{"id":117955,"date":"2018-03-11T10:47:44","date_gmt":"2018-03-11T10:47:44","guid":{"rendered":"https:\/\/www.deberes.net\/tesis\/sin-categoria\/interaccion-mnsv-planta-de-melon-una-aproximacion-transcriptomica-y-celular\/"},"modified":"2018-03-11T10:47:44","modified_gmt":"2018-03-11T10:47:44","slug":"interaccion-mnsv-planta-de-melon-una-aproximacion-transcriptomica-y-celular","status":"publish","type":"post","link":"https:\/\/www.deberes.net\/tesis\/fisiologia-vegetal\/interaccion-mnsv-planta-de-melon-una-aproximacion-transcriptomica-y-celular\/","title":{"rendered":"Interacci\u00f3n mnsv-planta de mel\u00f3n. una aproximaci\u00f3n transcript\u00f3mica y celular"},"content":{"rendered":"<h2>Tesis doctoral de <strong> Cristina Gomez Aix <\/strong><\/h2>\n<p>Resumen el virus de las manchas necr\u00f3ticas del mel\u00f3n (melon necrotic spot virus, mnsv; g\u00e9nero carmovirus; familia tombusviridae) es un virus de arn de sentido positivo end\u00e9mico en cultivos de mel\u00f3n. Este virus ha sido empleado como modelo de estudio de aspectos relacionados con la superaci\u00f3n de resistencias recesivas, la traducci\u00f3n de arns virales carentes de estructuras cap o el movimiento intercelular. Sin embargo, se sabe poco acerca de las respuestas que la infecci\u00f3n por este virus induce en las plantas o los genes alterados como consecuencia del desarrollo de su ciclo viral en su hu\u00e9sped natural, el mel\u00f3n. As\u00ed mismo, se desconocen los lugares, dentro de la c\u00e9lula vegetal, donde este virus lleva a cabo su replicaci\u00f3n viral. Todos estos procesos podr\u00edan aportar conocimiento sobre los genes necesarios en la interacci\u00f3n planta-virus que permiten el establecimiento de una infecci\u00f3n satisfactoria por parte del virus, o los factores necesarios para la puesta en marcha de una respuesta de defensa eficaz para contrarrestar la infecci\u00f3n y que podr\u00edan ser utilizados en estrategias antivirales. Con el objetivo de ahondar en el conocimiento de la interacci\u00f3n mnsv-mel\u00f3n, esta tesis ha sido estructurada en dos cap\u00edtulos. en el primer cap\u00edtulo, utilizando los microarrays como herramienta principal de an\u00e1lisis, se ha llevado a cabo un estudio de las alteraciones transcript\u00f3micas inducidas por mnsv en mel\u00f3n, con especial \u00e9nfasis en aquellos cambios asociados con: (i) una regi\u00f3n no codificante del genoma del virus, (ii) diferentes cultivares de mel\u00f3n (iii) diferentes tejidos de la planta, (iv) y con el desarrollo de infecciones de tipo local o sist\u00e9mico. Los resultados obtenidos han permitido identificar la desregulaci\u00f3n espec\u00edfica de dos grupos de genes asociados con una regi\u00f3n no traducible del genoma viral, as\u00ed como genes que responden de forma espec\u00edfica de cultivar. La comparaci\u00f3n entre tejidos ha mostrado una respuesta cuantitativa diferente entre hoja y cotiled\u00f3n, pero cualitativamente muy similar, aportando validez a los resultados obtenidos en cotiled\u00f3n. As\u00ed mismo, una aproximaci\u00f3n tradicional al estudio de la posible activaci\u00f3n de una respuesta sist\u00e9mica adquirida por parte de la planta, ha descartado la activaci\u00f3n de la misma. Finalmente, la comparaci\u00f3n entre estos resultados con los obtenidos en trabajos previos para otros virus, ha permitido identificar un s\u00f3lo gen compartido por los tres virus analizados, que podr\u00eda ser importante en las infecciones virales al menos en mel\u00f3n. Por otro lado, estos an\u00e1lisis han puesto de manifiesto la importancia de utilizar diferentes tiempos de muestreo a la hora de realizar comparaciones entre los cambios transcript\u00f3micos inducidos por diferentes virus. en el segundo cap\u00edtulo, se ha llevado a cabo el an\u00e1lisis tisular de las lesiones inducidas por mnsv para delimitar las regiones de inter\u00e9s donde realizar los an\u00e1lisis ultraestructurales. Este an\u00e1lisis identific\u00f3 una regi\u00f3n id\u00f3nea para tales efectos. El estudio ultraestructural mediante microscop\u00eda electr\u00f3nica de transmisi\u00f3n de la regi\u00f3n identificada como z2, ha puesto de manifiesto la existencia de grandes org\u00e1nulos originados como consecuencia de la infecci\u00f3n por mnsv. Estos org\u00e1nulos han sido identificados como mitocondrias modificadas estructuralmente y representan los lugares donde el virus lleva a cabo su replicaci\u00f3n ya que, a trav\u00e9s de experimentos de hibridaci\u00f3n in situ e inmunocitoqu\u00edmica, se han localizado los arns virales, la prote\u00edna de la c\u00e1psida viral (cp) as\u00ed como el dsrna (intermediario de replicaci\u00f3n) en estos org\u00e1nulos. La reconstrucci\u00f3n tridimensional de las mitocondrias modificadas