratamento farmacológico
7.
Int J Mol Sci ; 20(18)2019 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-31527404

RESUMO

Heat Shock Protein 90 (Hsp90) chaperone interacts with a broad range of client proteins involved in cancerogenesis and cancer progression. However, Hsp90 inhibitors were unsuccessful as anticancer agents due to their high toxicity, lack of selectivity against cancer cells and extrusion by membrane transporters responsible for multidrug resistance (MDR) such as P-glycoprotein (P-gp). Recognizing the potential of new compounds to inhibit P-gp function and/or expression is essential in the search for effective anticancer drugs. Eleven Hsp90 inhibitors containing an isoxazolonaphtoquinone core were synthesized and evaluated in two MDR models comprised of sensitive and corresponding resistant cancer cells with P-gp overexpression (human non-small cell lung carcinoma and colorectal adenocarcinoma). We investigated the effect of Hsp90 inhibitors on cell growth inhibition, P-gp activity and P-gp expression. Structure-activity relationship analysis was performed in respect to cell growth and P-gp inhibition. Compounds 5, 7, and 9 directly interacted with P-gp and inhibited its ATPase activity. Their potential P-gp binding site was identified by molecular docking studies. In addition, these compounds downregulated P-gp expression in MDR colorectal carcinoma cells, showed good relative selectivity towards cancer cells, while compound 5 reversed resistance to doxorubicin and paclitaxel in concentration-dependent manner. Therefore, compounds 5, 7 and 9 could be promising candidates for treating cancers with P-gp overexpression.


Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Antineoplásicos/farmacologia , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Perfilação da Expressão Gênica , Proteínas de Choque Térmico HSP90/química , Proteínas de Choque Térmico HSP90/genética , Humanos , Modelos Moleculares , Conformação Molecular , Estrutura Molecular , Relação Estrutura-Atividade
8.
Cell Physiol Biochem ; 53(3): 480-495, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31486323

RESUMO

BACKGROUND/AIMS: Hypoxia Inducible Factor-1α (HIF-1α) is involved in cancer progression and is stabilized by the chaperone HSP90 (Heat Shock Protein 90), preventing degradation. Previously identified HSP90 inhibitors bind to the N-terminal pocket of HSP90, which blocks binding to HIF-1α and induces HIF-1α degradation. N-terminal inhibitors have failed in the clinic as single therapy treatments partially because they induce a heat shock response. SM molecules are HSP90 inhibitors that bind to the C-terminus of HSP90 and do not induce a heat shock response. The effects of these C-terminal inhibitors on HIF-1α are unreported. METHODS: HCT116, MDA-MB-231, PC3, and HEK293T cells were treated with HSP90 inhibitors. qRT-PCR and western blotting was performed to assess mRNA and protein levels of HIF-1α, HSP- and RACK1-related genes. siRNA was used to knockdown RACK1, while MG262 was used to inhibit proteasome activity. Dimethyloxalylglycine (DMOG) was used to inhibit activity of the prolyl hydroxylases (PHDs). Anti-angiogenic activity of HSP90 inhibitors was assessed using a HUVEC tubule formation assay. RESULTS: We show that SM compounds decrease HIF-1α target expression at the mRNA and protein level under hypoxia in colorectal, breast and prostate cancer cells, leading to cell death, without inducing a heat shock response. Surprisingly, we found that when the C-terminal of HSP90 is inhibited, HIF-1α degradation occurs through the proteasome and prolyl hydroxylases in an oxygen-dependent manner even in very low levels of oxygen (tumor hypoxia levels). RACK1 was not required for proteasomal degradation of HIF-1α. CONCLUSION: Our results suggest that by targeting the C-terminus of HSP90 we can exploit the prolyl hydroxylase and proteasome pathway to induce HIF-1α degradation in hypoxic tumors.


