Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia

Detalhes bibliográficos
Autor(a) principal: Patricia Nogueira da Gama Silva
Data de Publicação: 2005
Tipo de documento: Tese
Idioma: por
Título da fonte: Repositório Institucional da UFMG
Texto Completo: http://hdl.handle.net/1843/42545
Resumo: Poxviruses are viruses that present a linear, double-stranded DNA genome and multiply exclusively in the cell cytoplasm, in both, vertebrates and invertebrates. The orthopoxvirus Vaccinia virus (VV), is the prototype of the Poxviridae family, having capacity to encode more than 200 gene products. MAPKs (mitogen activated protein kinases) are important mediators of signal transduction and have a crucial role in the regulation of several cellular functions such as growth, proliferation, differentiation and apoptosis. Due to this, several viruses, as part of their multiplication strategy, regulate the activation of different MAPKs, e.g., the hepatitis B virus, the coxsackievirus B3, the Influenza virus, the Epstein Barr virus and the VV, among others. One of the consequences of the MAPK signaling pathway activation, in particularly Ras/Raf/MEK/ERK/Elk, by mitogenic stimuli, is the expression of immediate early genes, including c-fos, c-jun and the early growth response gene (egr-1). Andrade et al. (2004) demonstrated that this signaling pathway is also stimulated by VV, leading to Egr-1 expression and being necessary for its mutiplication. Since the analysis of Egr-1 perfomed above did not contemplate all the viral multiplication cycle, we decided to do a more detailed study of it. We characterized Egr-1 expression (mensage and protein) in response to viral infection and verified that it iniciates 1 hour after infection and goes on until late times of infection, aproximately 36 hours after infection. We also verified that it is dependent of the multiplicity of infection (m.o.i.), “de novo” protein synthesis and of early expression of viral genes, and results of simultaneous stimulation of the signaling pathways involving MEK/ERK and serine-threonine kinases (STK). We also generated, selected and characterized cellular clones stably expressing, MEK-1 negative dominance and we were able to demonstrate that MEK/ERK is functionaly relevant, not only for the egr-1 expression induced by VV, but also for viral multiplication and viral plaque fenotype. To verify the functional significance of Egr-1 for the biology of this orthopoxvirus, we generated, selected and characterized cellular clones stably expressing, siRNA for Egr 1. We could demonstrate that this cellular protein, whose expression is regulated by the virus since early times of infection until late ones, is localized preferentially in the nucleus and has a crucial role in the multiplication economy of VV. The description of one signaling pathway, leading to the expression of cellular protein that affects multiplication and viral plaque fenotype, explicits the necessity of interaction of this microorganism with its host. Altogether, the results presented here, has no precedent in the literature of poxviruses.
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spelling Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus VacciniaPoxvírusOrthopoxvirusVaccíniaMicrobiologiaGenes PrecocesOrthopoxvirusVacciniaQuinases de Proteína Quinase Ativadas por MitógenoPoxviruses are viruses that present a linear, double-stranded DNA genome and multiply exclusively in the cell cytoplasm, in both, vertebrates and invertebrates. The orthopoxvirus Vaccinia virus (VV), is the prototype of the Poxviridae family, having capacity to encode more than 200 gene products. MAPKs (mitogen activated protein kinases) are important mediators of signal transduction and have a crucial role in the regulation of several cellular functions such as growth, proliferation, differentiation and apoptosis. Due to this, several viruses, as part of their multiplication strategy, regulate the activation of different MAPKs, e.g., the hepatitis B virus, the coxsackievirus B3, the Influenza virus, the Epstein Barr virus and the VV, among others. One of the consequences of the MAPK signaling pathway activation, in particularly Ras/Raf/MEK/ERK/Elk, by mitogenic stimuli, is the expression of immediate early genes, including c-fos, c-jun and the early growth response gene (egr-1). Andrade et al. (2004) demonstrated that this signaling pathway is also stimulated by VV, leading to Egr-1 expression and being necessary for its mutiplication. Since the analysis of Egr-1 perfomed above did not contemplate all the viral multiplication cycle, we decided to do a more detailed study of it. We characterized Egr-1 expression (mensage and protein) in response to viral infection and verified that it iniciates 1 hour after infection and goes on until late times of infection, aproximately 36 hours after infection. We also verified that it is dependent of the multiplicity of infection (m.o.i.), “de novo” protein synthesis and of early expression of viral genes, and results of simultaneous stimulation