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1.
Shanghai Kou Qiang Yi Xue ; 29(4): 350-354, 2020 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-33089280

RESUMO

PURPOSE: To investigate the expression and significance of chemokines CCL21, E-selectins and heat shock protein 90 (Hsp90) in periodontal tissues of rats with experimental periodontitis. METHODS: Forty 10-week-old male Wistar rats were significantly randomly divided into 4 groups with 10 rats in each group. Periodontitis models were established in groups A, B and C, and the rest were 10 blank control groups. Rats in group A, B and C were sacrificed at 4, 8 and 12 weeks after basic periodontal treatment, and the periodontal tissues of the first and second molars were taken for CCL21, E-selectins and Hsp90 protein expression detection. SPSS 25.0 software package was used to analyze the data. RESULTS: The levels of periodontal attachment in group A, B and C were higher than those in the control group(P<0.05). The levels of periodontal attachment, CCL21, E-selectins, Hsp90 mRNA and protein expression in periodontal tissues increased first and then decreased(P<0.05). The levels of periodontal attachment, CCL21, E-selectins, Hsp90 mRNA and protein expression in group B and C were significantly higher than those in group A(P<0.05). The levels of periodontal attachment, CCL21, E-selectins, Hsp90 and relative protein expression in periodontal tissues of group C were significantly lower than those of group B(P<0.05). CONCLUSIONS: The expression of CCL21, E-selectins and Hsp90 is up-regulated in periodontitis tissues. With local periodontal treatment, the expression level of CCL21, E-selectins and Hsp90 gradually decreases.


Assuntos
Selectina E , Periodontite , Animais , Quimiocina CCL21/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Masculino , Dente Molar , Periodontite/genética , Periodonto , Ratos , Ratos Wistar
2.
Anticancer Res ; 40(11): 6137-6150, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-33109551

RESUMO

BACKGROUND/AIM: Heat shock protein 90 (HSP90) controls maturation of oncogenic client proteins of cancer cells, and thus we studied the effect of HSP 90 inhibitors on cell survival and survival-related mediators in thyroid carcinoma cells. MATERIALS AND METHODS: Human TPC-1 and SW1736 thyroid carcinoma cells were utilized. Cell viability, cytotoxic activity and apoptosis were estimated using CCK-8 assay, cytotoxicity assay and FACS analysis, respectively. RESULTS: AUY922, BIIB021 and SNX5422 decreased cell viability, and increased cytotoxic activity and the proportion of apoptotic cells. The protein levels of cleaved PARP, cleaved caspase-3, Bax and Bim were elevated, and Bcl2 protein levels were reduced. Knockdown of Bax did not change cell viability, cytotoxic activity, the proportion of apoptotic cells and cleaved caspase-3 protein levels. Meanwhile, knockdown of Bim enhanced cell viability, and diminished cytotoxic activity, the proportion of apoptotic cells and cleaved caspase-3 protein levels. AUY922, BIIB021 and SNX5422 increased the protein levels of phospho-AMPK, and decreased those of phospho-ERK1/2, and total and phospho-AKT. CONCLUSION: AUY922, BIIB021 and SNX5422 induce cytotoxicity by modulating Bim and ERK1/2, AKT and AMPK signaling in thyroid carcinoma cells.


Assuntos
Adenina/análogos & derivados , Apoptose/efeitos dos fármacos , Proteína 11 Semelhante a Bcl-2/metabolismo , Benzamidas/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Indazóis/farmacologia , Isoxazóis/farmacologia , Piridinas/farmacologia , Resorcinóis/farmacologia , Neoplasias da Glândula Tireoide/patologia , Adenina/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Neoplasias da Glândula Tireoide/enzimologia
3.
Nat Commun ; 11(1): 4467, 2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-32948751

RESUMO

Recent studies have reported that upregulation of disulfide-bond A oxidoreductase-like protein (DsbA-L) prevented lipid-induced renal injury in diabetic nephropathy (DN). However, the role and regulation of proximal tubular DsbA-L for renal tubulointerstitial fibrosis (TIF) remains unclear. In current study, we found that a proximal tubules-specific DsbA-L knockout mouse (PT-DsbA-L-KO) attenuated UUO-induced TIF, renal cell apoptosis and inflammation. Mechanistically, the DsbA-L interacted with Hsp90 in mitochondria of BUMPT cells which activated the signaling of Smad3 and p53 to produce connective tissue growth factor (CTGF) and then resulted in accumulation of ECM of BUMPT cells and mouse kidney fibroblasts. In addition, the progression of TIF caused by UUO, ischemic/reperfusion (I/R), aristolochic acid, and repeated acute low-dose cisplatin was also alleviated in PT-DsbA-L-KO mice via the activation of Hsp90 /Smad3 and p53/CTGF axis. Finally, the above molecular changes were verified in the kidney biopsies from patients with obstructive nephropathy (Ob). Together, these results suggest that DsbA-L in proximal tubular cells promotes TIF via activation of the Hsp90 /Smad3 and p53/CTGF axis.


