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1.
Int J Mol Sci ; 22(20)2021 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-34681596

RESUMO

Epstein-Barr virus (EBV) is associated with several tumors and generates BamHI A rightward transcript (BART) microRNAs (miRNAs) from BART transcript introns. These BART miRNAs are expressed at higher levels in EBV-associated epithelial malignancies than in EBV-infected B lymphomas. To test the effects of EBV miRNA on the cell cycle and cell growth, we transfected miR-BART1-3p, a highly expressed EBV-associated miRNA, into gastric carcinoma cells. We found that miR-BART1-3p induced G0/G1 arrest and suppressed cell growth in gastric carcinoma cells. As our microarray analyses showed that E2F3, a cell cycle regulator, was inhibited by EBV infection, we hypothesized that miR-BART1-3p regulates E2F3. Luciferase assays revealed that miR-BART1-3p directly targeted the 3'-UTR of E2F3 mRNA. Both E2F3 mRNA and encoded protein levels were reduced following miR-BART1-3p transfection. In contrast, E2F3 expression in AGS-EBV cells transfected with a miR-BART1-3p inhibitor was enhanced. As E2F3 has been shown to regulate the expression of highly conserved miR-17-92 clusters in vertebrates, we examined whether this expression is affected by miR-BART1-3p, which can downregulate E2F3. The expression of E2F3, miR-17-92a-1 cluster host gene (MIR17HG), and miR-17-92 cluster miRNAs was significantly reduced in EBV-associated gastric carcinoma (EBVaGC) patients compared with EBV-negative gastric carcinoma (EBVnGC) patients. Further, miR-BART1-3p as well as the siRNA specific to E2F3 inhibited the expression of the miR-17-92 cluster, while inhibition of miR-BART1-3p enhanced the expression of the miR-17-92 cluster in cultured GC cells. Our results suggest a possible role of miR-BART1-3p in cell cycle regulation and in regulation of the miR-17-92 cluster through E2F3 suppression.


Assuntos
Fator de Transcrição E2F3/metabolismo , MicroRNAs/metabolismo , RNA Longo não Codificante/metabolismo , Regiões 3' não Traduzidas , Antagomirs/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Regulação para Baixo , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/genética , Pontos de Checagem da Fase G1 do Ciclo Celular , Regulação Neoplásica da Expressão Gênica , Herpesvirus Humano 4/isolamento & purificação , Humanos , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Interferência de RNA , RNA Longo não Codificante/genética , RNA Interferente Pequeno/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Neoplasias Gástricas/virologia
2.
Cell Signal ; 80: 109887, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33340658

RESUMO

Tumor cells switch from an epithelial to a mesenchymal-like phenotype, which represents a key hallmark of human cancer metastasis, including gallbladder cancer (GBC). A large set of microRNAs (miRNAs/miRs) have been studied to elucidate their functions in initiating or inhibiting this phenotypic switching in GBC cells. In this paper, we attempted to identify the expression pattern of the miR-214/-3120 cluster and its mode of action in the context of GBC, with a specific focus being placed on their effects on EMT and autophagy in GBC cells. Human GBC cells GBC-SD were assayed for their migration, invasion, and autophagy using the Transwell chamber system, MDC staining, and transmission electron microscopy. The tumorigenicity and metastatic behavior of GBC-SD cells were tested in nude mice. The expression of EMT- and autophagy-specific markers (E-cadherin, N-cadherin, vimentin, ATG5, LC3II/LC3I, and Beclin1) was analyzed in cultured GBC-SD cells and in human GBC-SD xenografts. The E2F3 luciferase reporter activity in the presence of miR-214/-3120 was evaluated by a dual luciferase assay. The miR-214/-3120 was downregulated in GBC. Exogenous miR-214/-3120 inhibited the phenotypic switching of GBC cells from epithelial to mesenchymal, prevented autophagy, and suppressed the tumorigenicity and metastatic behavior of GBC-SD cells in vitro and in vivo. E2F3 was demonstrated to be the target gene of miR-214/-3120, and its knockdown in part mimicked the effect of miR-214/-3120 on the EMT, autophagy, tumorigenicity, and metastatic behavior of GBC-SD cells. These results demonstrated that the miR-214/-3120 cluster blocks the process of EMT and autophagy to limit GBC metastasis by repressing E2F3 expression.


