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1.
Carcinogenesis ; 36(1): 122-32, 2015 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-25421723

RESUMO

Response to breast cancer chemoprevention can depend upon host genetic makeup and initiating events leading up to preneoplasia. Increased expression of aromatase and estrogen receptor (ER) is found in conjunction with breast cancer. To investigate response or resistance to endocrine therapy, mice with targeted overexpression of Esr1 or CYP19A1 to mammary epithelial cells were employed, representing two direct pathophysiological interventions in estrogen pathway signaling. Both Esr1 and CYP19A1 overexpressing mice responded to letrozole with reduced hyperplastic alveolar nodule prevalence and decreased mammary epithelial cell proliferation. CYP19A1 overexpressing mice were tamoxifen sensitive but Esr1 overexpressing mice were tamoxifen resistant. Increased ER expression occurred with tamoxifen resistance but no consistent changes in progesterone receptor, pSTAT3, pSTAT5, cyclin D1 or cyclin E levels in association with response or resistance were found. RNA-sequencing (RNA-seq) was employed to seek a transcriptome predictive of tamoxifen resistance using these models and a second tamoxifen-resistant model, BRCA1 deficient/Trp53 haploinsufficient mice. Sixty-eight genes associated with immune system processing were upregulated in tamoxifen-resistant Esr1- and Brca1-deficient mice, whereas genes related to aromatic compound metabolic process were upregulated in tamoxifen-sensitive CYP19A1 mice. Interferon regulatory factor 7 was identified as a key transcription factor regulating these 68 immune processing genes. Two loci encoding novel transcripts with high homology to human immunoglobulin lambda-like polypeptide 1 were uniquely upregulated in the tamoxifen-resistant models. Letrozole proved to be a successful alternative to tamoxifen. Further study of transcriptional changes associated with tamoxifen resistance including immune-related genes could expand our mechanistic understanding and lead to biomarkers predictive of escape or response to endocrine therapies.


Assuntos
Aromatase/metabolismo , Receptor alfa de Estrogênio/metabolismo , Fenômenos do Sistema Imunitário/genética , Neoplasias Mamárias Animais/tratamento farmacológico , Nitrilas/farmacologia , Lesões Pré-Cancerosas/tratamento farmacológico , Tamoxifeno/farmacologia , Triazóis/farmacologia , Animais , Antineoplásicos Hormonais/farmacologia , Inibidores da Aromatase/farmacologia , Proteína BRCA1/metabolismo , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Biomarcadores Tumorais/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Citometria de Fluxo , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Técnicas Imunoenzimáticas , Letrozol , Neoplasias Mamárias Animais/genética , Neoplasias Mamárias Animais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Análise de Sequência com Séries de Oligonucleotídeos , Lesões Pré-Cancerosas/genética , Lesões Pré-Cancerosas/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
2.
Arch Virol ; 156(5): 827-38, 2011 May.
Artigo em Inglês | MEDLINE | ID: mdl-21318310

RESUMO

Papillomavirus capsid proteins L1 and L2 mediate virion attachment, internalization and trafficking. In our studies of the capsid proteins, we identified an interaction of L2 with the E3 ligase Smad ubiquitin regulatory factor 2 (Smurf2). Smurf2 expression alters BPV1 virion trafficking and L2 protein levels. Using BPV1 pseudovirions (PSVs) containing a GFP or DSRed transgene encapsidated by L1 and L2 proteins, our data showed that although only BPV1 L2 interacts with Smurf2, both L1 and L2 levels decrease in a Smurf2- and ubiquitin-dependent manner. The decrease in L2 protein levels corresponded to a decrease in infection (i.e., loss of GFP or DSRed expression). We propose that Smurf2 regulates L2 protein cellular localization and therefore alters L2 protein levels. This change in trafficking and protein level decreases nuclear delivery and transcription of encapsidated pseudoviral transgenes and thus decreases BPV1 infection levels.