Assuntos
Fibrose/genética , Predisposição Genética para Doença/genética , Glutationa Transferase/genética , Glutationa Transferase/metabolismo , Nefropatias/genética , Idoso , Animais , Apoptose , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Nefropatias Diabéticas , Modelos Animais de Doenças , Feminino , Fibrose/patologia , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Inflamação , Rim/lesões , Nefropatias/patologia , Túbulos Renais Proximais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/metabolismo , Transdução de Sinais , Proteína Smad3/metabolismo , Proteína Supressora de Tumor p53/metabolismo
4.
Ecotoxicol Environ Saf ; 203: 110975, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32678756

RESUMO

Manganese (Mn) produces cholinergic neuronal loss in basal forebrain (BF) region that was related to cognitive dysfunction induced after single and repeated Mn treatment. All processes that generate cholinergic neuronal loss in BF remain to be understood. Mn exposure may produce the reduction of BF cholinergic neurons by increasing amyloid beta (Aß) and phosphorylated Tau (pTau) protein levels, altering heat shock proteins' (HSPs) expression, disrupting proteasome P20S activity and generating oxidative stress. These mechanisms, described to be altered by Mn in regions different than BF, could lead to the memory and learning process alteration produced after Mn exposure. The research performed shows that single and repeated Mn treatment of SN56 cholinergic neurons from BF induces P20S inhibition, increases Aß and pTau protein levels, produces HSP90 and HSP70 proteins expression alteration, and oxidative stress generation, being the last two effects mediated by NRF2 pathway alteration. The increment of Aß and pTau protein levels was mediated by HSPs and proteasome dysfunction. All these mechanisms mediated the cell decline observed after Mn treatment. Our results are relevant because they may assist to reveal the processes leading to the neurotoxicity and cognitive alterations observed after Mn exposure.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Prosencéfalo Basal/efeitos dos fármacos , Neurônios Colinérgicos/efeitos dos fármacos , Poluentes Ambientais/toxicidade , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Manganês/toxicidade , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas tau/metabolismo , Animais , Prosencéfalo Basal/metabolismo , Prosencéfalo Basal/patologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Neurônios Colinérgicos/metabolismo , Neurônios Colinérgicos/patologia , Relação Dose-Resposta a Droga , Poluentes Ambientais/metabolismo , Manganês/metabolismo , Camundongos , Estresse Oxidativo/efeitos dos fármacos
5.
Nucleic Acids Res ; 48(14): 7944-7957, 2020 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-32667666

RESUMO

Circadian clocks are endogenous oscillators that control ∼24-hour physiology and behaviors in virtually all organisms. The circadian oscillator comprises interconnected transcriptional and translational feedback loops, but also requires finely coordinated protein homeostasis including protein degradation and maturation. However, the mechanisms underlying the mammalian clock protein maturation is largely unknown. In this study, we demonstrate that necdin, one of the Prader-Willi syndrome (PWS)-causative genes, is highly expressed in the suprachiasmatic nuclei (SCN), the pacemaker of circadian clocks in mammals. Mice deficient in necdin show abnormal behaviors during an 8-hour advance jet-lag paradigm and disrupted clock gene expression in the liver. By using yeast two hybrid screening, we identified BMAL1, the core component of the circadian clock, and co-chaperone SGT1 as two necdin-interactive proteins. BMAL1 and SGT1 associated with the N-terminal and C-terminal fragments of necdin, respectively. Mechanistically, necdin enables SGT1-HSP90 chaperone machinery to stabilize BMAL1. Depletion of necdin or SGT1/HSP90 leads to degradation of BMAL1 through the ubiquitin-proteasome system, resulting in alterations in both clock gene expression and circadian rhythms. Taken together, our data identify the PWS-associated protein necdin as a novel regulator of the circadian clock, and further emphasize the critical roles of chaperone machinery in circadian clock regulation.


Assuntos
Fatores de Transcrição ARNTL/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Relógios Circadianos , Proteínas de Choque Térmico HSP90/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Proteínas Nucleares/metabolismo , Animais , Relógios Circadianos/genética , Expressão Gênica , Camundongos , Proteínas do Tecido Nervoso/genética , Proteínas Nucleares/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Estabilidade Proteica , Ubiquitina/metabolismo
6.
Ecotoxicol Environ Saf ; 201: 110861, 2020 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-32544748

