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1.
Zhonghua Zhong Liu Za Zhi ; 41(12): 918-922, 2019 Dec 23.
Artigo em Chinês | MEDLINE | ID: mdl-31874549

RESUMO

Objective: To investigate the expression level of antisense transcript of pseudogene, general transcription factor Ⅱi psedugen23 (GTF2IP23), in breast cancer and its effect on the host gene general transcription factor Ⅱi (GTF2I). Methods: The expressions of GTF2IP23 and GTF2I were detected in 40 cases of invasive breast cancer tumors and their counterparts by using quantitative real-time polymerase chain reaction (qRT-PCR). The effects of GTF2IP23 on the expression of GTF2I gene and cell proliferation and migration were analyzed by overexpression of GTF2IP23 in breast cancer cells. Results: The expression of GTF2IP23 mRNA in breast cancer tissues was significantly higher than that in adjacent tissues (P<0.001), while the expression of GTF2I mRNA was significantly lower than that in adjacent tissues (P=0.007). The expression of GTF2IP23 was negatively correlated with GTF2I (r=-0.335, P=0.025). The expression of GTF2IP23 in breast cancer cells was significantly higher than in normal breast cells (P<0.01), while GTF2I expression in breast cancer cells was significantly lower than that in normal breast cells (P<0.01). Overexpression of GTF2IP23 in ZR-75-30 cells significantly reduced the expression of GTF2I (P=0.034) and enhanced cell proliferation (P=0.017) and migration (P=0.026) capacity. Conclusions: GTF2IP23 is distinctly upregulated in breast cancer, it inhibits the expression of real gene GTF2I and promotes the proliferation of breast cancer cells.


Assuntos
Neoplasias da Mama/sangue , Proteínas Musculares/metabolismo , Proteínas Nucleares/metabolismo , Transativadores/metabolismo , Fatores de Transcrição TFII/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Proteínas Musculares/genética , Proteínas Nucleares/genética , Reação em Cadeia da Polimerase em Tempo Real , Transativadores/genética , Fatores de Transcrição TFII/metabolismo
2.
Zhonghua Gan Zang Bing Za Zhi ; 27(9): 693-697, 2019 Sep 20.
Artigo em Chinês | MEDLINE | ID: mdl-31594094

RESUMO

Objective: To investigate the effect and mechanism of XTP4 gene in apoptotic hepatoblastoma HepG2 cell line. Methods: HepG2 cells were transiently transfected with small interfering RNA of XTP4 genes, plasmid pcDNA3.1/myc-His(-) A-XTP4, and hepatitis B virus X protein transactivated x gene 4 (HBX protein trans-activate gene4, XTP4) and their respective negative controls. After 48h, the overexpression and interference expression condition of XTP4 in HepG2 cells were detected by Western blot. HepG2 cells apoptosis was detected by flow cytometry. The expression levels of apoptosis-related proteins P53, Bcl-2, Bax and Caspase-3 in HepG2 cells were detected by Western blot, and Bcl-2/Bax ratio was calculated. The chemiluminescence assay was used to detect activity of caspase-3 in HepG2 cells. The measured data were presented as (x ± s), and independent sample t-test was used for comparison between the two groups. Results: HepG2 cells had successfully achieved the overexpression and interference expression of XTP4 protein. Compared with the control group, the overexpression of XTP4 in HepG2 cells had significantly decreased the number of apoptotic cells (P < 0.05), and increased Bcl-2/Bax (P < 0.05) ratio, but decreased the expression of P53 protein (P < 0.05). The protein expression of Caspase-3 and activity of caspase-3 was decreased (P < 0.05). However, interference with XTP4 expression in HepG2 cells had significantly increased the number of apoptotic cells (P < 0.05) and decreased Bcl-2/Bax (P < 0.05) ratio, but increased the expression of P53 protein (P < 0.05). The protein expression of Caspase-3 and activity of caspase-3 was increased (P<0.05). Conclusion: In HepG2 apoptosis XTP4 has inhibitory effect, and its effect on inhibiting HepG2 apoptosis may be achieved by regulating the Bcl-2/Bax ratio, and the P53 protein may be involved.