Assuntos
Hipóxia Celular/fisiologia , Proteínas de Choque Térmico HSP90/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Aminoácidos Dicarboxílicos/metabolismo , Western Blotting , Hipóxia Celular/genética , Sobrevivência Celular/genética , Sobrevivência Celular/fisiologia , Células HCT116 , Células HEK293 , Proteínas de Choque Térmico HSP90/genética , Células Endoteliais da Veia Umbilical Humana , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Células PC-3 , Prolil Hidroxilases/genética , Prolil Hidroxilases/metabolismo , Complexo de Endopeptidases do Proteassoma/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
9.
Nat Commun ; 10(1): 3626, 2019 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-31399574

RESUMO

The molecular chaperone Hsp90 is an important regulator of proteostasis. It has remained unclear why S. cerevisiae possesses two Hsp90 isoforms, the constitutively expressed Hsc82 and the stress-inducible Hsp82. Here, we report distinct differences despite a sequence identity of 97%. Consistent with its function under stress conditions, Hsp82 is more stable and refolds more efficiently than Hsc82. The two isoforms also differ in their ATPases and conformational cycles. Hsc82 is more processive and populates closed states to a greater extent. Variations in the N-terminal ATP-binding domain modulate its dynamics and conformational cycle. Despite these differences, the client interactomes are largely identical, but isoform-specific interactors exist both under physiological and heat shock conditions. Taken together, changes mainly in the N-domain create a stress-specific, more resilient protein with a shifted activity profile. Thus, the precise tuning of the Hsp90 isoforms preserves the basic mechanism but adapts it to specific needs.


Assuntos
Proteínas de Choque Térmico HSP90/química , Chaperonas Moleculares/química , Isoformas de Proteínas/química , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/metabolismo , Adenosina Trifosfatases/metabolismo , Sequência de Aminoácidos , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Proteínas de Choque Térmico HSP90/fisiologia , Resposta ao Choque Térmico/fisiologia , Ligantes , Modelos Moleculares , Chaperonas Moleculares/metabolismo , Ligação Proteica , Conformação Proteica , Dobramento de Proteína , Isoformas de Proteínas/metabolismo , Estabilidade Proteica , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Alinhamento de Sequência , Estresse Fisiológico
10.
Nat Commun ; 10(1): 3613, 2019 08 09.
Artigo em Inglês | MEDLINE | ID: mdl-31399576

RESUMO

Small-molecule inhibitors for the 90-kDa heat shock protein (HSP90) have been extensively exploited in preclinical studies for the therapeutic interventions of human diseases accompanied with proteotoxic stress. By using an unbiased quantitative proteomic method, we uncover that treatment with three HSP90 inhibitors results in elevated expression of a large number of heat shock proteins. We also demonstrate that the HSP90 inhibitor-mediated increase in expression of DNAJB4 protein occurs partly through an epitranscriptomic mechanism, and is substantially modulated by the writer, eraser, and reader proteins of N6-methyladenosine (m6A). Furthermore, exposure to ganetespib leads to elevated modification levels at m6A motif sites in the 5'-UTR of DNAJB4 mRNA, and the methylation at adenosine 114 site in the 5'-UTR promotes the translation of the reporter gene mRNA. This m6A-mediated mechanism is also at play upon heat shock treatment. Cumulatively, we unveil that HSP90 inhibitors stimulate the translation of DNAJB4 through an epitranscriptomic mechanism.


Assuntos
Adenosina/análogos & derivados , Proteínas de Choque Térmico HSP40/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Proteômica , Regiões 5' não Traduzidas , Adenosina/metabolismo , Linhagem Celular Tumoral , Genes Reporter , Proteínas de Choque Térmico HSP40/genética , Proteínas de Choque Térmico HSP90/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Metilação , RNA Mensageiro/metabolismo , Triazóis
11.
Acupunct Med ; 37(6): 340-347, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31412703