RESUMO

Marine biota have been co-challenged with ocean warming and mercury (Hg) pollution over many generations because of human activities; however, the molecular mechanisms to explain their combined effects are not well understood. In this study, a marine planktonic copepod Pseudodiaptomus annandalei was acutely exposed to different temperature (22 and 25 °C) and Hg (0 and 118 µg/L) treatments in a 24-h cross-factored experiment. Hg accumulation and its subcellular fractions were determined in the copepods after exposure. The expression of the genes of superoxide dismutase (SOD), glutathione peroxidase (GPx), metallothionein1 (mt1), heat shock protein 70 (hsp70), hsp90, hexokinase (hk), and pyruvate kinase (pk) was also analyzed. Both the Hg treatment alone and the combined exposure of warmer temperature plus Hg pollution remarkably facilitated Hg bioaccumulation in the exposed copepods. Compared with the Hg treatment alone, the combined exposure increased total Hg accumulation and also the amount of Hg stored in the metal-sensitive fractions (MSF), suggesting elevated Hg toxicity in P. annandalei under a warmer environment, given that the MSF is directly related to metal toxicity. The warmer temperature significantly up-regulated the mRNA levels of mt1, hsp70, hsp90, and hk, indicating the copepods suffered from thermal stress. With exposure to Hg, the mRNA level of SOD increased strikingly but the transcript levels of hsp90, hk, and pk decreased significantly, indicating that Hg induced toxic events (e.g., oxidative damage and energy depletion). Particularly, in contrast to the Hg treatment alone, the combined exposure significantly down-regulated the mRNA levels of SOD and GPx but up-regulated the mRNA levels of mt1, hsp70, hsp90, hk, and pk. Collectively, the results of this study indicate that ocean warming will potentially boost Hg toxicity in the marine copepod P. annandalei, which is information that will increase the accuracy of the projections of marine ecosystem responses to the joint effects of climate change stressors and metal pollution on the future ocean.


Assuntos
Copépodes/efeitos dos fármacos , Temperatura Alta , Mercúrio/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Copépodes/genética , Copépodes/metabolismo , Glutationa Peroxidase/genética , Glutationa Peroxidase/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Proteínas de Choque Térmico HSP90/genética , Proteínas de Choque Térmico HSP90/metabolismo , Hexoquinase/genética , Hexoquinase/metabolismo , Mercúrio/farmacocinética , Metalotioneína/genética , Metalotioneína/metabolismo , Piruvato Quinase/genética , Piruvato Quinase/metabolismo , RNA Mensageiro/metabolismo , Superóxido Dismutase/genética , Superóxido Dismutase/metabolismo , Regulação para Cima , Poluentes Químicos da Água/farmacocinética
7.
Prostate ; 80(12): 993-1005, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32559345

RESUMO

BACKGROUND: Androgen deprivation therapy (ADT) is the mainstay of treatment for castration-resistant prostate cancer (CRPC). Unfortunately, although ADT initially prolongs survival, most patients relapse and develop resistance. Clinical failure of these treatments in CRPC highlights the urgent need to develop novel strategies to more effectively block androgen receptor (AR) signaling and target other oncogenic factors responsible for ADT resistance. METHODS: We developed a small-molecule compound LG1836 and investigated the in vitro and in vivo activity of LG1836 against CRPC in cellular and animal models. RESULTS: LG1836 exhibits potent in vitro cytotoxicity in CRPC cells. Mechanistic studies demonstrated that LG1836 inhibits the expression of AR and AR variant 7, partially mediated via proteasome-dependent protein degradation. LG1836 also suppresses survivin expression and effectively induces apoptosis in CRPC cells. Significantly, as a single agent, LG1836 is therapeutically efficacious in suppressing the in vivo growth of CRPC in the subcutaneous and intraosseous models and extends the survival of tumor-bearing mice. CONCLUSIONS: These preclinical studies indicate that LG1836 is a promising lead compound for the treatment of CRPC.


Assuntos
Piperidinas/farmacologia , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Bibliotecas de Moléculas Pequenas/farmacologia , Antagonistas de Receptores de Andrógenos/farmacologia , Animais , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Masculino , Camundongos , Camundongos Nus , Camundongos SCID , Neoplasias de Próstata Resistentes à Castração/patologia , Distribuição Aleatória , Receptores Androgênicos/biossíntese , Receptores Androgênicos/metabolismo , Survivina/antagonistas & inibidores , Survivina/biossíntese , Ubiquitinação , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Life Sci ; 256: 118000, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32585246