Assuntos
Apoptose , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/patologia , Transativadores/metabolismo , Caspase 3/metabolismo , Células Hep G2 , Humanos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Transfecção , Proteína Supressora de Tumor p53/metabolismo , Proteína X Associada a bcl-2/metabolismo
3.
Cancer Sci ; 110(10): 3110-3121, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31385398

RESUMO

Hepatocellular carcinoma (HCC) is one of the most common malignancies and the fourth leading cause of cancer-related death worldwide. Our previous study showed that EYA4 functioned by suppressing growth of HCC tumor cells, but its molecular mechanism is still not elucidated. Based on the results of gene microassay, EYA4 was inversely correlated with MYCBP and was verified in human HCC tissues by immunohistochemistry and western blot. Overexpressed and KO EYA4 in human HCC cell lines confirmed the negative correlation between EYA4 and MYCBP by qRT-PCR and western blot. Transfected siRNA of MYCBP in EYA4 overexpressed cells and overexpressed MYCBP in EYA4 KO cells could efficiently rescue the proliferation and G2/M arrest effects of EYA4 on HCC cells. Mechanistically, armed with serine/threonine-specific protein phosphatase activity, EYA4 reduced nuclear translocation of ß-catenin by dephosphorylating ß-catenin at Ser552, thereby suppressing the transcription of MYCBP which was induced by ß-catenin/LEF1 binding to the promoter of MYCBP. Clinically, HCC patients with highly expressed EYA4 and poorly expressed MYCBP had significantly longer disease-free survival and overall survival than HCC patients with poorly expressed EYA4 and highly expressed MYCBP. In conclusion, EYA4 suppressed HCC tumor cell growth by repressing MYCBP by dephosphorylating ß-catenin S552. EYA4 combined with MYCBP could be potential prognostic biomarkers in HCC.


Assuntos
Carcinoma Hepatocelular/metabolismo , Proteínas de Ligação a DNA/genética , Neoplasias Hepáticas/metabolismo , Transativadores/genética , Transativadores/metabolismo , Fatores de Transcrição/genética , beta Catenina/metabolismo , Adulto , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Núcleo Celular/metabolismo , Proliferação de Células , Proteínas de Ligação a DNA/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/genética , Masculino , Pessoa de Meia-Idade , Fosforilação , Prognóstico , Serina/metabolismo , Análise de Sobrevida , Fatores de Transcrição/metabolismo , Transcrição Genética , beta Catenina/química
4.
J Microbiol Biotechnol ; 29(8): 1299-1309, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31387340

RESUMO

As an opportunistic bacterial pathogen, Pseudomonas aeruginosa PAO1 contains two phenazineproducing gene operons, phzA1B1C1D1E1F1G1 (phz1) and phzA2B2C2D2E2F2G2 (phz2), each of which is independently capable of encoding all enzymes for biosynthesizing phenazines, including phenazine-1-carboxylic acid and its derivatives. Other previous study reported that the RpoS-deficient mutant SS24 overproduced pyocyanin, a derivative of phenazine-1- carboxylic acid. However, it is not known how RpoS mediates the expression of two phz operons and regulates pyocyanin biosynthesis in detail. In this study, with deletion of the rpoS gene in the PAΔphz1 mutant and the PAΔphz2 mutant respectively, we demonstrated that RpoS exerted opposite regulatory roles on the expression of the phz1and phz2 operons. We also confirmed that the phz1 operon played a critical role and especially biosynthesized much more phenazines than the phz2 operon when the rpoS gene was knocked out in P. aeruginosa. By constructing the translational reporter fusion vector lasR'-'lacZ and the chromosomal fusion mutant PAΔlasR::lacZ, we verified that RpoS deficiency caused increased expression of lasR, a transcription regulator gene in a first quorum sensing system (las) that activates overexpression of the phz1 operon, suggesting that in the absence of RpoS, LasR might act as an intermediate in overproduction of phenazine biosynthesis mediated by the phz1 operon in P. aeruginosa.


Assuntos
Proteínas de Bactérias/metabolismo , Fenazinas/metabolismo , Pseudomonas aeruginosa/metabolismo , Fator sigma/metabolismo , Transativadores/metabolismo , Proteínas de Bactérias/genética , Regulação Bacteriana da Expressão Gênica , Fusão Gênica , Óperon , Pseudomonas aeruginosa/genética , Piocianina/biossíntese , Fator sigma/genética
5.
Genes Dev ; 33(19-20): 1441-1455, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31467088

RESUMO

Rapid perturbation of protein function permits the ability to define primary molecular responses while avoiding downstream cumulative effects of protein dysregulation. The auxin-inducible degron (AID) system was developed as a tool to achieve rapid and inducible protein degradation in nonplant systems. However, tagging proteins at their endogenous loci results in chronic auxin-independent degradation by the proteasome. To correct this deficiency, we expressed the auxin response transcription factor (ARF) in an improved inducible degron system. ARF is absent from previously engineered AID systems but is a critical component of native auxin signaling. In plants, ARF directly interacts with AID in the absence of auxin, and we found that expression of the ARF PB1 (Phox and Bem1) domain suppresses constitutive degradation of AID-tagged proteins. Moreover, the rate of auxin-induced AID degradation is substantially faster in the ARF-AID system. To test the ARF-AID system in a quantitative and sensitive manner, we measured genome-wide changes in nascent transcription after rapidly depleting the ZNF143 transcription factor. Transcriptional profiling indicates that ZNF143 activates transcription in cis and regulates promoter-proximal paused RNA polymerase density. Rapidly inducible degradation systems that preserve the target protein's native expression levels and patterns will revolutionize the study of biological systems by enabling specific and temporally defined protein dysregulation.