RESUMO

OBJECTIVE: To study the effects of acupuncture on expression of heat shock protein (Hsp) 84 and 86, and brain ageing, in the senescence accelerated mouse prone 8 (SAMP8) model of Alzheimer's disease. METHODS: 7-month-old male senescence resistant mouse strain 1 (SAMR1) and SAMP8 mice were assigned to the following groups, with 15 animals in each group: SAMR1 control (Rc), SAMP8 control (Pc), SAMP8 acupuncture (Pa), SAMP8 sham-acupuncture (Psa). The Pa group was given acupuncture treatment once daily for 15 days. Neuromuscular coordination and cognitive function of the mice were evaluated by the tightrope test and Morris water maze test, respectively. The number of neurons in the CA1, CA3 and dentate gyrus (DG) regions of the hippocampus were measured. The levels of oxidative stress and protein carbonyl, mRNA and protein expression levels of Hsp84 and Hsp86 in the hippocampus were detected. RESULTS: Compared with the Rc group, in the Pc mice there was a lower success rate for the tightrope test, impaired cognitive abilities, a decline in neuron numbers, reduced levels of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), increased levels of superoxide anion and protein carbonyl, and decreased mRNA and protein levels of Hsp84 and Hsp86 (all P<0.05). After acupuncture treatment, the success rate for the tightrope test was elevated, cognitive function was improved, neuron numbers were enhanced, levels of SOD and GSH-Px were increased, levels of superoxide anion and protein carbonyl were decreased, and Hsp84 and Hsp86 mRNA and protein expression were increased in the Pa mice when compared with the Pc and Psa groups (all P<0.05). CONCLUSION: Acupuncture may delay brain ageing in SAMP8 mice by reducing oxidative protein damage and promoting Hsp84 and Hsp86 expression.


Assuntos
Terapia por Acupuntura , Doença de Alzheimer/psicologia , Doença de Alzheimer/terapia , Proteínas de Choque Térmico HSP90/metabolismo , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Animais , Cognição , Modelos Animais de Doenças , Proteínas de Choque Térmico HSP90/genética , Humanos , Masculino , Camundongos
13.
Nat Commun ; 10(1): 3499, 2019 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-31375671

RESUMO

Long non-coding RNAs (lncRNAs) contribute to colorectal cancer (CRC). However, the role of lncRNAs in CRC metabolism, especially glucose metabolism remains largely unknown. In this study, we identify a lncRNA, GLCC1, which is significantly upregulated under glucose starvation in CRC cells, supporting cell survival and proliferation by enhancing glycolysis. Mechanistically, GLCC1 stabilizes c-Myc transcriptional factor from ubiquitination by direct interaction with HSP90 chaperon and further specifies the transcriptional modification pattern on c-Myc target genes, such as LDHA, consequently reprogram glycolytic metabolism for CRC proliferation. Clinically, GLCC1 is associated with tumorigenesis, tumor size and predicts poor prognosis. Thus, GLCC1 is mechanistically, functionally, and clinically oncogenic in colorectal cancer. Targeting GLCC1 and its pathway may be meaningful for treating patients with colorectal cancer.


Assuntos
Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica , Glucose/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , RNA Longo não Codificante/metabolismo , Idoso , Animais , Carcinogênese/genética , Carcinogênese/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/mortalidade , Feminino , Glicólise/genética , Proteínas de Choque Térmico HSP90/genética , Humanos , Estimativa de Kaplan-Meier , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Prognóstico , Ubiquitinação/genética , Regulação para Cima , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Genes Cells ; 24(8): 524-533, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31273901

RESUMO

The outcome of epigenetic responses to stress depends strictly on genetic background, suggesting that altered phenotypes, when induced, are created by a combination of induced epigenetic factors and pre-existing allelic ones. When individuals with altered phenotypes are selected and subjected to successive breeding, alleles that potentiate epigenetic responses could accumulate in offspring populations. It is reasonable to suppose that many, if not all, of these allelic genes could also be involved in creating new phenotypes under nonstressful conditions. In this review, I discuss the possibility that the accumulation of such alleles in selected individuals with an epigenetic phenotype could give rise to individuals that exhibit the same phenotype even in the absence of stress.