RESUMO

AIMS: Hsp90 is regarded as an important therapeutic target in cancer treatment. Client proteins of Hsp90 like Beclin-1, PI3K, and AKT, are associated with tumor development, poor prognosis, and resistance to cancer therapies. This study aims to analyze the role of Gedunin, an Hsp-90 inhibitor, in mediation of crosstalk between apoptosis and autophagy by targeting Beclin-1:Bcl-2 interaction, and ER stress. MAIN METHODS: A549 cells were treated with different concentrations of gedunin, and inhibitory rate was evaluated by MTT assay. Effect of gedunin on generation of reactive oxygen species, mitochondrial membrane potential, and chromatin condensation was studied by staining methods like DCFH-DA, MitoTracker, and DAPI. Expression of EGFR, PIK3CA, AKT, marker genes for apoptosis and autophagy were studied using semi-quantitative RT-PCR. Interaction study of Hsp90:Beclin-1:Bcl-2 was done by immunoprecipitation analysis. Protein expression of autophagy and apoptosis markers along with Grp78, Hsp70, and Hsp90 was analyzed by immunoblotting. KEY FINDINGS: Gedunin exerts cytotoxic effects, causes increase in ROS generation, downregulates mitochondrial membrane potential and induces loss in DNA integrity. mRNA expression analysis revealed that gedunin sensitized A549 cells towards apoptosis by downregulating EGFR, PIK3CA, AKT, and autophagy. Gedunin also inhibited interaction between Hsp90:Beclin-1:Bcl-2, leading to downregulation of autophagy (Beclin-1, Atg5-12 complex, and LC3) and antiapoptotic protein Bcl-2, which may result in ER stress-induced apoptosis. Moreover, Hsp90 inhibition by gedunin did not cause upregulation of Hsp70 expression. SIGNIFICANCE: Gedunin induces apoptosis in lung cancer cells by disrupting Hsp90:Beclin-1:Bcl-2 interaction and autophagy downregulation, thus making gedunin a good drug lead for targeting lung cancer.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Limoninas/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Células A549 , Antineoplásicos Fitogênicos/administração & dosagem , Autofagia/efeitos dos fármacos , Proteína Beclina-1/metabolismo , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Limoninas/administração & dosagem , Neoplasias Pulmonares/patologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo
9.
J Med Chem ; 63(10): 5421-5441, 2020 05 28.
Artigo em Inglês | MEDLINE | ID: mdl-32352777

RESUMO

Herein, a series of HSP90 inhibitor-SN38 conjugates through ester and carbamate linkage in the 20-OH and 10-OH positions of SN38 were developed for improving the tumor-specific penetration and accumulation of SN38 via extracellular HSP90 (eHSP90)-mediated endocytosis. Mechanistic analyses confirmed that these novel conjugates could bind to eHSP90 and be selectively internalized into the tumor cells, which led to prolonged tumor regression in multiple models of cancer. Among all studied conjugates, compound 18b showed excellent in vitro activities, including acceptable HSP90α affinity and potent antitumor activity. Moreover, compound 18b exhibited superior antitumor activity and low toxicity in HCT116 and Capan-1 xenograft models. Pharmacokinetic analyses in HCT116 and Capan-1 xenografts further confirmed that compound 18b treatment could lead to effective cleavage and extended SN38 exposure at tumor sites. All these encouraging data indicate that this compound is a promising new candidate for cancer therapy and merits further chemical and biological evaluation.


Assuntos
Antineoplásicos/administração & dosagem , Antineoplásicos/síntese química , Sistemas de Liberação de Medicamentos/métodos , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Irinotecano/administração & dosagem , Irinotecano/síntese química , Células A549 , Animais , Antineoplásicos/metabolismo , Desenho de Fármacos , Células HCT116 , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Irinotecano/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
10.
Adv Gerontol ; 33(1): 40-45, 2020.
Artigo em Russo | MEDLINE | ID: mdl-32362082

RESUMO

The aim of this work was to examine the content of heat shock protein 90 (HSP90) in fibroblasts of human dermis from the development until deep aging (from 20 weeks of pregnancy until 85 years old), and defining of a role of HSP90 in age-dependent changes in the number of fibroblasts in the dermis. HSP90, proliferating cells nuclear antigen (PCNA) were detected with indirect immunohistochemical technique. Results showed that a portion of fibroblasts with positive staining for HSP90 in the dermis is not changed from 20 weeks of development to 20 years old. Percent of HSP90 positive fibroblasts in dermis is decreased from 21 to 60 years old. From 61 year, the number of HSP90 positive fibroblasts in dermis is increased. Age-related changes in the number of HSP90 positive fibroblasts is not statistically associated with an age-related decrease in a total number and percent of PCNA positive fibroblasts the dermis.