Assuntos
Técnicas Genéticas , Proteínas/metabolismo , Proteólise , Linhagem Celular , Inibidores de Cisteína Proteinase/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Ácidos Indolacéticos/farmacologia , Leupeptinas/farmacologia , Células MCF-7 , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteólise/efeitos dos fármacos , Transativadores/genética , Transativadores/metabolismo
6.
Biochim Biophys Acta Gene Regul Mech ; 1862(8): 759-770, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31269460

RESUMO

The majority of melanomas carry an oncogenic BRAF mutation (BRAFV600E), which results in constitutive kinase activity driving melanoma proliferation. While inhibitors of BRAFV600E (BRAFi) effectively lead to rapid tumor shrinkage, most patients treated with BRAFi develop acquired resistance. Identification of factors as regulators of melanoma growth and as potential sources of resistance is thus crucial for the design of improved therapies to treat advanced melanoma with more durable responses. Here, we show that KH-type splicing regulatory protein (KSRP) is critical for proliferation of melanoma cells without and with acquired resistance to vemurafenib. Silencing KSRP reduces cell proliferation and augments the growth suppressive effects of vemurafenib. We identify killin (KLLN), a p53-regulated DNA replication inhibitor, as a downstream effector of growth inhibition by KSRP silencing and demonstrate that KSRP promotes decay of KLLN mRNA through an RNA-protein interaction. Using heterologous mRNA reporters, we show that a U-rich element within the 3' untranslated region of KLLN is responsible for KSRP-dependent mRNA decay. These findings implicate that KSRP is an important regulator of melanoma cell growth in part through controlling KLLN mRNA stability.


Assuntos
Resistencia a Medicamentos Antineoplásicos , Melanoma/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Transativadores/genética , Transativadores/metabolismo , Proteínas Supressoras de Tumor/genética , Vemurafenib/uso terapêutico , Regiões 3' não Traduzidas , Animais , Linhagem Celular Tumoral , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Humanos , Melanoma/genética , Camundongos , Estabilidade de RNA , RNA Mensageiro/química , Proteínas Supressoras de Tumor/química , Regulação para Cima
7.
Nat Commun ; 10(1): 2999, 2019 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-31278262

RESUMO

The different genome-wide distributions of tri-methylation at H3K36 (H3K36me3) in various species suggest diverse mechanisms for H3K36me3 establishment during evolution. Here, we show that the transcription factor OsSUF4 recognizes a specific 7-bp DNA element, broadly distributes throughout the rice genome, and recruits the H3K36 methyltransferase SDG725 to target a set of genes including the key florigen genes RFT1 and Hd3a to promote flowering in rice. Biochemical and structural analyses indicate that several positive residues within the zinc finger domain are vital for OsSUF4 function in planta. Our results reveal a regulatory mechanism contributing to H3K36me3 distribution in plants.


Assuntos
Histona-Lisina N-Metiltransferase/metabolismo , Histonas/genética , Oryza/fisiologia , Proteínas de Plantas/metabolismo , Transativadores/metabolismo , Metilação de DNA/fisiologia , Flores/crescimento & desenvolvimento , Regulação da Expressão Gênica de Plantas/fisiologia , Regiões Promotoras Genéticas/genética , Ligação Proteica/genética
8.
DNA Cell Biol ; 38(8): 840-848, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31314587

RESUMO

microRNAs are a class of noncoding RNAs that play important roles in cancer progression. microRNA-183-3p (miR-183-3p) is a novel microRNA that is dysregulated in many kinds of cancers. Our previous studies found high expression and oncologic role of high-mobility group nucleosome binding domain 5 (HMGN5) in prostate cancer. In this study, we found that miR-183-3p was downregulated in prostate cancer cells and primary tissues compared with normal controls. In addition, miR-183-3p expression was negatively correlated with HMGN5 expression. On the basis of bioinformatics predication and quantitative polymerase chain reaction and Western blot verification, it is demonstrated that miR-183-3p regulated HMGN5 expression. Luciferase reporter assay confirmed that miR-183-3p directly targeted the 3'-untranslated region of HMGN5. Interestingly, cell proliferation and migration inhibition and apoptosis induction were also observed in miR-183-3p transfected human prostate cancer VCap and C4-2 cells. Moreover, overexpression of HMGN5 significantly reversed the inhibitory effect on cell proliferation and migration and promoted effect on cell apoptosis by miR-183-3p. Our data suggest that dysfunction of miR-183-3p-HMGN5 axis plays an oncogenic role and can be a therapeutic target for prostate cancer.