Assuntos
Papillomavirus Bovino 1/imunologia , Papillomavirus Bovino 1/patogenicidade , Proteínas do Capsídeo/metabolismo , Ubiquitina-Proteína Ligases/imunologia , Replicação Viral , Animais , Linhagem Celular , Chlorocebus aethiops , Humanos
3.
J Virol ; 83(16): 8221-32, 2009 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-19494002

RESUMO

Human papillomavirus type 16 (HPV16) has been identified as being the most common etiological agent leading to cervical cancer. Despite having a clear understanding of the role of HPV16 in oncogenesis, details of how HPV16 traffics during infection are poorly understood. HPV16 has been determined to enter via clathrin-mediated endocytosis, but the subsequent steps of HPV16 infection remain unclear. There is emerging evidence that several viruses take advantage of cross talk between routes of endocytosis. Specifically, JCV and bovine papillomavirus type 1 have been shown to enter cells by clathrin-dependent endocytosis and then require caveolin-1-mediated trafficking for infection. In this paper, we show that HPV16 is dependent on caveolin-1 after clathrin-mediated endocytosis. We provide evidence for the first time that HPV16 infection is dependent on trafficking to the endoplasmic reticulum (ER). This novel trafficking may explain the requirement for the caveolar pathway in HPV16 infection because clathrin-mediated endocytosis typically does not lead to the ER. Our data indicate that the infectious route for HPV16 following clathrin-mediated entry is caveolin-1 and COPI dependent. An understanding of the steps involved in HPV16 sorting and trafficking opens up the possibility of developing novel approaches to interfere with HPV16 infection and reduce the burden of papillomavirus diseases including cervical cancer.


Assuntos
Brefeldina A/farmacologia , Caveolina 1/metabolismo , Clatrina/metabolismo , Papillomavirus Humano 16/fisiologia , Queratinócitos/metabolismo , Infecções por Papillomavirus/metabolismo , Linhagem Celular , Endocitose , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/virologia , Papillomavirus Humano 16/efeitos dos fármacos , Papillomavirus Humano 16/genética , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/virologia , Infecções por Papillomavirus/virologia
4.
Virol J ; 6: 109, 2009 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-19619315

RESUMO

BACKGROUND: The infectious pathway of the non-enveloped Human Papillomavirus Type 16 (HPV16) includes binding to the cell surface, clathrin-mediated endocytosis, and penetration into an endosome. HPV16 infection was shown to decrease in the presence of the lysosomotrophic neutralizing agent ammonium chloride (NH4Cl). NH4Cl neutralizes acidic endo-lysosome compartments, thus suggesting that pH was responsible for PV capsid conformational changes leading endosome escape. RESULTS: However, our data suggested that NH4Cl blocked infection by preventing the movement of PV viral particles from the early endosome to the caveosome as was shown for JC virus 12. We have confirmed that HPV 16 infection requires the trafficking of reporter-virions to the caveosome as is the case for BPV1 34. In this manuscript we propose that the observed decrease in infection of PV in the presence of NH4Cl was due to a loss of movement of reporter-virions to caveosomes. We also demonstrate that cysteine proteases are involved in the infectious process, and that cathepsin B treatment of viral particles was shown to overcome the block of infection observed in the presence of furin inhibition. We confirmed the need for cathepsin B in HPV16 infection using cathepsin B null mouse embryonic fibroblasts. CONCLUSION: We present data that suggest HPV16 infection is in part mediated by cysteine proteases, and that NH4Cl blocks the intracellular trafficking of infectious viral particles. To our knowledge this is the first demonstration that cysteine proteases influence the infection of a non-enveloped virus.


Assuntos
Cloreto de Amônio/farmacologia , Antivirais/farmacologia , Cisteína Endopeptidases/metabolismo , Papillomavirus Humano 16/efeitos dos fármacos , Papillomavirus Humano 16/fisiologia , Internalização do Vírus , Animais , Linhagem Celular , Células Cultivadas , Endossomos/virologia , Interações Hospedeiro-Patógeno , Humanos , Camundongos , Camundongos Knockout
5.
Mol Biol Cell ; 30(14): 1770-1779, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31091168

RESUMO

Cell proliferation is essential for the development and maintenance of all organisms and is dysregulated in cancer. Using synchronized cells progressing through the cell cycle, pioneering microarray studies defined cell cycle genes based on cyclic variation in their expression. However, the concordance of the small number of synchronized cell studies has been limited, leading to discrepancies in definition of the transcriptionally regulated set of cell cycle genes within and between species. Here we present an informatics approach based on Boolean logic to identify cell cycle genes. This approach used the vast array of publicly available gene expression data sets to query similarity to CCNB1, which encodes the cyclin subunit of the Cdk1-cyclin B complex that triggers the G2-to-M transition. In addition to highlighting conservation of cell cycle genes across large evolutionary distances, this approach identified contexts where well-studied genes known to act during the cell cycle are expressed and potentially acting in nondivision contexts. An accessible web platform enables a detailed exploration of the cell cycle gene lists generated using the Boolean logic approach. The methods employed are straightforward to extend to processes other than the cell cycle.


