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1.
Sensors (Basel) ; 24(9)2024 Apr 29.
Artigo em Inglês | MEDLINE | ID: mdl-38732954

RESUMO

Biometric fingerprint identification hinges on the reliability of its sensors; however, calibrating and standardizing these sensors poses significant challenges, particularly in regards to repeatability and data diversity. To tackle these issues, we propose methodologies for fabricating synthetic 3D fingerprint targets, or phantoms, that closely emulate real human fingerprints. These phantoms enable the precise evaluation and validation of fingerprint sensors under controlled and repeatable conditions. Our research employs laser engraving, 3D printing, and CNC machining techniques, utilizing different materials. We assess the phantoms' fidelity to synthetic fingerprint patterns, intra-class variability, and interoperability across different manufacturing methods. The findings demonstrate that a combination of laser engraving or CNC machining with silicone casting produces finger-like phantoms with high accuracy and consistency for rolled fingerprint recordings. For slap recordings, direct laser engraving of flat silicone targets excels, and in the contactless fingerprint sensor setting, 3D printing and silicone filling provide the most favorable attributes. Our work enables a comprehensive, method-independent comparison of various fabrication methodologies, offering a unique perspective on the strengths and weaknesses of each approach. This facilitates a broader understanding of fingerprint recognition system validation and performance assessment.

2.
Soft Matter ; 16(34): 8078-8084, 2020 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-32789349

RESUMO

In a recent publication [Bergmann et al. Phys. Chem. Chem. Phys., 2018, 20, 5074-5083] we presented a method which enables to investigate the morphology of microgels by superresolution fluorescence microscopy. Here, this method is applied to three microgel species, based on N-isopropylmethacrylamide (NIPMAM), N-n-propylacrylamide (NNPAM) and N-n-propylmethacrylamide (NNPMAM)) with 5, 7.5 and 10 mol% cross-linker, respectively. Super-resolution microscopy reveals differences of the network morphology of the synthesized particles showing the importance of the monomer structure.

3.
Nanoscale Adv ; 2(1): 323-331, 2020 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-36134006

RESUMO

We investigate the internal morphology of smart core-shell microgels by super-resolution fluorescence microscopy exploiting a combination of 3D single molecule localization and structured illumination microscopy utilizing freely diffusing fluorescent dyes. This approach does not require any direct chemical labeling and does not perturb the network structure of these colloidal gels. Hence, it allows us to study the morphology of the particles with very high precision. We found that the structure of the core-forming seed particles is drastically changed by the second synthesis step necessary for making the shell, resulting in a core region with highly increased dye localization density. The present work shows that super-resolution microscopy has great potential with respect to the study of soft colloidal systems.

4.
Phys Chem Chem Phys ; 20(7): 5074-5083, 2018 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-29392265

RESUMO

We present a new method to resolve the network morphology of colloidal particles in an aqueous environment via super-resolution microscopy. By localization of freely diffusing fluorophores inside the particle network we can resolve the three dimensional structure of one species of colloidal particles (thermoresponsive microgels) without altering their chemical composition through copolymerization with fluorescent monomers. Our approach utilizes the interaction of the fluorescent dye rhodamine 6G with the polymer network to achieve an indirect labeling. We calculate the 3D structure from the 2D images and compare the structure to previously published models for the microgel morphology, e.g. the fuzzy sphere model. To describe the differences in the data an extension of this model is suggested. Our method enables the tailor-made fabrication of colloidal particles which are used in various applications, such as paints or cosmetics, and are promising candidates for drug delivery, smart surface coatings, and nanocatalysis. With the precise knowledge of the particle morphology an understanding of the underlying structure-property relationships for various colloidal systems is possible.

5.
Phys Chem Chem Phys ; 19(6): 4887-4890, 2017 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-28116366

RESUMO

Certain fluorophores, in particular those that can undergo photoinduced radical pair reactions are known to exhibit a magnetic field dependent fluorescence summarized in the term magnetic field effect (MFE). We tried to reproduce experiments that reported magnetic field enhanced fluorescence for commonly used organic dyes with a high quantum yield suitable for single molecule localization microscopy. We find that the enhanced fluorescence is due to fluorescence reflected by the magnet's surface rather than MFE.