Assuntos
Proteínas HMGN/genética , MicroRNAs/genética , Neoplasias da Próstata/genética , Transativadores/genética , Regiões 3' não Traduzidas , Idoso , Apoptose/genética , Estudos de Casos e Controles , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Proteínas HMGN/metabolismo , Humanos , Masculino , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Neoplasias da Próstata/patologia , Transativadores/metabolismo
9.
Medicine (Baltimore) ; 98(26): e15954, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31261501

RESUMO

Early diagnosis of pancreatic cancer (PC) is based on endoscopic ultrasound (EUS). However, EUS is invasive and requires a high level of technical skill. Recently, liquid biopsies have achieved the same sensitivity and specificity for the diagnosis of numerous pathologies, including cancer. Insulin-promoting factor 1 (PDX1) and Msh-homeobox 2 (MSX2), 2 homeotic genes, have been confirmed to be related to pancreatic oncogenesis.The aim of this study is to establish the diagnostic utility of circulating serum levels of MSX2 and PDX1 expression in patients with PC.A prospective study was conducted from January 2014 to February 2017. Patients with a suspected diagnosis of PC who underwent fine needle aspiration biopsy guided by EUS (EUS-FNA) were included in the study, in addition to non-PC control subjects. Both tissue and blood serum samples were submitted to histopathological analysis and measurement of PDX1 and MSX2 gene expression by means of qRT-PCR.Patients were divided into non-PC, malignant pathology (MP), or benign pathology (BP) groups. Significant differences in both MSX2 [2.05 (1.66-4.60) vs 0.83 (0.49-1.60), P = .006] and PDX1 [2.59 (1.28-10.12) vs 1.02 (0.81-1.17), P = .036] gene expression were found in blood samples of PC compared with non-PC subjects. We also observed a significant increase in MSX2 transcripts in tissue biopsy samples of patients diagnosed with MP compared with those with BP [1.98 (1.44-4.61) and 0.66 (0.45-1.54), respectively, P = .012]. The ROC curves indicate a sensitivity and specificity of 80% for PDX1 and 86% for MSX2.Gene expression of MSX2 in tissue samples obtained by EUS-FNA and serum expression of MSX2 and PDX1 were higher in patients with PC.


Assuntos
Proteínas de Homeodomínio/metabolismo , Neoplasias Pancreáticas/metabolismo , Transativadores/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/metabolismo , Biópsia por Agulha Fina , Estudos de Casos e Controles , Aspiração por Agulha Fina Guiada por Ultrassom Endoscópico , Feminino , Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Pâncreas/metabolismo , Pâncreas/patologia , Neoplasias Pancreáticas/patologia , Estudos Prospectivos , Sensibilidade e Especificidade
10.
Nat Commun ; 10(1): 3049, 2019 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-31296872

RESUMO

The transcription factor p63 is a master regulator of ectoderm development. Although previous studies show that p63 triggers epidermal differentiation in vitro, the roles of p63 in developing embryos remain poorly understood. Here, we use zebrafish embryos to analyze in vivo how p63 regulates gene expression during development. We generate tp63-knock-out mutants that recapitulate human phenotypes and show down-regulated epidermal gene expression. Following p63-binding dynamics, we find two distinct functions clearly separated in space and time. During early development, p63 binds enhancers associated to neural genes, limiting Sox3 binding and reducing neural gene expression. Indeed, we show that p63 and Sox3 are co-expressed in the neural plate border. On the other hand, p63 acts as a pioneer factor by binding non-accessible chromatin at epidermal enhancers, promoting their opening and epidermal gene expression in later developmental stages. Therefore, our results suggest that p63 regulates cell fate decisions during vertebrate ectoderm specification.