mostr\u00f3 la presencia de grandes dilataciones internas, interconectadas entre ellas y con el citoplasma circundante a trav\u00e9s de poros y\/o estructuras complejas as\u00ed como su conexi\u00f3n con cuerpos lip\u00eddicos. As\u00ed mismo, se llev\u00f3 a cabo el estudio de la implicaci\u00f3n de la prote\u00edna p29 de mnsv en la localizaci\u00f3n y generaci\u00f3n de los sitios de replicaci\u00f3n de mnsv. Para ello se generaron construcciones de la prote\u00edna p29 fusionada a gfp que fueron localizadas mediante microscop\u00eda laser confocal en mitocondrias. El an\u00e1lisis de la ultraestructura de las c\u00e9lulas que expresaron la prote\u00edna de fusi\u00f3n p29-gfp y su localizaci\u00f3n por inmunocitoqu\u00edmica identific\u00f3 la presencia de esta prote\u00edna en mitocondrias, as\u00ed como la inducci\u00f3n de modificaciones estructurales en estos org\u00e1nulos muy semejantes a las inducidas por el virus.    summary melon necrotic spot virus (mnsv) (genus carmovirus, family tombusviridae) is a single-stranded, positive-sense rna virus endemic of cucurbit crops worldwide. This virus has become an experimental model for the analysis of cell-to-cell virus movement and translation of uncapped viral rnas, whereas little is known about its replication or the transcriptome changes induced in melon plant by the virus during the viral cycle and in response to the infection. So far it is unknown the cellular compartment in which this virus carries out its viral replication. Addressing all these processes could provide information about genes involved in the plant-virus interaction that lead to a successful viral infection and the cellular factors involved in the defense response to counteract the infection. This knowledge may be used in future antiviral strategies. To clarify all these questions about mnsv-melon interaction this thesis has been structured into two chapters.  the first chapter is focused on transcriptomic alterations induced by mnsv in melon plants, with special emphasis on those changes associated with: (i) a non-coding region of the genome virus, (ii) different melon cultivars, (iii) different plant tissues, and, (iv) development of local or systemic infections. Microarray analyses have led to identify a specific deregulation of two groups of genes associated with an untranslated region of the viral genome, and also several sets of genes that respond specifically in each cultivar. The comparison between tissues has shown a different quantitative response between leaf and cotyledon, but qualitatively similar, providing validity to the results obtained in cotyledon. Also, a traditional approach to the study of the putative activation of a systemic acquired response from the plant has dismissed the activation of such response. Finally, the comparison of these results with those obtained in previous work for other viruses, has identified one gene shared by all the three viruses tested. This gene could play a key role in melon viralinfection. On the other hand, these results have shown the need of using different sampling times when comparing transcriptome changes induced by different viruses. in the second chapter, an histological analysis of the mnsv infected tissues was performed in order to define the most adequate region where to focuses the ultrastructural analysis. This analysis identified an ideal region for this purpose. The ultrastructural study by transmission electron microscopy of the region identified as z2, has revealed the existence of large organelles originated as a result of mnsv infection. Immunolocalization of the glycine decarboxylase complex (gdc) p protein in these organelles confirmed their mitochondrial origin. In situ hybridization and immunolocalization experiments showed the specific localization of positive-sense viral rna, capsid protein (cp), and double-stranded (ds)rna (a replication intermediate) in these organelles meaning that replication of the virus takes place in association with them. The three-dimensional reconstructions of the altered mitochondria showed the presence of large, interconnected, internal dilations which appeared to be linked to the outside cytoplasmic environment through pores and\/or complex structures, and with lipid bodies. Transient expression of mnsv p29 revealed that its specific target is mitochondria. Our data document the extensive reorganization of host mitochondria induced by mnsv, which provides a protected environment to viral replication, and show that the mnsv p29 protein is the primary determinant of this effect in the host.