Assuntos
Adaptação Biológica/genética , Evolução Biológica , Epigênese Genética , Alelos , Animais , Cromatina/genética , Cromatina/metabolismo , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Padrões de Herança , Modelos Genéticos , Mutação , Fenótipo , Seleção Genética , Estresse Fisiológico
15.
BMC Cancer ; 19(1): 663, 2019 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-31277598

RESUMO

BACKGROUND: Liver cancer is among top deadly cancers worldwide with a very poor prognosis, and the liver is a vulnerable site for metastases of other cancers. Early diagnosis is crucial for treatment of the predominant liver cancers, namely hepatocellular carcinoma (HCC). Here we developed a novel computational framework for the stage-specific analysis of HCC. METHODS: Using publicly available clinical and RNA-Seq data of cancer samples and controls and the AJCC staging system, we performed a linear modelling analysis of gene expression across all stages and found significant genome-wide changes in the log fold-change of gene expression in cancer samples relative to control. To identify genes that were stage-specific controlling for confounding differential expression in other stages, we developed a set of six pairwise contrasts between the stages and enforced a p-value threshold (< 0.05) for each such contrast. Genes were specific for a stage if they passed all the significance filters for that stage. The monotonicity of gene expression with cancer progression was analyzed with a linear model using the cancer stage as a numeric variable. RESULTS: Our analysis yielded two stage-I specific genes (CA9, WNT7B), two stage-II specific genes (APOBEC3B, FAM186A), ten stage-III specific genes including DLG5, PARI, NCAPG2, GNMT and XRCC2, and 35 stage-IV specific genes including GABRD, PGAM2, PECAM1 and CXCR2P1. Overexpression of DLG5 was found to be tumor-promoting contrary to the cancer literature on this gene. Further, GABRD was found to be signifincantly monotonically upregulated across stages. Our work has revealed 1977 genes with significant monotonic patterns of expression across cancer stages. NDUFA4L2, CRHBP and PIGU were top genes with monotonic changes of expression across cancer stages that could represent promising targets for therapy. Comparison with gene signatures from the BCLC staging system identified two genes, HSP90AB1 and ARHGAP42. Gene set enrichment analysis indicated overrepresented pathways specific to each stage, notably viral infection pathways in HCC initiation. CONCLUSIONS: Our study identified novel significant stage-specific differentially expressed genes which could enhance our understanding of the molecular determinants of hepatocellular carcinoma progression. Our findings could serve as biomarkers that potentially underpin diagnosis as well as pinpoint therapeutic targets.


Assuntos
Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/genética , Detecção Precoce de Câncer/métodos , Perfilação da Expressão Gênica/métodos , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/genética , Transcriptoma , Adulto , Idoso , Biomarcadores Tumorais/genética , Feminino , Proteínas Ativadoras de GTPase/genética , Expressão Gênica , Proteínas de Choque Térmico HSP90/genética , Humanos , Modelos Lineares , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Adulto Jovem
16.
Biochim Biophys Acta Mol Cell Res ; 1866(10): 1544-1555, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31326539

RESUMO

Plasma membrane transporter SLC6A14 transports all neutral and basic amino acids in a Na/Cl - dependent way and it is up-regulated in many types of cancer. Mass spectrometry analysis of overexpressed SLC6A14-associated proteins identified, among others, the presence of cytosolic heat shock proteins (HSPs) and co-chaperones. We detected co-localization of overexpressed and native SLC6A14 with HSP90-beta and HSP70 (HSPA14). Proximity ligation assay confirmed a direct interaction of overexpressed SLC6A14 with both HSPs. Treatment with radicicol and VER155008, specific inhibitors of HSP90 and HSP70, respectively, attenuated these interactions and strongly reduced transporter presence at the cell surface, what resulted from the diminished level of the total transporter protein. Distortion of SLC6A14 proper folding by both HSPs inhibitors directed the transporter towards endoplasmic reticulum-associated degradation pathway, a process reversed by the proteasome inhibitor - bortezomib. As demonstrated in an in vitro ATPase assay of recombinant purified HSP90-beta, the peptides corresponding to C-terminal amino acid sequence following the last transmembrane domain of SLC6A14 affected the HSP90-beta activity. These results indicate that a plasma membrane protein folding can be controlled not only by chaperones in the endoplasmic reticulum, but also those localized in the cytosol.