Assuntos
Ectoderma/embriologia , Desenvolvimento Embrionário/genética , Regulação da Expressão Gênica no Desenvolvimento , Placa Neural/embriologia , Fosfoproteínas/metabolismo , Transativadores/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Animais , Animais Geneticamente Modificados , Sistemas CRISPR-Cas/genética , Diferenciação Celular/genética , Cromatina/metabolismo , Regulação para Baixo , Ectoderma/metabolismo , Embrião não Mamífero , Elementos Facilitadores Genéticos/genética , Epiderme/embriologia , Epiderme/metabolismo , Técnicas de Inativação de Genes , Modelos Animais , Placa Neural/metabolismo , Fosfoproteínas/genética , Ligação Proteica/genética , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Transativadores/genética , Peixe-Zebra/embriologia , Proteínas de Peixe-Zebra/genética
11.
Chemosphere ; 234: 338-345, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31228835

RESUMO

Silkworm (Bombyx mori) is one of the most important economic insects in the world, while pesticides impact its economic benefits. Tebuconazole is a fungicide that has been frequently detected in agriculture systems at concentrations that affect endocrine function in organisms. In the present study, silkworm larvae at different instar stages were exposed to tebuconazole, respectively. Cocoon weight, cocoon shell weight and cocoon shell rate were significantly decreased by 6.8%, 11.8% and 4.4% respectively, after exposure to 0.40 mg/L tebuconazole at 2nd -3rd instar stage. Vacuolization was found in the exposure silkworm under histopathological study at all stages exposures, indicating potential damage to silk gland. Downregulation of genes transcription (Fibh, Fibl, P25, Ser2, Ser3) involved with protein synthesis in the silk gland were further observed, and the results showed significant decreasing in mRNA expression among the tebuconazole treatments. Ecdysteroid levels in silkworm were changed with pronounced decreases after exposed to tebuconazole. In contrast, exposure to tebuconazole significantly increased juvenile hormone 1 concentrations and the maximum increasing fold of juvenile hormone 1 was up to 3.73 which was observed at stage I exposure. In addition, co-exposure to 2 and 10 mg/L forskolin able to mitigate tebuconazole-induced downregulate of mRNA expression of Sgf1 in the present study, indicating the potential mechanism of tebuconazole-induced chronic toxicity in silkworm may relative to PI3K/AKT/TORC1/Sgf1 pathway.


Assuntos
Bombyx/efeitos dos fármacos , Seda/genética , Triazóis/toxicidade , Animais , Regulação para Baixo , Ecdisteroides/metabolismo , Expressão Gênica/efeitos dos fármacos , Proteínas de Insetos/genética , Larva/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Biossíntese de Proteínas , Sesquiterpenos/metabolismo , Transativadores/metabolismo
12.
Nat Commun ; 10(1): 2723, 2019 06 20.
Artigo em Inglês | MEDLINE | ID: mdl-31222014

RESUMO

Non-genetic drug resistance is increasingly recognised in various cancers. Molecular insights into this process are lacking and it is unknown whether stable non-genetic resistance can be overcome. Using single cell RNA-sequencing of paired drug naïve and resistant AML patient samples and cellular barcoding in a unique mouse model of non-genetic resistance, here we demonstrate that transcriptional plasticity drives stable epigenetic resistance. With a CRISPR-Cas9 screen we identify regulators of enhancer function as important modulators of the resistant cell state. We show that inhibition of Lsd1 (Kdm1a) is able to overcome stable epigenetic resistance by facilitating the binding of the pioneer factor, Pu.1 and cofactor, Irf8, to nucleate new enhancers that regulate the expression of key survival genes. This enhancer switching results in the re-distribution of transcriptional co-activators, including Brd4, and provides the opportunity to disable their activity and overcome epigenetic resistance. Together these findings highlight key principles to help counteract non-genetic drug resistance.


Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Leucemia Mieloide Aguda/tratamento farmacológico , Transativadores/antagonistas & inibidores , Animais , Antineoplásicos/uso terapêutico , Medula Óssea/patologia , Sistemas CRISPR-Cas/genética , Linhagem Celular Tumoral , Epigênese Genética/efeitos dos fármacos , Feminino , Células HEK293 , Humanos , Estimativa de Kaplan-Meier , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/mortalidade , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Endogâmicos C57BL , Análise de Sequência de RNA , Análise de Célula Única , Transativadores/genética , Transativadores/metabolismo , Transcrição Genética/efeitos dos fármacos , Resultado do Tratamento , Ensaios Antitumorais Modelo de Xenoenxerto
13.
J Biomed Sci ; 26(1): 44, 2019 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-31170980