<\/p>\n<p>&nbsp;<\/p>\n<h3>Datos acad\u00e9micos de la tesis doctoral \u00ab<strong>Interacci\u00f3n mnsv-planta de mel\u00f3n. una aproximaci\u00f3n transcript\u00f3mica y celular<\/strong>\u00ab<\/h3>\n<ul>\n<li><strong>T\u00edtulo de la tesis:<\/strong>\u00a0 Interacci\u00f3n mnsv-planta de mel\u00f3n. una aproximaci\u00f3n transcript\u00f3mica y celular <\/li>\n<li><strong>Autor:<\/strong>\u00a0 Cristina Gomez Aix <\/li>\n<li><strong>Universidad:<\/strong>\u00a0 Murcia<\/li>\n<li><strong>Fecha de lectura de la tesis:<\/strong>\u00a0 24\/07\/2015<\/li>\n<\/ul>\n<p>&nbsp;<\/p>\n<h3>Direcci\u00f3n y tribunal<\/h3>\n<ul>\n<li><strong>Director de la tesis<\/strong>\n<ul>\n<li>Miguel Angel Aranda Regules<\/li>\n<\/ul>\n<\/li>\n<li><strong>Tribunal<\/strong>\n<ul>\n<li>Presidente del tribunal: vicente Pall\u00e1s benet <\/li>\n<li>cristina Risco ortiz (vocal)<\/li>\n<li>  (vocal)<\/li>\n<li>  (vocal)<\/li>\n<\/ul>\n<\/li>\n<\/ul>\n<p>&nbsp;<\/p>\n","protected":false},"excerpt":{"rendered":"<p>Tesis doctoral de Cristina Gomez Aix Resumen el virus de las manchas necr\u00f3ticas del mel\u00f3n (melon necrotic spot virus, mnsv; [&hellip;]<\/p>\n","protected":false},"author":1,"featured_media":0,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"site-sidebar-layout":"default","site-content-layout":"","ast-site-content-layout":"","site-content-style":"default","site-sidebar-style":"default","ast-global-header-display":"","ast-banner-title-visibility":"","ast-main-header-display":"","ast-hfb-above-header-display":"","ast-hfb-below-header-display":"","ast-hfb-mobile-header-display":"","site-post-title":"","ast-breadcrumbs-content":"","ast-featured-img":"","footer-sml-layout":"","theme-transparent-header-meta":"","adv-header-id-meta":"","stick-header-meta":"","header-above-stick-meta":"","header-main-stick-meta":"","header-below-stick-meta":"","astra-migrate-meta-layouts":"default","ast-page-background-enabled":"default","ast-page-background-meta":{"desktop":{"background-color":"var(--ast-global-color-4)","background-image":"","background-repeat":"repeat","background-position":"center center","background-size":"auto","background-attachment":"scroll","background-type":"","background-media":"","overlay-type":"","overlay-color":"","overlay-gradient":""},"tablet":{"background-color":"","background-image":"","background-repeat":"repeat","background-position":"center center","background-size":"auto","background-attachment":"scroll","background-type":"","background-media":"","overlay-type":"","overlay-color":"","overlay-gradient":""},"mobile":{"background-color":"","background-image":"","background-repeat":"repeat","background-position":"center center","background-size":"auto","background-attachment":"scroll","background-type":"","background-media":"","overlay-type":"","overlay-color":"","overlay-gradient":""}},"ast-content-background-meta":{"desktop":{"background-color":"var(--ast-global-color-5)","background-image":"","background-repeat":"repeat","background-position":"center center","background-size":"auto","background-attachment":"scroll","background-type":"","background-media":"","overlay-type":"","overlay-color":"","overlay-gradient":""},"tablet":{"background-color":"var(--ast-global-color-5)","background-image":"","background-repeat":"repeat","background-position":"center center","background-size":"auto","background-attachment":"scroll","background-type":"","background-media":"","overlay-type":"","overlay-color":"","overlay-gradient":""},"mobile":{"background-color":"var(--ast-global-color-5)","background-image":"","background-repeat":"repeat","background-position":"center center","background-size":"auto","background-attachment":"scroll","background-type":"","background-media":"","overlay-type":"","overlay-color":"","overlay-gradient":""}},"footnotes":""},"categories":[849,36483,37372,479,8235],"tags":[232145,73507,191225,8398],"class_list":["post-117955","post","type-post","status-publish","format-standard","hentry","category-biologia-celular","category-biologia-molecular","category-biologia-molecular-de-plantas","category-fisiologia-vegetal","category-murcia","tag-cristina-gomez-aix","tag-cristina-risco-ortiz","tag-miguel-angel-aranda-regules","tag-vicente-pallas-benet"],"_links":{"self":[{"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/posts\/117955","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/users\/1"}],"replies":[{"embeddable":true,"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/comments?post=117955"}],"version-history":[{"count":0,"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/posts\/117955\/revisions"}],"wp:attachment":[{"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/media?parent=117955"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/categories?post=117955"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/www.deberes.net\/tesis\/wp-json\/wp\/v2\/tags?post=117955"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}