Assuntos
Envelhecimento , Derme/citologia , Fibroblastos/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Adolescente , Adulto , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Recém-Nascido , Pessoa de Meia-Idade , Gravidez , Adulto Jovem
11.
Life Sci ; 254: 117737, 2020 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-32376268

RESUMO

Tumor necrosis factor receptor-associated protein 1 (TRAP1), a molecular chaperone, is a major member of the mitochondrial heat shock protein 90 (Hsp90) family. Studies have shown that TRAP1 can prevent hypoxia-induced damage to cardiomyocytes, maintain cardiomyocytes viability and mitochondrial membrane potential, and protect cardiomyocytes. In addition, it can also protect astrocytes from ischemic damage in vitro. In recent years, there have been many new discoveries in tumors. The abnormal expression of TRAP1 is closely related to the occurrence and development of various tumors. TRAP1 protein seems to be a central regulatory protein, involved in the activation of various oncogenic proteins and signaling pathways, and has a balanced function at tumor transformation and the intersection of different metabolic processes. Targeting its chaperone activity and molecular interactions can destroy the metabolism and survival adaptability of tumor cells, paving the way for the development of highly selective mitochondrial anti-tumor drugs. Moreover, the combination of TRAP1 inhibition and current traditional cancer therapies has shown promising applications. These findings have important implications for the diagnosis and treatment of tumors. Therefore, we reviewed the recently identified functions of the molecular chaperone TRAP1 in cancer development and progression, as well as the discovery and recent advances in selective TRAP1 inhibitors as anticancer drug therapies, opening up new attractive prospects for exploring strategies for targeting TRAP1 as a tumor cell target.


Assuntos
Antineoplásicos/uso terapêutico , Proteínas de Choque Térmico HSP90/efeitos dos fármacos , Chaperonas Moleculares/efeitos dos fármacos , Neoplasias/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Proteínas de Choque Térmico HSP90/fisiologia , Humanos , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/fisiologia , Neoplasias/tratamento farmacológico
12.
Toxicol Appl Pharmacol ; 400: 115075, 2020 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-32470352

RESUMO

NLRP3, one of the HSP-90 clients, has been defined as a critical component of IBD. In a rat model of DSS-induced colitis, we investigated the anti-inflammatory potential of the combined therapy with CP-456773 (CP), an NLRP3 inhibitor, and celastrol (CSR), an NF-κB inhibitor. Our results revealed that the CSR/CP combined therapy (CCCT) attenuated colon shortening, DAI and MDI in addition to improvement of the colonic histological picture. Moreover, the CCCT increased the antioxidant defense machinery of the colonic tissue and decreased MPO activity. Furthermore, the inflammation markers such as TNF-α and IL-6 were downregulated. These effects might be attributed to the inhibitory effect of CSR on the priming step of the NLRP3 inflammasome activation by interrupting NF-κB signalling and inhibition of HSP-90 (at the protein and mRNA levels) along with inhibitory effect of CP on the expression of the NLRP3. These latter effects resulted in decreased tissue expression and activity of the caspase-1 and repressing the subsequent release of the active forms of IL-1ß and IL-18, hence, the pyroptosis process is restrained. Additionally, the CCCT resulted in inducing autophagy by AMPK/mTOR-dependent mechanisms leading to the accumulation of BECN1 protein and a significant decrease in the levels of p62 SQSTM1. The inhibitory effect on HSP-90 in conjunction with induction of autophagy suggest increased autophagic degradation of NLRP3. This novel approach provides a basis for the clinical application of this combination in IBD treatment and might also be promising for the pharmacological intervention of other NLRP3 inflammasome-dependent inflammatory conditions.


Assuntos
Anti-Inflamatórios/farmacologia , Autofagia/efeitos dos fármacos , Colite/tratamento farmacológico , Proteínas de Choque Térmico HSP90/metabolismo , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Sulfonas/farmacologia , Triterpenos/farmacologia , Animais , Anti-Inflamatórios/administração & dosagem , Colite/induzido quimicamente , Colite/metabolismo , Colite/patologia , Citocinas/metabolismo , Sulfato de Dextrana/farmacologia , Modelos Animais de Doenças , Quimioterapia Combinada , Proteínas de Choque Térmico HSP90/sangue , Compostos Heterocíclicos de 4 ou mais Anéis/administração & dosagem , Compostos Heterocíclicos de 4 ou mais Anéis/uso terapêutico , Inflamassomos/efeitos dos fármacos , Inflamassomos/imunologia , Masculino , Ratos Sprague-Dawley , Sulfonas/administração & dosagem , Sulfonas/uso terapêutico , Triterpenos/administração & dosagem , Triterpenos/uso terapêutico
13.
Nucleic Acids Res ; 48(11): 6280-6293, 2020 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-32396196