Assuntos
Autofagia , Transição Epitelial-Mesenquimal/genética , MicroRNAs/metabolismo , Animais , Antagomirs/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Regulação para Baixo , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/genética , Fator de Transcrição E2F3/metabolismo , Neoplasias da Vesícula Biliar/metabolismo , Neoplasias da Vesícula Biliar/patologia , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Camundongos Nus , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Transplante Heterólogo
3.
Biochem Biophys Res Commun ; 524(4): 816-824, 2020 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-32044038

RESUMO

Nasopharyngeal carcinoma (NPC) is a malignant tumor from head and neck with characteristics in remarkable geographic and racial distribution worldwide, which has the important features of vigorous proliferation and inflammatory cells infiltration. By analyzing the expression profile data of NPC, we found that the E2F-related gene sets were highly enriched in NPC tissues. E2F transcription factor family is an important cycle regulator, which can promote the malignant proliferation and tumorigenesis. Here, we showed that E2Fs accelerated malignant phenotypes of NPC cells. RNA sequencing revealed that E2Fs can significantly up-regulate the inflammatory pathways in NPC cells. E2F1, as a transcription factor, can active the transcription activity of IL-6 promoter, and modulate macrophage function through a microenvironment manner. Thus, this study characterized a significant role of E2Fs in inflammation and tumorigenesis of NPC, which provided a promising anti-tumor target in NPC, since E2Fs are highly expressed and activated in NPC.


Assuntos
Carcinogênese/genética , Fator de Transcrição E2F1/genética , Fator de Transcrição E2F3/genética , Regulação Neoplásica da Expressão Gênica , Carcinoma Nasofaríngeo/genética , Neoplasias Nasofaríngeas/genética , Animais , Apoptose/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , Ciclo Celular/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Estudos de Coortes , Fator de Transcrição E2F1/antagonistas & inibidores , Fator de Transcrição E2F1/metabolismo , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/metabolismo , Humanos , Inflamação , Interleucina-6/genética , Interleucina-6/metabolismo , Masculino , Camundongos , Camundongos Nus , Carcinoma Nasofaríngeo/metabolismo , Carcinoma Nasofaríngeo/patologia , Neoplasias Nasofaríngeas/metabolismo , Neoplasias Nasofaríngeas/patologia , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Células THP-1 , Carga Tumoral , Microambiente Tumoral/genética
4.
J BUON ; 23(5): 1492-1499, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30570877

RESUMO

PURPOSE: MicroRNA (miR)-194-5p is downregulated in bladder cancer (BC), but its role in BC has not been determined mechanistically. METHODS: The expression levels of miR-194-5p and E2F transcription factor 3 (E2F3) were determined by means of quantitative reverse transcription and polymerase chain reaction in BC specimens. In addition, T24 BC cells were transfected with a miR-194-5p mimic, a miR-194-5p inhibitor, or E2F3 small interfering (si)RNA, and the level of E2F3 protein expressed by these cells was assessed by western blotting. A dual luciferase reporter assay was applied to verify the binding site between miR-194-5p and the 3' untranslated region of E2F3. Transwell assays were performed to examine cell migration and invasion. RESULTS: Dysregulation of miR-194-5p in BC was closely associated with node metastasis and differentiation. In BC specimens and cell lines, miR-194-5p mRNA was downregulated, while E2F3 mRNA was upregulated. Overexpression of miR-194-5p suppressed the expression of E2F3 mRNA and protein. By regulating E2F3, miR-194-5p inhibited cell migration and invasion in BC. Treatment of BC cells with E2F3 siRNA had the same effect as did overexpression of miR-194-5p. CONCLUSIONS: MiR-194-5p directly targets E2F3 and inhibits cell migration and invasion in BC.