6.
ACS Nano ; 9(8): 8122-30, 2015 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-26173009

RESUMO

We describe a method for the deposition of minute amounts of fluorophore-labeled oligonucleotides with high local precision in conductive and transparent solid layers of poly(vinyl alcohol) (PVA) doped with glycerin and cysteamine (PVA-G-C layers). Deposition of negatively charged fluorescent molecules was accomplished with a setup based on a scanning ion conductance microscope (SICM) using nanopipettes with tip diameters of ∼100 nm by using the ion flux flowing between two electrodes through the nanopipette. To investigate the precision of the local deposition process, we performed in situ super-resolution microscopy by direct stochastic optical reconstruction microscopy (dSTORM). Exploiting the single-molecule sensitivity and reliability of dSTORM, we determine the number of fluorescent molecules deposited in single spots. The correlation of applied charge and number of deposited molecules enables the quantification of delivered molecules by measuring the charge during the delivery process. We demonstrate the reproducible deposition of 3-168 fluorescent molecules in single spots and the creation of fluorescent structures. The fluorescent structures are highly stable and can be reused several times.


Assuntos
Microscopia de Fluorescência/métodos , Nanotecnologia/métodos , Oligonucleotídeos/química , Coloração e Rotulagem/métodos , Carbocianinas/química , Cisteamina/química , Eletrodos , Corantes Fluorescentes/química , Glicerol/química , Microscopia de Fluorescência/instrumentação , Nanotecnologia/instrumentação , Álcool de Polivinil/química , Eletricidade Estática , Processos Estocásticos
7.
J Exp Med ; 203(11): 2409-12, 2006 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-17043147

RESUMO

CD40 was initially identified as a receptor expressed by B cells that is crucial for inducing an effective adaptive immune response. CD40 was subsequently shown to be expressed by endothelial cells and to promote angiogenesis. New data now show that in tumor-prone transgenic mice, CD40-mediated neovascularization is essential for early stage tumorigenicity. This suggests, at least in this mouse model, that CD40 has an important role in the angiogenic process that is coupled to carcinogenesis, a finding that could lead to novel therapeutic opportunities.


Assuntos
Antígenos CD40/fisiologia , Neoplasias Mamárias Experimentais/imunologia , Neovascularização Patológica/imunologia , Animais , Antígenos CD40/biossíntese , Humanos , Neoplasias Mamárias Experimentais/etiologia
8.
Oncogene ; 24(37): 5693-700, 2005 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-16123802

RESUMO

Cellular homeostasis is tightly controlled by the various pathways that regulate cell proliferation and cell death. Breaking this balance is often associated with cancer development. The transforming growth factor-beta (TGF-beta) pathway plays an important role in cellular homeostasis by regulating cell growth inhibition, cellular senescence, differentiation and apoptosis. Deregulated TGF-beta signaling is known to be involved in a variety of human cancers, including those of the colon, pancreas, breast and prostate. While TGF-beta is a potent negative regulator of hematopoiesis, the role of aberrant TGF-beta signaling in leukemogenesis remains largely unknown. Recently, evidence demonstrating deregulated TGF-beta signaling in leukemogenesis, particularly in acute promyelocytic leukemia (APL), has started to emerge. In this review, we summarize the current progress towards the understanding of the molecular mechanisms by which aberrant TGF-beta signaling may participate in leukemogenesis.


Assuntos
Leucemia/etiologia , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta/fisiologia , Endossomos/fisiologia , Células-Tronco Hematopoéticas/fisiologia , Humanos , Leucemia Promielocítica Aguda/etiologia , Microdomínios da Membrana/fisiologia , Proteínas de Neoplasias/fisiologia , Proteínas Nucleares/fisiologia , Proteína da Leucemia Promielocítica , Fatores de Transcrição/fisiologia , Proteínas Supressoras de Tumor/fisiologia
9.
Cancer Res ; 65(10): 4078-87, 2005 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-15899797

RESUMO

YB-1 protein levels are elevated in most human breast cancers, and high YB-1 levels have been correlated with drug resistance and poor clinical outcome. YB-1 is a stress-responsive, cell cycle-regulated transcription factor with additional functions in RNA metabolism and translation. In this study, we show in a novel transgenic mouse model that human hemagglutinin-tagged YB-1 provokes remarkably diverse breast carcinomas through the induction of genetic instability that emerges from mitotic failure and centrosome amplification. The increase of centrosome numbers proceeds during breast cancer development and explanted tumor cell cultures show the phenotype of ongoing numerical chromosomal instability. These data illustrate a mechanism that might contribute to human breast cancer development.