RESUMO

Although originally identified as the components of the complex aiding the cytosolic chaperonin CCT in the folding of actins and tubulins in the cytosol, prefoldins (PFDs) are emerging as novel regulators influencing gene expression in the nucleus. Work conducted mainly in yeast and animals showed that PFDs act as transcriptional regulators and participate in the nuclear proteostasis. To investigate new functions of PFDs, we performed a co-expression analysis in Arabidopsis thaliana. Results revealed co-expression between PFD and the Sm-like (LSM) genes, which encode the LSM2-8 spliceosome core complex, in this model organism. Here, we show that PFDs interact with and are required to maintain adequate levels of the LSM2-8 complex. Our data indicate that levels of the LSM8 protein, which defines and confers the functional specificity of the complex, are reduced in pfd mutants and in response to the Hsp90 inhibitor geldanamycin. We provide biochemical evidence showing that LSM8 is a client of Hsp90 and that PFD4 mediates the interaction between both proteins. Consistent with our results and with the role of the LSM2-8 complex in splicing through the stabilization of the U6 snRNA, pfd mutants showed reduced levels of this snRNA and altered pre-mRNA splicing patterns.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Chaperonas Moleculares/metabolismo , Complexos Multiproteicos/metabolismo , Proteínas de Ligação a RNA/metabolismo , Spliceossomos/metabolismo , Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Complexos Multiproteicos/química , Mutação , Ligação Proteica , Processamento de RNA , Spliceossomos/química
14.
Adv Exp Med Biol ; 1243: 135-146, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32297216

RESUMO

Molecular chaperones are responsible for maintaining intracellular protein quality control by facilitating the conformational maturation of new proteins as well as the refolding of denatured proteins. While there are several classes of molecular chaperones in the cell, this chapter will focus solely on the small molecule modulation of Hsp90, the 90 kDa heat shock protein. Hsp90 is not only responsible for folding nascent proteins, but it also regulates the triage of numerous client proteins through partnering with the ubiquitin-proteasome pathway. Consequently, Hsp90 plays critical role in maintaining the protein homeostasis (proteostasis) network within the cell and is required for the activation/maturation of more than 300 client protein substrates. Many of the clients that depend upon Hsp90 are overexpressed or mutated during malignant transformation. This often renders the clients thermodynamically unstable and dependent on Hsp90 for stability. This phenomenon results in an oncogenic 'addiction' to the Hsp90 protein folding machinery as Hsp90 maintains onco-client proteins. Furthermore, Hsp90-dependent substrates are associated with all ten hallmarks of cancer, making Hsp90 an attractive target for the development of cancer chemotherapeutics. In fact, 17 small molecule inhibitors of Hsp90 have been developed and clinically evaluated for the treatment of cancer. Unfortunately, most of these molecules have failed for various reasons, necessitating a new approach to modulate the Hsp90 protein folding machine.


Assuntos
Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Animais , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Neoplasias/metabolismo , Dobramento de Proteína/efeitos dos fármacos
15.
Life Sci ; 252: 117676, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32304763

RESUMO

AIMS: Many µ-opioid receptor (MOR)-associated proteins can regulate the MOR signaling pathway. Using a bacterial two-hybrid screen, we found that the C-terminal of the MOR associated with heat shock protein 90 isoform ß (Hsp90ß). Here, we explored the effect of Hsp90ß on MOR signaling transduction and function. MAIN METHODS: The interaction of Hsp90ß with MOR was detected by co-immunoprecipitation and immunofluorescence. The effects of Hsp90ß on MOR signaling induced by opioids were studied in vitro and in vivo. The effects of the Hsp90ß inhibitor 17-N-allylamino-17-demethoxygeldanamycin (17-AAG) on morphine tolerance and dependence were studied via a hot plate test and CPP test. KEY FINDINGS: Hsp90ß, instead of Hsp90α, interacted with the MOR in HEK293 cells and SH-SY5Y cells, and the interaction was augmented after morphine pretreatment. The interaction of Hsp90ß and MOR increased the inhibition of cAMP and decreased PKA activity under opioid treatment. The functional Hsp90ß-MOR complex also promoted the phosphorylation and internalization of the MOR induced by DAMGO in MOR-CHO cells. 17-AAG blocked Hsp90ß-MOR interactions and decreased the effect of Hsp90ß on the MOR signal transduction. In C57BL/6 mice, 17-AAG decreased morphine-induced acute anti-nociception in the hot plate test, with an increase in phosphorylated PKA and phosphorylated JNK and a decrease in phosphorylated CREB and phosphorylated ERK in murine brains. Chronic morphine treatment induced tolerance, and dependence was inhibited by 17-AAG co-administration. SIGNIFICANCE: Hsp90ß is a positive co-regulator of the MOR via the activation of a G-protein-dependent and ß-arrestin-dependent pathway. Hsp90ß has the potential to improve the pharmacologic profile of existing opiates. It is conceivable that in future clinical treatments, the Hsp90ß inhibitor, 17-AAG, could decrease the tolerance and dependence in cancer patients induced by opioids.