Assuntos
Sistemas de Transporte de Aminoácidos/metabolismo , Membrana Celular/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Transporte Proteico/fisiologia , Adenosina Trifosfatases/metabolismo , Sistemas de Transporte de Aminoácidos/genética , Biotinilação , Bortezomib/farmacologia , Proteínas de Transporte/efeitos dos fármacos , Proteínas de Transporte/metabolismo , Citosol/metabolismo , Retículo Endoplasmático/metabolismo , Degradação Associada com o Retículo Endoplasmático , Proteínas de Choque Térmico HSP70/antagonistas & inibidores , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Proteínas de Choque Térmico HSP90/genética , Humanos , Células MCF-7 , Macrolídeos/farmacologia , Chaperonas Moleculares/metabolismo , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Dobramento de Proteína , Transporte Proteico/efeitos dos fármacos , Nucleosídeos de Purina/farmacologia
17.
PLoS Biol ; 17(7): e3000358, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31283755

RESUMO

Hsp90 is a conserved molecular chaperone that assists in the folding and function of diverse cellular regulators, with a profound impact on biology, disease, and evolution. As a central hub of protein interaction networks, Hsp90 engages with hundreds of protein-protein interactions within eukaryotic cells. These interactions include client proteins, which physically interact with Hsp90 and depend on the chaperone for stability or function, as well as co-chaperones and partner proteins that modulate chaperone function. Currently, there are no methods to accurately predict Hsp90 interactors and there has been considerable network rewiring over evolutionary time, necessitating experimental approaches to define the Hsp90 network in the species of interest. This is a pressing challenge for fungal pathogens, for which Hsp90 is a key regulator of stress tolerance, drug resistance, and virulence traits. To address this challenge, we applied a novel biochemical fractionation and quantitative proteomic approach to examine alterations to the proteome upon perturbation of Hsp90 in a leading human fungal pathogen, Candida albicans. In parallel, we performed affinity purification coupled to mass spectrometry to define physical interacting partners for Hsp90 and the Hsp90 co-chaperones and identified 164 Hsp90-interacting proteins, including 111 that are specific to the pathogen. We performed the first analysis of the Hsp90 interactome upon antifungal drug stress and demonstrated that Hsp90 stabilizes processing body (P-body) and stress granule proteins that contribute to drug tolerance. We also describe novel roles for Hsp90 in regulating posttranslational modification of the Rvb1-Rvb2-Tah1-Pih1 (R2TP) complex and the formation of protein aggregates in response to thermal stress. This study provides a global view of the Hsp90 interactome in a fungal pathogen, demonstrates the dynamic role of Hsp90 in response to environmental perturbations, and highlights a novel connection between Hsp90 and the regulation of mRNA-associated protein granules.


Assuntos
Candida albicans/metabolismo , Proteínas Fúngicas/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Chaperonas Moleculares/metabolismo , Proteômica/métodos , Candida albicans/genética , Candida albicans/patogenicidade , Candidíase/microbiologia , Proteínas Fúngicas/genética , Redes Reguladoras de Genes , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Choque Térmico HSP90/genética , Humanos , Microscopia Confocal , Chaperonas Moleculares/genética , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Ligação Proteica , Proteoma/genética , Proteoma/metabolismo , Virulência/genética
18.
Nat Commun ; 10(1): 2574, 2019 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-31189925

RESUMO

Complex conformational dynamics are essential for function of the dimeric molecular chaperone heat shock protein 90 (Hsp90), including transient, ATP-biased N-domain dimerization that is necessary to attain ATPase competence. The intrinsic, but weak, ATP hydrolyzing activity of human Hsp90 is markedly enhanced by the co-chaperone Aha1. However, the cellular concentration of Aha1 is substoichiometric relative to Hsp90. Here we report that initial recruitment of this cochaperone to Hsp90 is markedly enhanced by phosphorylation of a highly conserved tyrosine (Y313 in Hsp90α) in the Hsp90 middle domain. Importantly, phosphomimetic mutation of Y313 promotes formation of a transient complex in which both N- and C-domains of Aha1 bind to distinct surfaces of the middle domains of opposing Hsp90 protomers prior to ATP-directed N-domain dimerization. Thus, Y313 represents a phosphorylation-sensitive conformational switch, engaged early after client loading, that affects both local and long-range conformational dynamics to facilitate initial recruitment of Aha1 to Hsp90.