Assuntos
Neoplasias da Mama/genética , Transformação Celular Neoplásica/genética , Centrossomo/fisiologia , Proteínas de Ligação a DNA/genética , Neoplasias Mamárias Experimentais/genética , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Transformação Celular Neoplásica/patologia , Instabilidade Cromossômica , Proteínas de Ligação a DNA/biossíntese , Modelos Animais de Doenças , Feminino , Amplificação de Genes , Humanos , Neoplasias Mamárias Experimentais/metabolismo , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Transgênicos , Proteínas Nucleares , Proteína 1 de Ligação a Y-Box
10.
Oncogene ; 24(22): 3606-18, 2005 May 19.
Artigo em Inglês | MEDLINE | ID: mdl-15750632

RESUMO

Vaults have been suggested to play a direct role in multidrug resistance (MDR) to anticancer drugs. The human major vault protein (MVP) also known as lung resistance-related protein (LRP) represents the predominant component of vaults that may be involved in the defense against xenobiotics. Here, we demonstrate that besides MDR-related cytostatics, also the non-MDR-related drug 5-fluorouracil (5-FU) was able to induce MVP mRNA and protein expression. Treatment with 5-FU amplified the binding activity and interaction of the transcription factor Y-box binding protein-1 (YB-1) with the Y-box of the human MVP gene promoter in a time-dependent manner. 5-FU also induced reporter expressions driven by a panel of newly generated MVP promoter deletion mutants. Interestingly, stably YB-1 overexpressing cell clones showed enhanced binding of YB-1 to the Y-box motif, associated with enhanced basal as well as 5-FU-inducible MVP promoter-driven reporter expressions. Moreover, transduction of YB-1 cDNA led to increased expression of endogenous MVP protein. Under physiological conditions, we observed a strong coexpression of MVP and YB-1 in human colon carcinoma specimen. In summary, our data demonstrate a direct involvement of YB-1 in controlling basal and 5-FU-induced MVP promoter activity. Therefore, YB-1 is directly linked to MVP-mediated drug resistance.


Assuntos
Antimetabólitos Antineoplásicos/farmacologia , Proteínas de Ligação a DNA/metabolismo , Resistencia a Medicamentos Antineoplásicos/fisiologia , Fluoruracila/farmacologia , Partículas de Ribonucleoproteínas em Forma de Abóbada/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Proteínas de Ligação a DNA/efeitos dos fármacos , Resistência a Múltiplos Medicamentos , Ensaio de Desvio de Mobilidade Eletroforética , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Proteínas Nucleares , Regiões Promotoras Genéticas , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Partículas de Ribonucleoproteínas em Forma de Abóbada/genética , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Proteína 1 de Ligação a Y-Box
11.
Blood ; 105(9): 3686-90, 2005 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-15626733

RESUMO

The promyelocytic leukemia (PML) gene, a tumor suppressor inactivated in acute promyelocytic leukemia (APL), regulates apoptosis induced by DNA damage. However, the molecular mechanisms by which PML modulates apoptosis following genotoxic stress are only partially elucidated. PML is essential for p53-dependent induction of programmed cell death upon gamma-irradiation through PML-nuclear body (NB)-mediated control of p53 acetylation. Here, we show that PML selectively regulates proapoptotic transcription factors upon different types of DNA damage. We find that Pml inactivation protects fibroblasts from UV-induced apoptosis in a p53-independent manner. We demonstrate that c-Jun is required for UV-induced apoptosis and that PML is essential for both c-Jun transcriptional activation and DNA binding upon UV radiation. We find that PML physically interacts with c-Jun and that upon UV radiation the PML-NBs reorganize into novel nuclear microspeckled structures (UV-NBs), where PML and c-Jun dynamically accumulate. These data identify a novel PML-dependent pathway for c-Jun transcriptional activation and induction of apoptosis in response to DNA damage and shed new light on the role of PML in tumor suppression.