Assuntos
Fator de Transcrição E2F3/metabolismo , MicroRNAs/metabolismo , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/metabolismo , Movimento Celular/fisiologia , Regulação para Baixo , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/biossíntese , Fator de Transcrição E2F3/genética , Humanos , MicroRNAs/biossíntese , MicroRNAs/genética , Invasividade Neoplásica , Transfecção , Neoplasias da Bexiga Urinária/patologia
5.
Cell Death Dis ; 9(5): 509, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29724991

RESUMO

HOXB9, as a HOX family transcription factor, playing a significant role in embryonic development and cancer progression. However, the function of HOXB9 and its precise mechanism in regulating endometrial cancer progression remains unknown. Here, we demonstrated that the expression of HOXB9 was increased in endometrial cancer, and associated with histological grade and lymph node metastasis. In addition, elevated HOXB9 predicts a poor prognosis in endometrial cancer patients. Interestingly, bioinformatics analysis of TCGA cancer database showed that HOXB9 expression is positively correlated with E2F3 expression. Moreover, HOXB9 promoted E2F3 expression by directly targeting to its promoter. Furthermore, we found that knocking down E2F3 abolished the ability of HOXB9 in enhancing cell migration. Taken together, for the first, we demonstrated the function and mechanism of HOXB9 in regulating endometrial cancer progression, and indicated HOXB9 may be a novel prognostic marker of endometrial cancer.


Assuntos
Adenocarcinoma/genética , Fator de Transcrição E2F3/genética , Neoplasias do Endométrio/genética , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/mortalidade , Adenocarcinoma/patologia , Adulto , Idoso , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Progressão da Doença , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/metabolismo , Neoplasias do Endométrio/metabolismo , Neoplasias do Endométrio/mortalidade , Neoplasias do Endométrio/patologia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Proteínas de Homeodomínio/antagonistas & inibidores , Proteínas de Homeodomínio/metabolismo , Humanos , Metástase Linfática , Pessoa de Meia-Idade , Gradação de Tumores , Estadiamento de Neoplasias , Prognóstico , Regiões Promotoras Genéticas , Ligação Proteica , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Análise de Sobrevida
6.
Eur Rev Med Pharmacol Sci ; 22(3): 743-749, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29461605

RESUMO

OBJECTIVE: To detect the change in miRNA-210 expression of cardiomyocytes under hypoxia/reoxygenation status. Also, the effect of miR-210 on the apoptosis of cardiomyocytes induced by oxygen-glucose deprivation/reperfusion (OGD/R) and its mechanism through establishing the OGD/R injury model of primary cardiomyocytes in this experiment were investigated. MATERIALS AND METHODS: The cell model of OGD/R injury was established. The cell apoptosis in each group was detected by methyl thiazolyl tetrazolium (MTT) assay and detection of Caspase-3 activity. The change in miR-210 expression in each group was detected by Real-time fluorescence quantitative polymerase chain reaction (PCR). The high-expression and low-expression miR-210 models were established through the transient transfection of miR-210 mimic and inhibitor to detect the relevant indexes of cell apoptosis. At the same time, changes in mRNA and protein expressions of E2F3 were detected by RT-PCR and Western blotting, respectively. The E2F3 overexpression vector was constructed, and the overexpression vector plasmid and miR-210 mimic were jointly transfected into the cells to detect the relevant indexes of cell apoptosis. RESULTS: After OGD/R treatment, the activity of Caspase-3 was increased, the survival of cardiomyocytes was significantly inhibited and the expression level of miR-210 was up-regulated in OGD/R injury. Transfection of miR-210 mimic for miR-210 overexpression could alleviate the OGD/R-induced cardiomyocyte injury, while the decrease of miR-210 expression could aggravate the apoptosis of cardiomyocytes. In addition, the high expression of miR-210 could inhibit the protein expression of E2F3, and co-transfection of E2F3 plasmid and miR-210 mimic could reverse the inhibiting effect of miR-210 on the apoptosis of cardiomyocytes. CONCLUSIONS: We confirmed that miR-210 can inhibit the OGD/R-induced apoptosis of cardiomyocytes, and miR-210, as an upstream factor, plays a protective role in cardiomyocytes through directly inhibiting the protein expression of its target gene E2F3.