RESUMO

BACKGROUND: Our previous report suggested that centrosomal P4.1-associated protein (CPAP) is required for Hepatitis B virus (HBV) encoded non-structure protein X (HBx)-mediated nuclear factor kappa light chain enhancer of activated B cells (NF-κB) activation. CPAP is overexpressed in HBV-associated hepatocellular carcinoma (HCC); however, the interaction between CPAP and HBx in HBV-HCC remains unclear. METHODS: The mRNA expression of CPAP and HBx was analyzed by quantitative-PCR (Q-PCR). NF-κB transcriptional activity and CPAP promoter activity were determined using a reporter assay in Huh7 and Hep3B cells. Immunoprecipitation (IP) and in situ proximal ligation assay (PLA) were performed to detect the interaction between CPAP and HBx. Chromatin-IP was used to detect the association of cAMP response element binding protein (CREB) and HBx with the CPAP promoter. Cell proliferation was measured using cell counting kit CCK-8, Bromodeoxyuridine (5-bromo-2'-deoxyuridine, BrdU) incorporation, and clonogenic assays. The tumorigenic effects of CPAP were determined using xenograft animal models. RESULTS: HBx can transcriptionally up-regulate CPAP via interacting with CREB. Overexpressed CPAP directly interacted with HBx to promote HBx-mediated cell proliferation and migration; SUMO modification of CPAP was involved in interacting with HBx. Knocked-down expression of CPAP decreased the HBx-mediated tumorigenic effects, including cytokines secretion. Interestingly, overexpressed CPAP maintained the HBx protein stability in an NF-κB-dependent manner; and the expression levels of CPAP and HBx were positively correlated with the activation status of NF-κB in HCC. Increased expression of CPAP and CREB mRNAs existed in the high-risk group with a lower survival rate in HBV-HCC. CONCLUSION: The interaction between CPAP and HBx can provide a microenvironment to facilitate HCC development via enhancing NF-κB activation, inflammatory cytokine production, and cancer malignancies. This study not only sheds light on the role of CPAP in HBV-associated HCC, but also provides CPAP as a potential target for blocking the hyper-activated NF-κB in HCC.


Assuntos
Carcinogênese/genética , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Proteínas Associadas aos Microtúbulos/farmacologia , Transativadores/metabolismo , Animais , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID
14.
Nat Commun ; 10(1): 2458, 2019 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-31165730

RESUMO

During stress, prompt export of stress-inducible transcripts is critical for cell survival. Here, we characterize a function of the SAGA (Spt-Ada-Gcn5 acetyltransferase) deubiquitylating module (DUBm) in monitoring messenger ribonucleoprotein (mRNP) biogenesis to regulate non-canonical mRNA export of stress-inducible transcripts. Our genetic and biochemical analyses suggest that there is a functional relationship between Sgf73p of DUBm and the essential mRNA export factor, Yra1p. Under physiological conditions, Sgf73p is critical for the proper chromatin localization and RNA binding of Yra1p, while also quality controlling the biogenesis of mRNPs in conjunction with the nuclear exosome exonuclease, Rrp6p. Under environmental stress, when immediate transport of stress-inducible transcripts is imperative, Sgf73p facilitates the bypass of canonical surveillance and promotes the timely export of necessary transcripts. Overall, our results show that the Sgf73p-mediated plasticity of gene expression is important for the ability of cells to tolerate stress and regulate proteostasis to survive under environmental uncertainty.


Assuntos
Adaptação Fisiológica , Regulação Fúngica da Expressão Gênica , Histona Acetiltransferases/metabolismo , Proteínas Nucleares/metabolismo , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Ribonucleoproteínas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Estresse Fisiológico , Cromatina/metabolismo , Complexo Multienzimático de Ribonucleases do Exossomo/metabolismo , Proteostase , Transporte de RNA , Saccharomyces cerevisiae , Transativadores/metabolismo
15.
PLoS Pathog ; 15(5): e1007774, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31095645

RESUMO

The structural proteins of DNA viruses are generally encoded by late genes, whose expression relies on recruitment of the host transcriptional machinery only after the onset of viral genome replication. ß and γ-herpesviruses encode a unique six-member viral pre-initiation complex (vPIC) for this purpose, although how the vPIC directs specific activation of late genes remains largely unknown. The specificity underlying late transcription is particularly notable given that late gene promoters are unusually small, with a modified TATA-box being the only recognizable element. Here, we explored the basis for this specificity using an integrative approach to evaluate vPIC-dependent gene expression combined with promoter occupancy during Kaposi's sarcoma-associated herpesvirus (KSHV) infection. This approach distinguished the direct and indirect targets of the vPIC, ultimately revealing a novel promoter motif critical for KSHV vPIC binding. Additionally, we found that the KSHV vPIC component ORF24 is required for efficient viral DNA replication and identified a ORF24 binding element in the origin of replication that is necessary for late gene promoter activation. Together, these results identify an elusive element that contributes to vPIC specificity and suggest novel links between KSHV DNA replication and late transcription.