Assuntos
Analgésicos Opioides/farmacologia , Benzoquinonas/farmacologia , Proteínas de Choque Térmico HSP90/metabolismo , Lactamas Macrocíclicas/farmacologia , Morfina/farmacologia , Receptores Opioides mu/efeitos dos fármacos , Analgésicos Opioides/administração & dosagem , Animais , Células CHO , Cricetinae , Cricetulus , Tolerância a Medicamentos , Feminino , Células HEK293 , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Morfina/administração & dosagem , Nociceptividade/efeitos dos fármacos , Receptores Opioides mu/metabolismo
16.
Biochim Biophys Acta Mol Cell Res ; 1867(8): 118728, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32343987

RESUMO

Acquired multidrug resistance of cancer cells challenges the chemotherapeutic interventions. To understand the role of molecular chaperone, Hsp90 in drug adapted tumor cells, we have used in vitro drug adapted epidermoid tumor cells as a model system. We found that chemotherapeutic drug adaptation of tumor cells is mediated by induced activities of both Hsp90 and P-glycoprotein (P-gp). Although the high-affinity conformation of Hsp90 has correlated with the enhanced drug efflux activity, we did not observe a direct interaction between P-gp and Hsp90. The enrichment of P-gp and Hsp90 at the cholesterol-rich membrane microdomains is found obligatory for enhanced drug efflux activity. Since inhibition of cholesterol biosynthesis is not interfering with the drug efflux activity, it is presumed that the net cholesterol redistribution mediated by Hsp90 regulates the enhanced drug efflux activity. Our in vitro cholesterol and Hsp90 interaction studies have furthered our presumption that Hsp90 facilitates cholesterol redistribution. The drug adapted cells though exhibited anti-proliferative and anti-tumor effects in response to 17AAG treatment, drug treatment has also enhanced the drug efflux activity. Our findings suggest that drug efflux activity and metastatic potential of tumor cells are independently regulated by Hsp90 by distinct mechanisms. We expose the limitations imposed by Hsp90 inhibitors against multidrug resistant tumor cells.


Assuntos
Antineoplásicos/farmacologia , Colesterol/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/metabolismo , Membro 1 da Subfamília B de Cassetes de Ligação de ATP , Indutores da Angiogênese , Animais , Apoptose/efeitos dos fármacos , Benzoquinonas/farmacologia , Linhagem Celular Tumoral , Progressão da Doença , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Proteínas de Choque Térmico HSP90/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/genética , Humanos , Lactamas Macrocíclicas/farmacologia , Masculino , Camundongos Nus , Transcriptoma , Ensaios Antitumorais Modelo de Xenoenxerto
17.
Cancer Invest ; 38(5): 310-328, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32274949

RESUMO

Heat shock protein 90 (HSP90), a highly and unique chaperone, presents as a double-edged sword. It plays an essential role in many physiological and pathological processes, including tumor development. The current review highlights a recent understanding of the roles of HSP90 in molecular mechanisms underlying cancer survival and progression. HSP90 and its client proteins through the regulation of oncoproteins including signaling proteins, receptors, and transcriptional factors involved in tumorigenesis. It also has potential clinical application as diagnostic and prognostic biomarkers for assessing cancer progression. In this way, using HSP90 to develop new anticancer therapeutic agents including HSP90 inhibitors, anti-HSP90 antibody, and HSP90-based vaccines has been promising.


Assuntos
Proteínas de Choque Térmico HSP90/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/metabolismo , Carcinogênese/metabolismo , Carcinogênese/patologia , Progressão da Doença , Humanos , Terapia de Alvo Molecular/métodos , Neoplasias/tratamento farmacológico
18.
Sheng Li Xue Bao ; 72(2): 157-166, 2020 Apr 25.
Artigo em Chinês | MEDLINE | ID: mdl-32328609

RESUMO

This study was aimed to investigate the regulatory mechanism of heat shock protein 90 (Hsp90) on transcription factor EB (TFEB) during autophagy in liver cancer cells. Human hepatocellular carcinoma cell line HepG2 was treated with Hsp90 N- and C-terminal inhibitors (STA9090 and Novobiocin), respectively. Western blot and RT-PCR were used to detect the expression levels of TFEB and autophagy-related proteins. Chromatin immunoprecipitation (ChIP) assay was used to observe the ability of Hsp90α binding to the TFEB proximal promoter region. The double-luciferase gene reporter experiment was used to determine the activity of TFEB promoter. The results showed that hypoxia induced up-regulation of TFEB protein and mRNA expression levels in the HepG2 cells. The protein expression levels of TFEB, LC3 and P62 were down-regulated significantly by either STA9090 or Novobiocin, under both normoxic and hypoxic conditions. Transfection of Hsp90α-overexpressing plasmids up-regulated TFEB protein levels in either wild-type or Hsp90α knockout HepG2 cells. Hsp90 bound to the TFEB proximal promoter region and was involved in regulating TFEB transcriptional process. Whereas both STA9090 and Novobiocin inhibited Hsp90 to bind to the TFEB proximal promoter region, and decreased the activity of TFEB promoter. These results suggest that Hsp90 promotes TFEB transcription in human hepatocellular carcinoma cells by binding to the proximal promoter region, thereby up-regulating the expression levels of autophagy-related proteins.