of the signaling pathways involving MEK/ERK and serine-threonine kinases (STK). We also generated, selected and characterized cellular clones stably expressing, MEK-1 negative dominance and we were able to demonstrate that MEK/ERK is functionaly relevant, not only for the egr-1 expression induced by VV, but also for viral multiplication and viral plaque fenotype. To verify the functional significance of Egr-1 for the biology of this orthopoxvirus, we generated, selected and characterized cellular clones stably expressing, siRNA for Egr 1. We could demonstrate that this cellular protein, whose expression is regulated by the virus since early times of infection until late ones, is localized preferentially in the nucleus and has a crucial role in the multiplication economy of VV. The description of one signaling pathway, leading to the expression of cellular protein that affects multiplication and viral plaque fenotype, explicits the necessity of interaction of this microorganism with its host. Altogether, the results presented here, has no precedent in the literature of poxviruses.Os poxvírus são vírus que apresentam genoma DNA, dupla fita, linear e que se multiplicam exclusivamente no citoplasma de células, tanto de vertebrados, quanto de invertebrados. O orthopoxvirus Vaccínia (VV) é o protótipo da família Poxviridae, tendo a capacidade de codificar mais de 200 polipetídeos. As MAPKs (proteínas quinases ativadas por mitógenos) são importantes mediadoras de transdução de sinais e têm papel crucial na regulação de várias funções celulares como crescimento, proliferação, diferenciação e apoptose. Graças a isto, diversos vírus, como parte de suas estratégias multiplicativas, manipulam a ativação de diferentes MAPKs, como por exemplo: o vírus da hepatite B, o coxsackievirus B3, o vírus Influenza, o vírus Epstein Barr e o HIV, dentre outros. Uma das conseqüências da ativação da via sinalizadora MAPK, em particular Ras/Raf/MEK/ERK/Elk, por estímulos mitogênicos, é a expressão de genes imediatamente precoces, incluindo c-fos, c-jun e o gene de resposta precoce ao crescimento (egr-1). Andrade e colaboradores, 2004, demonstraram que esta via sinalizadora também é estimulada pelo VV, levando a expressão de Egr-1 e sendo necessária à sua multiplicação. Como a análise de Egr-1, realizada acima, não contemplava todo o ciclo de multiplicação viral, decidimos fazer um estudo mais detalhado da mesma. Caracterizamos a expressão de Egr-1 (mensagem e proteína) em resposta à infecção viral e verificamos que a mesma se inicia 1 hora pós-infecção e se prolonga até tempos tardios da infecção, isto é, aproximadamente, 36 horas pós-infecção. Verificamos também que a mesma é dependente da multiplicidade de infecção (m.o.i.), da síntese proteica “de novo” e da expressão de genes precoces virais e resulta da estimulação simultânea das vias sinalizadoras envolvendo MEK/ERK e serina-treonina quinases (STK). Também geramos, selecionamos e caracterizamos clones celulares expressando, de forma estável, dominância negativa para MEK-1 e pudemos demonstrar que MEK/ERK é funcionalmente relevante, tanto para a expressão de Egr-1 induzida pelo VV, quanto para a multiplicação e o fenótipo de placa de lise viral. Para verificar o significado funcional de Egr-1 para a biologia deste orthopoxvírus, geramos, selecionamos e caracterizamos, clones celulares expressando, de forma estável, siRNA para Egr-1. Pudemos demonstrar que esta proteína celular, cuja expressão é regulada pelo vírus desde os tempos precoces até tardios da infecção, e se localiza preferencialmente no núcleo, desempenha papel crucial na economia multiplicativa do VV. A descrição de uma via sinalizadora, levando a expressão de proteína celular que tenha reflexos, tanto na multiplicação, quanto no fenótipo da placa de lise viral, explicita a necessidade da interação deste microrganismo com seu hospedeiro. O conjunto de resultados apresentados nesta tese não tem precedentes na literatura dos poxvírus.Universidade Federal de Minas GeraisBrasilICB - DEPARTAMENTO DE MICROBIOLOGIAPrograma de Pós-Graduação em MicrobiologiaUFMGCláudio Antônio Bonjardimhttp://lattes.cnpq.br/9624031110564127Patricia Nogueira da Gama Silva2022-06-15T18:31:13Z2022-06-15T18:31:13Z2005-02-25info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/doctoralThesisapplication/pdfhttp://hdl.handle.net/1843/42545porinfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UFMGinstname:Universidade Federal de Minas Gerais (UFMG)instacron:UFMG2022-06-15T18:31:13Zoai:repositorio.ufmg.br:1843/42545Repositório InstitucionalPUBhttps://repositorio.ufmg.br/oairepositorio@ufmg.bropendoar:2022-06-15T18:31:13Repositório Institucional da UFMG - Universidade Federal de Minas Gerais (UFMG)false
dc.title.none.fl_str_mv Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia
title Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia
spellingShingle Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia
Patricia Nogueira da Gama Silva
Poxvírus
Orthopoxvirus
Vaccínia
Microbiologia
Genes Precoces
Orthopoxvirus