Assuntos
Regulação Viral da Expressão Gênica , Herpesvirus Humano 8/fisiologia , Regiões Promotoras Genéticas , Transativadores/metabolismo , Iniciação da Transcrição Genética , Proteínas Virais/genética , Replicação Viral , DNA Viral/genética , Genoma Viral , Células HEK293 , Interações Hospedeiro-Patógeno , Humanos , Ligação Proteica , RNA Polimerase II/metabolismo , Sarcoma de Kaposi/genética , Sarcoma de Kaposi/metabolismo , Sarcoma de Kaposi/virologia , Transativadores/genética , Proteínas Virais/metabolismo , Latência Viral
16.
Artif Cells Nanomed Biotechnol ; 47(1): 2003-2009, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31106596

RESUMO

Hepatitis B virus is one of the main causes of hepatitis and hepatocellular carcinoma (HCC). Hepatitis B virus-encoded X protein (HBx) has been shown to be involved in many aspects of the pathogenicity of liver diseases. Orexin A is a small peptide produced in the hippocampus. Orexin A and its receptor have become important therapeutic targets for certain metabolic disorders. In this study, we show that orexin A has a protective role against HBx-induced cytotoxicity and inflammation in hepatocytes. The ectopic expression of HBx in hepatocytes reduces orexin A receptor 1 (OX1R) expression. When orexin A is added to the cells, it mitigates HBx-induced oxidative stress indicator 4-hydroxynonenal (4-HNE) and reactive oxygen species (ROS) as well the NADPH subunit NADPH oxidase 4 (NOX-4). Orexin A also ameliorates HBx-mediated mitochondrial membrane potential and adenosine triphosphate (ATP) reduction. Moreover, orexin A significantly inhibits HBx-induced production of pro-inflammatory cytokines including interleukin 8 (IL-8), tumour necrosis factor α (TNF-α) and chemokine ligand 2 (CXCL2). The presence of orexin A ameliorates HBx-induced lactate dehydrogenase (LDH) release, indicating that it could protect hepatocytes from cytotoxicity. Mechanistically, we found that orexin A suppresses c-Jun N-terminal kinase (JNK) phosphorylation, accumulation of nuclear factor-κB (NF-κB) protein p65 in nuclei, and NF-κB promoter activity, suggesting that orexin A suppresses JNK and NF-κB pathway activation. In conclusion, our study demonstrates that orexin A peptide possesses a protective role against HBx-mediated cytotoxicity and inflammation in hepatocytes.


Assuntos
Citoproteção/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Orexinas/farmacologia , Transativadores/metabolismo , Linhagem Celular , Citocinas/metabolismo , Ativação Enzimática/efeitos dos fármacos , Hepatócitos/citologia , Hepatócitos/metabolismo , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Mitocôndrias/efeitos dos fármacos , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Transativadores/genética
17.
Int J Mol Sci ; 20(10)2019 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-31108974

RESUMO

Soil salinity represents a major constraint in the growth of chrysanthemum. Therefore, improving salinity tolerance of chrysanthemum has become an important research direction in tolerance breeding. Multiprotein bridging factor 1 (MBF1) is an evolutionarily highly conserved transcriptional co-activator in archaea and eukaryotes and has been reported to play important roles to respond to abiotic stresses. Here, a MBF1 gene induced by salt stress was isolated and functionally characterized from Dendranthema grandiflorum and name as DgMBF1. Overexpression of DgMBF1 in chrysanthemum increased the tolerance of plants to high salt stress compared to wild type (WT). It also showed fewer accumulations of hydrogen peroxide (H2O2), superoxide anion (O2-), higher activities of antioxidant enzymes, more content of proline and soluble sugar (SS) and more favorable K+/Na+ ratio than those of WT under salt stress. In addition, the expression level of genes related to antioxidant biosynthesis, proline biosynthesis, glyco-metabolism and K+/Na+ homeostasis was statistically significant higher in the DgMBF1-overexpressed lines than that in WT. These results demonstrated that DgMBF1 is a positive regulator in response to salt stress and could serve as a new candidate gene for salt-tolerant plant breeding.