Assuntos
Dano ao DNA , Proteínas de Neoplasias/fisiologia , Proteínas Nucleares/fisiologia , Proteínas Proto-Oncogênicas c-jun/fisiologia , Fatores de Transcrição/fisiologia , Animais , Apoptose , Células Cultivadas , Dano ao DNA/efeitos da radiação , Embrião de Mamíferos/citologia , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Proteína da Leucemia Promielocítica , Ligação Proteica , Proteínas Proto-Oncogênicas c-jun/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Ativação Transcricional/efeitos da radiação , Transfecção , Proteína Supressora de Tumor p53 , Proteínas Supressoras de Tumor , Raios Ultravioleta
12.
Nature ; 431(7005): 205-11, 2004 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-15356634

RESUMO

Transforming growth factor beta (TGF-beta) is a pluripotent cytokine that controls key tumour suppressive functions, but cancer cells are often unresponsive to it. The promyelocytic leukaemia (PML) tumour suppressor of acute promyelocytic leukaemia (APL) accumulates in the PML nuclear body, but cytoplasmic PML isoforms of unknown function have also been described. Here we show that cytoplasmic Pml is an essential modulator of TGF-beta signalling. Pml-null primary cells are resistant to TGF-beta-dependent growth arrest, induction of cellular senescence and apoptosis. These cells also have impaired phosphorylation and nuclear translocation of the TGF-beta signalling proteins Smad2 and Smad3, as well as impaired induction of TGF-beta target genes. Expression of cytoplasmic Pml is induced by TGF-beta. Furthermore, cytoplasmic PML physically interacts with Smad2/3 and SARA (Smad anchor for receptor activation) and is required for association of Smad2/3 with SARA and for the accumulation of SARA and TGF-beta receptor in the early endosome. The PML-RARalpha oncoprotein of APL can antagonize cytoplasmic PML function and APL cells have defects in TGF-beta signalling similar to those observed in Pml-null cells. Our findings identify cytoplasmic PML as a critical TGF-beta regulator, and further implicate deregulated TGF-beta signalling in cancer pathogenesis.


Assuntos
Citoplasma/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linfócitos B/citologia , Linfócitos B/efeitos dos fármacos , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Linhagem Celular Tumoral , Senescência Celular/efeitos dos fármacos , Citoplasma/efeitos dos fármacos , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Endossomos/efeitos dos fármacos , Endossomos/metabolismo , Fibroblastos , Proteínas de Ligação ao GTP , Humanos , Leucemia Promielocítica Aguda/metabolismo , Leucemia Promielocítica Aguda/patologia , Camundongos , Mutação/genética , Proteínas de Neoplasias/deficiência , Proteínas de Neoplasias/genética , Proteínas Nucleares/deficiência , Proteínas Nucleares/genética , Proteínas de Fusão Oncogênica/metabolismo , Proteína da Leucemia Promielocítica , Ligação Proteica , Transporte Proteico , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Proteína Smad2 , Proteína Smad3 , Transativadores/genética , Transativadores/metabolismo , Fatores de Transcrição/deficiência , Fatores de Transcrição/genética , Transcrição Gênica/efeitos dos fármacos , Proteínas Supressoras de Tumor
13.
J Neurosci ; 24(26): 5966-73, 2004 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-15229244

RESUMO

The serotonin transporter (5-HTT) gene contains a variable number tandem repeat (VNTR) domain within intron 2 that is often associated with a number of neurological conditions, including affective disorders. The implications of this polymorphism are not yet understood, however, we have previously demonstrated that the 5-HTT VNTR is a transcriptional regulatory domain, and the allelic variation supports differential reporter gene expression in vivo and in vitro. The aim of this study was to identify transcription factors responsible for the regulation of this VNTR. Using a yeast one-hybrid screen, we found the transcription factor Y box binding protein 1 (YB-1) interacts with the 5-HTT VNTR. Consistent with this, we demonstrate in a reporter gene assay that the polymorphic VNTR domains differentially respond to exogenous YB-1 and that YB-1 will bind to the VNTR in vitro in a sequence-specific manner. Interestingly, the transcription factor CCTC-binding factor (CTCF), previously shown to interact with YB-1, interferes with the ability of the VNTR to support YB-1-directed reporter gene expression. In addition, CTCF blocks the binding of YB-1 to its DNA recognition sequences in vitro, thus providing a possible mechanism of regulation of YB-1 activation of the VNTR by CTCF. Therefore, we have identified YB-1 and CTCF as transcription factors responsible, at least in part, for modulation of VNTR function as a transcriptional regulatory domain. Our data suggest a novel mechanism that explains, in part, the ability of the distinct VNTR copy numbers to support differential reporter gene expression based on YB-1 binding sites.