Assuntos
Ciclo Celular/genética , Bases de Dados Genéticas , Regulação da Expressão Gênica , Animais , Drosophila/genética , Humanos , Camundongos , Plantas/genética
6.
Endocr Relat Cancer ; 21(3): R195-208, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24481326

RESUMO

The majority of human breast cancers are estrogen receptor-positive (ER+), but this has proven challenging to model in genetically engineered mice. This review summarizes information on 21 mouse models that develop ER+ mammary cancer. Where available, information on cancer pathology and gene expression profiles is referenced to assist in understanding which histological subtype of ER+ human cancer each model might represent. ESR1, CCDN1, prolactin, TGFα, AIB1, ESPL1, and WNT1 overexpression, PIK3CA gain of function, as well as loss of P53 (Trp53) or STAT1 are associated with ER+ mammary cancer. Treatment with the PPARγ agonist efatutazone in a mouse with Brca1 and p53 deficiency and 7,12-dimethylbenz(a)anthracene exposure in combination with an activated myristoylated form of AKT1 also induce ER+ mammary cancer. A spontaneous mutant in nude mice that develops metastatic ER+ mammary cancer is included. Age of cancer development ranges from 3 to 26 months and the percentage of cancers that are ER+ vary from 21 to 100%. Not all models are characterized as to their estrogen dependency and/or response to anti-hormonal therapy. Strain backgrounds include C57Bl/6, FVB, BALB/c, 129S6/SvEv, CB6F1, and NIH nude. Most models have only been studied on one strain background. In summary, while a range of models are available for studies of pathogenesis and therapy of ER+ breast cancers, many could benefit from further characterization, and opportunity for development of new models remains.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/prevenção & controle , Modelos Animais de Doenças , Receptor alfa de Estrogênio/metabolismo , Animais , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos
7.
Endocr Relat Cancer ; 21(3): 443-57, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24692510

RESUMO

Transformation-related protein 63 (Trp63), the predominant member of the Trp53 family, contributes to epithelial differentiation and is expressed in breast neoplasia. Trp63 features two distinct promoters yielding specific mRNAs encoding two major TRP63 isoforms, a transactivating transcription factor and a dominant negative isoform. Specific TRP63 isoforms are linked to cell cycle arrest, apoptosis, survival, and epithelial mesenchymal transition (EMT). Although TRP63 overexpression in cultured cells is used to elucidate functions, little is known about Trp63 regulation in normal and cancerous mammary tissues. This study used ChIP-seq to interrogate transcription factor binding and histone modifications of the Trp63 locus in mammary tissue and RNA-seq and immunohistochemistry to gauge gene expression. H3K4me2 and H3K4me3 marks coincided only with the proximal promoter, supporting RNA-seq data showing the predominance of the dominant negative isoform. STAT5 bound specifically to the Trp63 proximal promoter and Trp63 mRNA levels were elevated upon deleting Stat5 from mammary tissue, suggesting its role as a negative regulator. The dominant negative TRP63 isoform was localized to nuclei of basal mammary epithelial cells throughout reproductive cycles and retained in a majority of the triple-negative cancers generated from loss of full-length Brca1. Increased expression of dominant negative isoforms was correlated with developmental windows of increased progesterone receptor binding to the proximal Trp63 promoter and decreased expression during lactation was correlated with STAT5 binding to the same region. TRP63 is present in the majority of triple-negative cancers resulting from loss of Brca1 but diminished in less differentiated cancer subtypes and in cancer cells undergoing EMT.


Assuntos
Diferenciação Celular , Glândulas Mamárias Animais/crescimento & desenvolvimento , Glândulas Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/patologia , Fosfoproteínas/fisiologia , Fator de Transcrição STAT5/metabolismo , Transativadores/fisiologia , Animais , Proteína BRCA1/fisiologia , Western Blotting , Células Cultivadas , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Histonas/metabolismo , Técnicas Imunoenzimáticas , Camundongos , Camundongos Endogâmicos C57BL , Isoformas de Proteínas , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reprodução , Reação em Cadeia da Polimerase Via Transcriptase Reversa
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