Assuntos
Chrysanthemum/crescimento & desenvolvimento , Tolerância ao Sal , Transativadores/genética , Chrysanthemum/genética , Chrysanthemum/metabolismo , Clonagem Molecular , Regulação da Expressão Gênica de Plantas , Peróxido de Hidrogênio/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Análise de Sequência de DNA , Açúcares/metabolismo , Superóxidos/metabolismo , Transativadores/metabolismo
18.
J Med Food ; 22(5): 444-450, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31084542

RESUMO

Studies have identified the potential of chemopreventive effects of sulforaphane (SFN); however, the underlying mechanisms of its effect on breast cancer require further elucidation. This study investigated the anticancer effects of SFN that specifically induces G1/S arrest in breast ductal carcinoma (ZR-75-1) cells. The proliferation of the cancer cells after treatment with SFN was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. DNA content and cell cycle status were analyzed through flow cytometry. Our results demonstrated the inhibition of growth in ZR-75-1 cells upon SFN exposure. In addition, SERTAD1 (SEI-1) caused the accumulation of SFN-treated G1/S-phase cells. The downregulation of SEI-1, cyclin D2, and histone deacetylase 3 suggested that in addition to the identified effects of SFN against breast cancer prevention, it may also exert antitumor activities in established breast cancer cells. In conclusion, SFN can inhibit growth of and induce cell cycle arrest in cancer cells, suggesting its potential role as an anticancer agent.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Isotiocianatos/farmacologia , Proteínas Nucleares/genética , Transativadores/genética , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/fisiopatologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ciclina D2/genética , Ciclina D2/metabolismo , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Humanos , Proteínas Nucleares/metabolismo , Extratos Vegetais/farmacologia , Pontos de Checagem da Fase S do Ciclo Celular/efeitos dos fármacos , Transativadores/metabolismo , Fatores de Transcrição , Verduras/química
19.
Cell Physiol Biochem ; 52(6): 1503-1516, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31112017

RESUMO

BACKGROUND/AIMS: Zinc Finger Protein 281 (ZNF281) was recently identified as a novel oncogene in several human carcinomas. However, the clinical significance of ZNF281 in colorectal cancer (CRC) and the molecular mechanisms by which ZNF281 promotes the growth and metastasis of CRC remain unknown. METHODS: ZNF281 expression in CRC tissues was assessed, and the outcomes were analyzed to determine the clinical importance of ZNF281 expression. Cell Transwell assays and a wound healing assay were performed to assess the effects of ZNF281 on CRC cell migration and invasion in vitro. Western blotting was applied to analyze the potential mechanisms. RESULTS: ZNF281 mRNA and protein levels were significantly increased in CRC tissues compared with normal colon tissues, and high ZNF281 expression was associated with advanced T stage, N stage, TNM stage and differentiation. Therefore, ZNF281 expression might be an independent prognostic indicator in CRC patients. Moreover, knockdown of ZNF281 expression suppressed cell proliferation, migration and invasion by inhibiting the Wnt/ß-catenin pathway. CONCLUSION: Our study indicates that ZNF281 plays a critical role in the progression and metastasis of CRC and could represent a potential therapeutic target for CRC.


Assuntos
Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Transativadores/metabolismo , Idoso , Linhagem Celular Tumoral , Movimento Celular/genética , Movimento Celular/fisiologia , Proliferação de Células/genética , Proliferação de Células/fisiologia , Neoplasias Colorretais/genética , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Células HT29 , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Invasividade Neoplásica/fisiopatologia , Prognóstico , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Neoplásico/genética , RNA Neoplásico/metabolismo , RNA Interferente Pequeno/genética , Transativadores/antagonistas & inibidores , Transativadores/genética , Regulação para Cima , Via de Sinalização Wnt , beta Catenina/metabolismo
20.
Acta Crystallogr D Struct Biol ; 75(Pt 5): 498-504, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-31063152

RESUMO

Bovine meat and milk factors (BMMFs) are circular, single-stranded episomal DNAs that have been detected in bovine meat and milk products. BMMFs are thought to have roles in human malignant and degenerative diseases. BMMFs encode a replication initiator protein (Rep) that is actively transcribed and translated in human cells. In this study, a Rep WH1 domain encoded on a BMMF (MSBI1.176) isolated from a multiple sclerosis human brain sample was determined to 1.53 Šresolution using X-ray crystallography. The overall structure of the MSBI1.176 WH1 domain was remarkably similar to other Rep structures, despite having a low (28%) amino-acid sequence identity. The MSBI1.176 WH1 domain contained elements common to other Reps, including five α-helices, five ß-strands and a hydrophobic pocket. These new findings suggest that the MSBI1.176 Rep might have comparable roles and functions to other known Reps of different origins.


Assuntos
Encéfalo/metabolismo , DNA Helicases/química , DNA Helicases/metabolismo , Esclerose Múltipla/metabolismo , Plasmídeos/isolamento & purificação , Plasmídeos/metabolismo , Transativadores/química , Transativadores/metabolismo , Sequência de Aminoácidos , Animais , Bovinos , Cristalografia por Raios X , Humanos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Domínios Proteicos , Homologia de Sequência
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