Assuntos
Proteínas de Ligação a DNA/fisiologia , Regulação da Expressão Gênica/fisiologia , Glicoproteínas de Membrana/genética , Proteínas de Membrana Transportadoras/genética , Repetições Minissatélites , Proteínas do Tecido Nervoso/genética , Proteínas Repressoras/fisiologia , Fatores de Transcrição/fisiologia , Alelos , Animais , Sítios de Ligação , Fator de Ligação a CCCTC , Células COS , Linhagem Celular , Galinhas , Chlorocebus aethiops , DNA Complementar/genética , Genes Reporter , Humanos , Íntrons/genética , Rim , Glicoproteínas de Membrana/biossíntese , Proteínas de Membrana Transportadoras/biossíntese , Transtornos do Humor/genética , Transtornos do Humor/metabolismo , Proteínas do Tecido Nervoso/biossíntese , Proteínas Nucleares , Polimorfismo Genético , Ligação Proteica/efeitos dos fármacos , Mapeamento de Interação de Proteínas , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/metabolismo , Proteínas da Membrana Plasmática de Transporte de Serotonina , Transcrição Gênica , Transfecção , Técnicas do Sistema de Duplo-Híbrido , Proteína 1 de Ligação a Y-Box , Dedos de Zinco
14.
Nat Cell Biol ; 6(7): 665-72, 2004 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15195100

RESUMO

The promyelocytic leukaemia (PML) tumour-suppressor protein potentiates p53 function by regulating post-translational modifications, such as CBP-dependent acetylation and Chk2-dependent phosphorylation, in the PML-Nuclear Body (NB). PML was recently shown to interact with the p53 ubiquitin-ligase Mdm2 (refs 4-6); however, the mechanism by which PML regulates Mdm2 remains unclear. Here, we show that PML enhances p53 stability by sequestering Mdm2 to the nucleolus. We found that after DNA damage, PML and Mdm2 accumulate in the nucleolus in an Arf-independent manner. In addition, we found that the nucleolar localization of PML is dependent on ATR activation and phosphorylation of PML by ATR. Notably, in Pml(-/-) cells, sequestration of Mdm2 to the nucleolus was impaired, as well as p53 stabilization and the induction of apoptosis. Furthermore, we demonstrate that PML physically associates with the nucleolar protein L11, and that L11 knockdown impairs the ability of PML to localize to nucleoli after DNA damage. These findings demonstrate an unexpected role of PML in the nucleolar network for tumour suppression.


Assuntos
Nucléolo Celular/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Fator 1 de Ribosilação do ADP/genética , Fator 1 de Ribosilação do ADP/metabolismo , Transporte Ativo do Núcleo Celular/genética , Animais , Proteínas Mutadas de Ataxia Telangiectasia , Compartimento Celular/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Transformada , Nucléolo Celular/genética , Células Cultivadas , Fibroblastos , Humanos , Camundongos , Células NIH 3T3 , Proteínas de Neoplasias/genética , Proteínas Nucleares/genética , Fosforilação , Proteína da Leucemia Promielocítica , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Transporte Proteico/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-mdm2 , Estabilidade de RNA/genética , Proteínas Ribossômicas/genética , Proteínas Ribossômicas/metabolismo , Fatores de Transcrição/genética , Proteína Supressora de Tumor p53/genética , Proteínas Supressoras de Tumor/genética
15.
Cancer Res ; 64(1): 322-8, 2004 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-14729641

RESUMO

Resistance to chemotherapy is responsible for a failure of current treatment regimens in cancer patients. We have reported previously that the Y-box protein YB-1 regulates expression of the P-glycoprotein gene mdr1, which plays a major role in the development of a multidrug resistant-tumor phenotype. YB-1 predicts drug resistance and patient outcome in breast cancer. Thus, YB-1 is a promising target for new therapeutic approaches to defeat multidrug resistance. In drug-resistant cancer cells and in adenovirus-infected cells YB-1 is found in the nucleus. Nuclear accumulation of YB-1 in adenovirus-infected cells is a function of the E1 region, and we have shown that YB-1 facilitates adenovirus replication. Here we report that E1A-deleted or mutant adenovirus vectors, such as Ad312 and Ad520, replicate efficiently in multidrug-resistant (MDR) cancer cells and induce an adenovirus cytopathic effect resulting in host cell lysis. Thus, replication-defective adenoviruses are a previously unrecognized vector system for a selective elimination of MDR cancer cells. Our work forms the basis for the development of novel oncolytic adenovirus vectors for the treatment of MDR malignant diseases in the clinical setting.