Assuntos
Fator de Transcrição E2F3/biossíntese , Glucose/deficiência , MicroRNAs/biossíntese , Miócitos Cardíacos/metabolismo , Oxigênio/metabolismo , Traumatismo por Reperfusão/metabolismo , Apoptose/fisiologia , Hipóxia Celular/fisiologia , Células Cultivadas , Fator de Transcrição E2F3/antagonistas & inibidores , Glucose/metabolismo , Humanos , Traumatismo por Reperfusão/prevenção & controle
7.
Eur Rev Med Pharmacol Sci ; 21(18): 4050-4057, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29028097

RESUMO

OBJECTIVE: Pancreatic cancer (PC) is the most malignant tumor among all the tumors in the digestive system. MiR-217 has been reported to take a critical part in various malignant tumors. The aim of this study was to explore the function of MiR-217 in pancreatic cancer and its target genes. PATIENTS AND METHODS: Twenty pairs of PC tissues and matched normal adjacent pancreatic tissues were collected. The expression of miR-217 in PC tissues and normal pancreatic tissues was detected by Real-time polymerase chain reaction (PCR). PC cells were transfected with miR-217 mimics, inhibitors and negative control, respectively. Cell Counting Kit-8 (CCK-8) assay was used to detect cell viability. Cell apoptosis was checked via Annexin V-FITC/PI apoptosis kit. The protein expression of E2F3 was detected by Western blot. To detect repression by miR-217, HEK293T cells were co-transfected with the indicated E2F3 3'-UTR luciferase reporter. RESULTS: The expression of miR-217 was reduced in PC tissues comparing to normal pancreatic tissues. Meantime, the in-vitro study revealed that miR-217 suppressed PC cell growth, invasion but promoted apoptosis. Next, we proved that E2F3 was the target of miR-217 on PC cell function. CONCLUSIONS: miR-217 suppresses PC cell growth, invasion but promotes apoptosis in vitro through targeting E2F3. The miR-217-E2F3 axis may be used for PC therapy.


Assuntos
Fator de Transcrição E2F3/metabolismo , MicroRNAs/metabolismo , Neoplasias Pancreáticas/patologia , Regiões 3' não Traduzidas , Antagomirs/metabolismo , Apoptose , Sequência de Bases , Estudos de Casos e Controles , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação para Baixo , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/genética , Células HEK293 , Humanos , MicroRNAs/química , MicroRNAs/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Prognóstico , Alinhamento de Sequência
8.
Cell Physiol Biochem ; 35(5): 2033-42, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25871967

RESUMO

BACKGROUND: MicroRNA-449a is a tumor suppressor that is down-regulated in multiple tumors types. However, the role of miR-449a in gastric cancer (GC) remains largely unknown. METHODS: MiR-449a expression was up-regulated using miR-449a mimics, and the role of miR-449a in GC was assessed using cell viability and apoptosis assays. miR-449a target genes were confirmed using luciferase activity, RT-PCR and western blot assays. RESULTS: miR-449a was downregulated in gastric cancer cell lines and gastric cancer tissues. Restoration of miR-449a expression inhibited gastric cancer cell proliferation and colony formation. Significant G0/G1 arrest was observed in gastric cancer cells transfected with miR-449a mimics. Furthermore, combination therapy with miR-449a with cisplatin displayed greater anti-tumor effects than treatment with cisplatin alone. We also identified E2F3 (E2F transcription factor 3), an important transcription factor involved in the proliferation and metastasis of tumor cells, as a direct target gene of miR-449a. Furthermore, silencing E2F3 elicits similar a repressive effect as overexpression of miR-449a in gastric cancer cells, and E2F3 overexpression rescued the repressing effects of miR-449a mimics. CONCLUSIONS: This study indicates that the miR-449a/E2F3 axis plays an important role in proliferation and apoptosis in gastric cancer. Therefore, miR-449a represents a novel target for gastric cancer therapy.


Assuntos
Apoptose , Fator de Transcrição E2F3/metabolismo , MicroRNAs/metabolismo , Regiões 3' não Traduzidas , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Sequência de Bases , Western Blotting , Proliferação de Células/efeitos dos fármacos , Cisplatino/farmacologia , Regulação para Baixo/efeitos dos fármacos , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/genética , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Humanos , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Oligonucleotídeos Antissenso/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Alinhamento de Sequência , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia
9.
Oncotarget ; 6(6): 3918-31, 2015 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-25714015