Assuntos
Adenosina Trifosfatases/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Chaperonas Moleculares/metabolismo , Domínios Proteicos/genética , Adenosina Trifosfatases/genética , Ácido Glutâmico/genética , Células HEK293 , Proteínas de Choque Térmico HSP90/genética , Humanos , Modelos Moleculares , Ressonância Magnética Nuclear Biomolecular , Fosforilação/fisiologia , Relação Estrutura-Atividade , Tirosina/genética , Tirosina/metabolismo
19.
J Exp Clin Cancer Res ; 38(1): 204, 2019 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-31101057

RESUMO

BACKGROUND: The treatment for advanced primary hepatocellular carcinoma (HCC) is sorafenib (SORA), while HCC has become increasingly drug resistant with enhanced aerobic glycolysis. The present study aimed to examine the chemotherapeutic effects of a flavonoid proanthocyanidin B2 (PB2) on HCC. METHODS: Five kinds of HCC cell lines and LO2 were used to test the effect of PB2 on aerobic glycolysis. The proliferation, cell cycle, apoptosis and a xenograft mouse model were analyzed. Lentivirus overexpressed pyruvate kinase M2 (PKM2) or sh-PKM2 was used to verify the target of PB2. The detailed mechanism was investigated by immunofluorescence, co-immunoprecipitation, and western blotting. RESULTS: PB2 inhibited the proliferation, induced cell cycle arrest, and triggered apoptosis of HCC cells in vivo and in vitro. PB2 also suppressed glucose uptake and lactate levels via the direct inhibition of the key glycolytic enzyme, PKM2. In addition, PKM2 inhibited the nuclear translocation of PKM2 and co-localization of PKM2/HIF-1α in the nucleus, leading to the inhibition of aerobic glycolysis. Co-treatment with PB2 was also effective in enhancing the chemosensitivity of SORA. CONCLUSIONS: PB2 inhibited the expression and nuclear translocation of PKM2, therefore disrupting the interaction between PKM2/HSP90/HIF-1α, to suppress aerobic glycolysis and proliferation, and trigger apoptosis in HCC via HIF-1α-mediated transcription suppression.


Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Proteínas de Transporte/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Neoplasias Hepáticas/tratamento farmacológico , Proteínas de Membrana/genética , Proantocianidinas/farmacologia , Hormônios Tireóideos/genética , Transporte Ativo do Núcleo Celular/genética , Animais , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/genética , Xenoenxertos , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Camundongos , Sorafenibe/farmacologia
20.
PLoS Pathog ; 15(5): e1007749, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31121004

RESUMO

The regulation of paramyxovirus RNA synthesis by host proteins is poorly understood. Here, we identified a novel regulation mechanism of paramyxovirus RNA synthesis by the Hsp90 co-chaperone R2TP complex. We showed that the R2TP complex interacted with the paramyxovirus polymerase L protein and that silencing of the R2TP complex led to uncontrolled upregulation of mumps virus (MuV) gene transcription but not genome replication. Regulation by the R2TP complex was critical for MuV replication and evasion of host innate immune responses. The R2TP complex also regulated measles virus (MeV) RNA synthesis, but its function was inhibitory and not beneficial to MeV, as MeV evaded host innate immune responses in the absence of the R2TP complex. The identification of the R2TP complex as a critical host factor sheds new light on the regulation of paramyxovirus RNA synthesis.


Assuntos
Proteínas de Choque Térmico HSP90/metabolismo , Interações Hospedeiro-Patógeno , Vírus da Caxumba/genética , Caxumba/genética , RNA Viral/biossíntese , Proteínas Virais/metabolismo , Replicação Viral , Células A549 , Proteínas de Choque Térmico HSP90/genética , Humanos , Caxumba/virologia , Proteínas Virais/genética
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