Vaccinia
Quinases de Proteína Quinase Ativadas por Mitógeno
title_short Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia
title_full Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia
title_fullStr Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia
title_full_unstemmed Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia
title_sort Significado funcional do gene de resposta precoce ao crescimento - EGR-1 - para a biologia do Orthopoxvírus Vaccinia
author Patricia Nogueira da Gama Silva
author_facet Patricia Nogueira da Gama Silva
author_role author
dc.contributor.none.fl_str_mv Cláudio Antônio Bonjardim
http://lattes.cnpq.br/9624031110564127
dc.contributor.author.fl_str_mv Patricia Nogueira da Gama Silva
dc.subject.por.fl_str_mv Poxvírus
Orthopoxvirus
Vaccínia
Microbiologia
Genes Precoces
Orthopoxvirus
Vaccinia
Quinases de Proteína Quinase Ativadas por Mitógeno
topic Poxvírus
Orthopoxvirus
Vaccínia
Microbiologia
Genes Precoces
Orthopoxvirus
Vaccinia
Quinases de Proteína Quinase Ativadas por Mitógeno
description Poxviruses are viruses that present a linear, double-stranded DNA genome and multiply exclusively in the cell cytoplasm, in both, vertebrates and invertebrates. The orthopoxvirus Vaccinia virus (VV), is the prototype of the Poxviridae family, having capacity to encode more than 200 gene products. MAPKs (mitogen activated protein kinases) are important mediators of signal transduction and have a crucial role in the regulation of several cellular functions such as growth, proliferation, differentiation and apoptosis. Due to this, several viruses, as part of their multiplication strategy, regulate the activation of different MAPKs, e.g., the hepatitis B virus, the coxsackievirus B3, the Influenza virus, the Epstein Barr virus and the VV, among others. One of the consequences of the MAPK signaling pathway activation, in particularly Ras/Raf/MEK/ERK/Elk, by mitogenic stimuli, is the expression of immediate early genes, including c-fos, c-jun and the early growth response gene (egr-1). Andrade et al. (2004) demonstrated that this signaling pathway is also stimulated by VV, leading to Egr-1 expression and being necessary for its mutiplication. Since the analysis of Egr-1 perfomed above did not contemplate all the viral multiplication cycle, we decided to do a more detailed study of it. We characterized Egr-1 expression (mensage and protein) in response to viral infection and verified that it iniciates 1 hour after infection and goes on until late times of infection, aproximately 36 hours after infection. We also verified that it is dependent of the multiplicity of infection (m.o.i.), “de novo” protein synthesis and of early expression of viral genes, and results of simultaneous stimulation of the signaling pathways involving MEK/ERK and serine-threonine kinases (STK). We also generated, selected and characterized cellular clones stably expressing, MEK-1 negative dominance and we were able to demonstrate that MEK/ERK is functionaly relevant, not only for the egr-1 expression induced by VV, but also for viral multiplication and viral plaque fenotype. To verify the functional significance of Egr-1 for the biology of this orthopoxvirus, we generated, selected and characterized cellular clones stably expressing, siRNA for Egr 1. We could demonstrate that this cellular protein, whose expression is regulated by the virus since early times of infection until late ones, is localized preferentially in the nucleus and has a crucial role in the multiplication economy of VV. The description of one signaling pathway, leading to the expression of cellular protein that affects multiplication and viral plaque fenotype, explicits the necessity of interaction of this microorganism with its host. Altogether, the results presented here, has no precedent in the literature of poxviruses.
publishDate 2005
dc.date.none.fl_str_mv 2005-02-25
2022-06-15T18:31:13Z
2022-06-15T18:31:13Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/doctoralThesis
format doctoralThesis
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://hdl.handle.net/1843/42545
url http://hdl.handle.net/1843/42545
dc.language.iso.fl_str_mv por
language por
dc.rights.driver.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv Universidade Federal de Minas Gerais
Brasil
ICB - DEPARTAMENTO DE MICROBIOLOGIA
Programa de Pós-Graduação em Microbiologia
UFMG
publisher.none.fl_str_mv Universidade Federal de Minas Gerais
Brasil
ICB - DEPARTAMENTO DE MICROBIOLOGIA
Programa de Pós-Graduação em Microbiologia
UFMG
dc.source.none.fl_str_mv reponame:Repositório Institucional da UFMG
instname:Universidade Federal de Minas Gerais (UFMG)
instacron:UFMG
instname_str Universidade Federal de Minas Gerais (UFMG)
instacron_str UFMG
institution UFMG
reponame_str Repositório Institucional da UFMG
collection Repositório Institucional da UFMG
repository.name.fl_str_mv Repositório Institucional da UFMG - Universidade Federal de Minas Gerais (UFMG)
repository.mail.fl_str_mv repositorio@ufmg.br
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