RESUMO

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths. Protoporphyrin IX (PPIX) has been used for photodynamic therapy. Mesenchymal cancer cells adapt to tumor microenvironments for growth and metastasis possibly in association with miRNA dysregulation. In view of the effect of PPIX on cancer-related genes, and its potential to inhibit tumor growth and migration/invasion, this study investigated whether PPIX enables mesenchymal liver tumor to restore dysregulated miRNAs, and if so, whether it sensitizes the cancer cells to chemotherapy. In addition, we explored new target(s) of the miRNA(s) that contribute to the anti-cancer effects. Of the ten miRNAs predicted by the 3'-UTR of HIF-1α mRNA, PPIX treatment increased miR-199a-5p, leading to the inhibition of E2F3 expression which is upregulated in mesenchymal liver tumor. miR-199a-5p levels were downregulated in HCC with E2F3 overexpression. An approach modulating epithelial-mesenchymal transition provided the expected changes in miR-199a-5p and E2F3 in vivo. PPIX prevented tumor cell growth and migration/invasion, and had a synergistic anti-cancer effect when combined with chemotherapeutics. In a xenograft model, PPIX treatment decreased overall growth and average tumor volume, which paralleled E2F3 inhibition. Overall, PPIX inhibited growth advantage and migratory ability of cancer cells and sensitized mesenchymal liver tumor cells to chemotherapeutics.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Carcinoma Hepatocelular/tratamento farmacológico , Fator de Transcrição E2F3/antagonistas & inibidores , Neoplasias Hepáticas/tratamento farmacológico , Células-Tronco Mesenquimais/efeitos dos fármacos , MicroRNAs/biossíntese , Fármacos Fotossensibilizantes/farmacologia , Protoporfirinas/farmacologia , Animais , Sequência de Bases , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Movimento Celular/efeitos dos fármacos , Sinergismo Farmacológico , Fator de Transcrição E2F3/genética , Fator de Transcrição E2F3/metabolismo , Células Hep G2 , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/biossíntese , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/genética , MicroRNAs/metabolismo , Dados de Sequência Molecular , Protoporfirinas/administração & dosagem , Distribuição Aleatória , Transfecção , Regulação para Cima/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Mol Cell Biochem ; 392(1-2): 289-96, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24671492

RESUMO

The poor prognosis of hepatocellular carcinoma (HCC) is mainly due to the development of invasion and metastasis. Recent data strongly suggests the important role of miRNAs in cancer progression, including invasion and metastasis. Here, we found miR-217 expression was much lower in highly invasive MHCC-97H HCC cells and metastatic HCC tissues. Restored miR-217 expression with miR-217 mimics inhibited invasion of MHCC-97H cells. Inversely, miR-217 inhibition enhanced the invasive ability of Huh7 and MHCC-97L cells. Mechanically, bioinformatics analysis combined with experimental analysis demonstrated E2F3 was a novel direct target of miR-217. Moreover, E2F3 protein level was positively associated with HCC metastasis and functional analysis confirmed the positive role of E2F3 in HCC cell invasion. Our findings suggest miR-217 function as a potential tumor suppressor in HCC progression and miR-217-E2F3 axis may be a novel candidate for developing rational therapeutic strategies.


Assuntos
Carcinoma Hepatocelular/patologia , Fator de Transcrição E2F3/antagonistas & inibidores , Neoplasias Hepáticas/patologia , MicroRNAs/fisiologia , Invasividade Neoplásica/genética , Sequência de Bases , Linhagem Celular Tumoral , Primers do DNA , Humanos , MicroRNAs/genética , Reação em Cadeia da Polimerase em Tempo Real
11.
Pak J Pharm Sci ; 25(4): 793-801, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23009996

RESUMO

The transcription factor-based therapeutic approaches are the mainstay of current anticancer drug design options to develop highly selective agents with novel modes of action. In this paper, a homology model of DNA-binding domain of transcription factor E2F3 was generated according to X-ray structure of E2F4. As a first step of our proposed project aspired towards exploration of highly selective potential E2F3 ligands, we performed structure-based virtual screening of ZINC 3D chemical database by using Dock Blaster server. Then 31 compounds, selected by filtration step, were docked against the prominent DNA binding site residues of E2F3 model. Two of them have shown a promising interaction with respect to binding poses. The aim is to propose new active ligands against neoplasias characterized by overexpression of E2F3 transcription factor.