Assuntos
Autofagia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Carcinoma Hepatocelular/patologia , Proteínas de Choque Térmico HSP90/metabolismo , Neoplasias Hepáticas/patologia , Carcinoma Hepatocelular/metabolismo , Células Hep G2 , Humanos , Neoplasias Hepáticas/metabolismo , Regiões Promotoras Genéticas
19.
mBio ; 11(2)2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32317319

RESUMO

Protein homeostasis is critical for proliferation and viability of all organisms. For Candida albicans, protein homeostasis also modulates the transition between yeast and filamentous forms, which is critical for virulence. A key regulator of morphogenesis is the molecular chaperone Hsp90, which mediates proteostasis under physiological and stress conditions. Hsp90 regulates morphogenesis by repressing cyclic AMP-protein kinase A (cAMP-PKA) signaling, such that inhibition of Hsp90 causes filamentation in the absence of an inducing cue. We explored the effect of perturbation of another facet of protein homeostasis and discovered that morphogenesis is also regulated by the proteasome, a large 33-subunit protein complex consisting of a 20S catalytic core and two 19S regulatory particles, which controls degradation of intracellular proteins. We identified a conserved role of the proteasome in morphogenesis as pharmacological inhibition of the proteasome induced filamentation of C. albicans and the related species Candida dubliniensis, Candida tropicalis, Candida krusei, and Candida parapsilosis For C. albicans, genetic depletion of any of 29 subunits of the 19S or 20S particle induced filamentation. Filaments induced by inhibition of either the proteasome or Hsp90 have shared structural characteristics, such as aberrant nuclear content, and shared genetic dependencies, such as intact cAMP-PKA signaling. Consistent with a functional connection between these facets of protein homeostasis that modulate morphogenesis, we observed that proteasome inhibition results in an accumulation of ubiquitinated proteins that overwhelm Hsp90 function, relieving Hsp90-mediated repression of morphogenesis. Together, our findings provide a mechanism whereby interconnected facets of proteostasis regulate C. albicans morphogenesis.IMPORTANCE Fungi cause life-threatening infections and pose a serious threat to human health as there are very few effective antifungal drugs. Candida albicans is a major human fungal pathogen and cause of morbidity and mortality in immunocompromised individuals. A key trait that enables C. albicans virulence is its ability to transition between yeast and filamentous forms. Understanding the mechanisms regulating this virulence trait can facilitate the development of much-needed, novel therapeutic strategies. A key regulator of morphogenesis is the molecular chaperone Hsp90, which is crucial for proteostasis. Here, we expanded our understanding of how proteostasis regulates fungal morphogenesis and identified the proteasome as a repressor of filamentation in C. albicans and related species. Our work suggests that proteasome inhibition overwhelms Hsp90 function, thereby inducing morphogenesis. This work provides a foundation for understanding the role of the proteasome in fungal virulence and offers potential for targeting the proteasome to disarm fungal pathogens.


Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Proteínas Fúngicas/metabolismo , Fungos/fisiologia , Proteínas de Choque Térmico HSP90/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Transdução de Sinais , AMP Cíclico/metabolismo , Fungos/citologia
20.
Chemistry ; 26(43): 9459-9465, 2020 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-32167602

RESUMO

Protein folding quality control in cells requires the activity of a class of proteins known as molecular chaperones. Heat shock protein-90 (Hsp90), a multidomain ATP driven molecular machine, is a prime representative of this family of proteins. Interactions between Hsp90, its co-chaperones, and client proteins have been shown to be important in facilitating the correct folding and activation of clients. Hsp90 levels and functions are elevated in tumor cells. Here, we computationally predict the regions on the native structures of clients c-Abl, c-Src, Cdk4, B-Raf and Glucocorticoid Receptor, that have the highest probability of undergoing local unfolding, despite being ordered in their native structures. Such regions represent potential ideal interaction points with the Hsp90-system. We synthesize mimics spanning these regions and confirm their interaction with partners of the Hsp90 complex (Hsp90, Cdc37 and Aha1) by Nuclear Magnetic Resonance (NMR). Designed mimics selectively disrupt the association of their respective clients with the Hsp90 machinery, leaving unrelated clients unperturbed and causing apoptosis in cancer cells. Overall, selective targeting of Hsp90 protein-protein interactions is achieved without causing indiscriminate degradation of all clients, setting the stage for the development of therapeutics based on specific chaperone:client perturbation.


Assuntos
Carcinógenos/química , Proteínas de Ciclo Celular/química , Chaperoninas/química , Proteínas de Choque Térmico HSP90/química , Chaperonas Moleculares/química , Carcinógenos/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Dobramento de Proteína
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