Assuntos
Proteínas E1A de Adenovirus/genética , Resistência a Múltiplos Medicamentos , Terapia Genética/métodos , Replicação Viral/genética , Adenovírus Humanos/genética , Adenovírus Humanos/isolamento & purificação , Sequência de Bases , Linhagem Celular , Linhagem Celular Tumoral , Primers do DNA , DNA Complementar/genética , Deleção de Genes , Células HeLa , Humanos , Neoplasias/genética , Neoplasias/terapia , Transfecção/métodos
16.
J Biol Chem ; 278(30): 27988-96, 2003 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-12695516

RESUMO

Expression of the Y-box protein YB-1 is increased in proliferating normal and cancer cells, but its role in cell proliferation and cell cycle progression is unclear. We have identified a cell cycle-dependent relocalization of YB-1 from the cytoplasm to the nucleus at the G1/S phase transition and demonstrate that both the charged zipper and the cold shock domain are involved in regulating this process. Using cell lines that constitutively overexpress YB-1, we show that nuclear accumulation of YB-1 is associated with increased cyclin A and cyclin B1 mRNA and protein expression. We provide evidence that deregulated YB-1 expression is linked to adhesion-independent cell proliferation through the induction of cyclin A. Thus, we have identified YB-1 as a cell cycle stage-specific transcription factor important for cell proliferation.


Assuntos
Proteínas Estimuladoras de Ligação a CCAAT/fisiologia , Ciclina A/metabolismo , Ciclina B/metabolismo , Proteínas de Ligação a DNA , Regulação Neoplásica da Expressão Gênica , Fatores de Transcrição/metabolismo , Motivos de Aminoácidos , Neoplasias da Mama/metabolismo , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Adesão Celular , Ciclo Celular , Divisão Celular , Linhagem Celular , Núcleo Celular/metabolismo , Separação Celular , Ciclina A/genética , Ciclina B/genética , Ciclina B1 , Citoplasma/metabolismo , Citometria de Fluxo , Fase G1 , Genes Reporter , Células HeLa , Humanos , Immunoblotting , Imuno-Histoquímica , Cinética , Microscopia de Fluorescência , Mutação , Fatores de Transcrição NFI , Proteínas Nucleares , Regiões Promotoras Genéticas , Estrutura Terciária de Proteína , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fase S , Frações Subcelulares/metabolismo , Transfecção , Proteína 1 de Ligação a Y-Box
17.
J Biol Chem ; 277(12): 10427-34, 2002 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-11788582

RESUMO

The adenovirus early proteins E1A and E1B-55kDa are key regulators of viral DNA replication, and it was thought that targeting of p53 by E1B-55kDa is essential for this process. Here we have identified a previously unrecognized function of E1B for adenovirus replication. We found that E1B-55kDa is involved in targeting the transcription factor YB-1 to the nuclei of adenovirus type 5-infected cells where it is associated with viral inclusion bodies believed to be sites of viral transcription and replication. We show that YB-1 facilitates E2 gene expression through the E2 late promoter thus controlling E2 gene activity at later stages of infection. The role of YB-1 for adenovirus replication was demonstrated with an E1-minus adenovirus vector containing a YB-1 transgene. In infected cells, AdYB-1 efficiently replicated and produced infectious progeny particles. Thus, adenovirus E1B-55kDa protein and the host cell factor YB-1 act jointly to facilitate adenovirus replication in the late phase of infection.


Assuntos
Adenoviridae/metabolismo , Proteínas E2 de Adenovirus/genética , Antígenos de Bactérias , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Núcleo Celular/metabolismo , Proteínas de Ligação a DNA , Regiões Promotoras Genéticas , Fatores de Transcrição , Transporte Ativo do Núcleo Celular , Adenoviridae/genética , Proteínas de Bactérias/metabolismo , Northern Blotting , Southern Blotting , Western Blotting , Linhagem Celular , DNA Complementar/metabolismo , Regulação da Expressão Gênica , Células HeLa , Humanos , Microscopia Confocal , Microscopia Eletrônica , Microscopia de Fluorescência , Fatores de Transcrição NFI , Proteínas Nucleares , Ligação Proteica , Transgenes , Replicação Viral , Proteína 1 de Ligação a Y-Box
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