Assuntos
Antineoplásicos/química , Desenho de Fármacos , Fator de Transcrição E2F3/química , Sequência de Aminoácidos , Antineoplásicos/farmacologia , Sítios de Ligação , Desenho Assistido por Computador , Cristalografia por Raios X , DNA/química , Bases de Dados de Compostos Químicos , Fator de Transcrição E2F3/antagonistas & inibidores , Humanos , Ligantes , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Molecular , Conformação Proteica , Relação Estrutura-Atividade
12.
Oncogene ; 26(7): 1028-37, 2007 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-16909110

RESUMO

Amplification and overexpression of the E2F3 gene at 6p22 in human bladder cancer is associated with increased tumour stage, grade and proliferation index, and in prostate cancer E2F3 overexpression is linked to tumour aggressiveness. We first used small interfering RNA technology to confirm the potential importance of E2F3 overexpression in bladder cancer development. Knockdown of E2F3 expression in bladder cells containing the 6p22 amplicon strongly reduced the extent of bromodeoxyuridine (BrdU) incorporation and the rate of cellular proliferation. In contrast, knockdown of CDKAL1/FLJ20342, another proposed oncogene, from this amplicon had no effect. Expression cDNA microarray analysis on bladder cancer cells following E2F3 knockdown was then used to identify genes regulated by E2F3, leading to the identification of known E2F3 targets such as Cyclin A and CDC2 and novel targets including pituitary tumour transforming gene 1, Polo-like kinase 1 (PLK1) and Caveolin-2. For both bladder and prostate cancer, we have proposed that E2F3 protein overexpression may cooperate with removal of the E2F inhibitor retinoblastoma tumor suppressor protein (pRB) to drive cellular proliferation. In support of this model, we found that ectopic expression of E2F3a enhanced the BrdU incorporation, a marker of cellular proliferation rate, of prostate cancer DU145 cells, which lack pRB, but had no effect on the proliferation rate of PC3 prostate cancer cells that express wild-type pRB. BrdU incorporation in PC3 cells could, however, be increased by overexpressing E2F3a in cells depleted of pRB. When taken together, these observations indicate that E2F3 levels have a critical role in modifying cellular proliferation rate in human bladder and prostate cancer.


Assuntos
Proliferação de Células , Fator de Transcrição E2F3/fisiologia , Neoplasias da Próstata/patologia , Neoplasias da Bexiga Urinária/patologia , Linhagem Celular Tumoral , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/biossíntese , Fator de Transcrição E2F3/genética , Humanos , Masculino , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/metabolismo
13.
Oncogene ; 26(3): 321-7, 2007 Jan 18.
Artigo em Inglês | MEDLINE | ID: mdl-16909124

RESUMO

Functions encoded by single genes in lower organisms are often represented by multiple related genes in the mammalian genome. An example is the retinoblastoma and E2F families of proteins that regulate transcription during the cell cycle. Analysis of gene function using germline mutations is often confounded by overlapping function resulting in compensation. Indeed, in cells deleted of the E2F1 or E2F3 genes, there is an increase in the expression of the other family member. To avoid complications of compensatory effects, we have used small-interfering RNAs that target individual E2F proteins to generate a temporary loss of E2F function. We find that both E2F1 and E2F3 are required for cells to enter the S phase from a quiescent state, whereas only E2F3 is necessary for the S phase in growing cells. We also find that the acute loss of E2F3 activity affects the expression of genes encoding DNA replication and mitotic activities, whereas loss of E2F1 affects a limited number of genes that are distinct from those regulated by E2F3. We conclude that the long-term loss of E2F activity does lead to compensation by other family members and that the analysis of acute loss of function reveals specific and distinct roles for these proteins.


Assuntos
Biomarcadores/metabolismo , Fator de Transcrição E2F1/fisiologia , Fator de Transcrição E2F3/fisiologia , Animais , Western Blotting , Bromodesoxiuridina/metabolismo , Proliferação de Células , Replicação do DNA , Fator de Transcrição E2F1/antagonistas & inibidores , Fator de Transcrição E2F1/genética , Fator de Transcrição E2F3/antagonistas & inibidores , Fator de Transcrição E2F3/genética , Embrião de Mamíferos/citologia , Embrião de Mamíferos/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Camundongos , Mitose , Análise de Sequência com Séries de Oligonucleotídeos , RNA Interferente Pequeno/farmacologia , Ratos , Fase S , Transcrição Gênica
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