Transformation of Arabidopsis thaliana with heat stress-related genes from a Copaifera officinalis expressed sequence tag library

This dissertation has three components: (1) discovery of genes involved in heat stress-related processes derived from a tropical plant known as the diesel tree (C. officinalis) by means of an expressed sequence tag (EST) library, (2) stable transformation of Arabidopsis thaliana with C. officinalis heat stress-related genes and (3) assessing the degree to which C. officinalis heat-stress associated genes provide thermotolerance in transgenic A. thaliana. This dissertation project has important plant biotechnological components as well as agriculturally important themes. Global climate change is predicted to result in elevated temperatures over the next century and so studying the influence of heat stress upon plants is becoming increasingly important. Subsequently, in this work, it was hypothesized that heat stress-related transgenes derived from C. officinalis would provide some level of thermotolerance to A. thaliana. Various stress assays, which included both in vitro and in vivo assays, were performed to help determine the degree of thermotolerance in each transgenic line. A novel and key component in this project is that for the EST library C. officinalis cDNA was ligated into a plant binary vector, which eliminated the need to perform individual ligation reactions for each gene of interest (GOI). Sequencing plates were obtained and stored at -80C so that individual wells containing the E. coli harboring the binary vector could be picked and grown overnight for a subsequent plasmid miniprep. Plasmids were transfered into Agrobacterium tumefaciens for generation transgenic lines. This created a streamlined and highthroughput method of generating multiple transgenic plants, each with a unique GOI. Although others have overexpressed heat stress-related genes in A. thaliana, this project overexpressed C. officinialis plant genes, which is novel. The results suggest that overexpression of C. officinalis heat stress-related genes in A. thaliana help confer thermotolerance. Furthermore, the library clones obtained can be investigated in future studies, which offers a rich resource for transgenic studies

In silico analysis of terpene synthase genes in Arabidopsis thaliana

Terpenes are defense chemicals found in wide groups of plants. Terpenoids play a large role in plant development and stress response. The terpene synthase family comprises a diverse set of genes, all which contribute to production of terpenoids. We have used tools of bioinformatics and performed an in silico analysis of developmental and tissue specific terpene synthase gene expression in Arabidopsis thaliana, as well as those expressed due to biotic and abiotic environmental stimuli. Using software tools from Genevestigator, a powerful microarray analyzer, we used multiple tool sets to better understand terpene synthase expression in Arabidopsis, which will hopefully open the genetic door to further wet laboratory investigations. The data can be used to predict roles of terpene synthase genes in plant cell division and growth. The data presented here can be used to model for terpene synthesis expression in other plant species and can also be used to integrate basic plant physiology, and ‘omics’ disciplines

In Silico Analysis of Terpene Synthase Genes in Arabidopsis Thaliana

Terpenes are defense chemicals found in wide groups of plants. Terpenoids play a large role in plant development and stress response. The terpene synthase family comprises a diverse set of genes, all which contribute to production of terpenoids. We have used tools of bioinformatics and performed an in silico analysis of developmental and tissue specific terpene synthase gene expression in Arabidopsis thaliana, as well as those expressed due to biotic and abiotic environmental stimuli. Using software tools from Genevestigator, a powerful microarray analyzer, we used multiple tool sets to better understand terpene synthase expression in Arabidopsis, which will hopefully open the genetic door to further wet laboratory investigations. The data can be used to predict roles of terpene synthase genes in plant cell division and growth. The data presented here can be used to model for terpene synthesis expression in other plant species and can also be used to integrate basic plant physiology, and ‘omics’ disciplines

Die Kaschmirfrage: Nachbeben der Diplomatie: indisch-pakistanische Annäherung - neue Qualität in den bilateralen Beziehungen?

Inhalt: Annäherung im Zeichen der Katastrophe; Dimensionen der Kaschmirproblematik; Wurzeln der Gewalt; Radikalisierung des Konflikts in den 1990er Jahren; Von der Eskalation zur Entspannung: Der "Kampf gegen den Terrorismus"; Dialog und Annäherung: Erste Gehversuche; Politisches Kalkül und Skepsis bestehen fort; Das Erdbeben und seine Folgen: Chance und Herausforderung zugleich; Fazit: Hoffnung auf weitere Annäherung

Estudio de las mucinas y de antígenos carbohidratos asociados al cáncer colorrectal

El cáncer colorrectal (CCR) es la tercera causa de muerte por cáncer en la Argentina. Las mucinas y sus antígenos carbohidratos asociados han sido relacionados con la proliferación, invasión, metástasis y mal pronóstico. Objetivo: Estudiar la utilidad de MUC1, MUC2, MUC5AC y los antígenos carbohidratos asociados como marcadores tumorales pronósticos y predictivos de respuesta al tratamiento en pacientes con CCR. Materiales: Se incluyeron 248 muestras histológicas: 90 CCR, 64 ganglios linfáticos, 68 muestras de mucosa colorrectal normal, 26 adenomas y 120 muestras sanguíneas. Métodos: Se empleó la técnica de inmunohistoquímica en los tejidos tumorales y radioinmunoanálisis en las sanguíneas. Se utilizó un índice de expresión antigénica (intensidad por el porcentaje de la reacción). Los pacientes fueron estadificados según el TNM (AJCC, edición 2002). Resultados: Independientemente del estadío y del tratamiento instaurado, los pacientes tuvieron una menor sobrevida global (SG) y tiempo libre de progresión (TLP) cuando sus tumores primarios (TP) o ganglios linfáticos metastáticos (GLM) expresaron MUC1, Tn, Lewis x/y, sialil Lewis x y cuando en el núcleo de las células tumorales se halló presente β-catenina. Los pT avanzados se asociaron con mayor expresión de MUC1 y MUC5AC. Estos tumores presentaron mayor expresión de sialil Lewis a/x y Lewis y en sus GLM. El estadío pN2 se asoció con mayor expresión de sialil Lewis a/x. Se observó una mayor expresión de MUC1 y sialil Lewis a/x en los TP de los pacientes con M1. Los TP indiferenciados presentaron una elevada expresión de MUC1 y MUC5AC y una disminución de MUC2, los antígenos del grupo Lewis sialil y CEA. Los TP grado nuclear 3 tuvieron mayor expresión de MUC5AC, Tn, Lewis x y CEA. Un elevado índice mitótico en los TP se correlacionó con una alta expresión de MUC1 y CEA y una baja expresión de MUC2 y MUC5AC. Los GLM con alto índice mitótico expresaron en su mayoría Lewis x/y y sialil Lewis x. MUC1 se halló asociada a la invasión linfática y perineural tumoral. En esta última también se encontró elevada MUC5AC, sialil Lewis a y CEA. Los tumores mayores de 5 cm presentaron una elevada expresión de sialil Lewis x y CEA. El aumento de la expresión de MUC1 se relacionó con cáncer familiar, diabetes mellitus, hábito alcohólico y tabáquico, hipertensión arterial, colecistectomía previa y una pérdida de peso mayor a 10 kg en los dos meses previos al diagnóstico. Se demostró una variación en la expresión de las mucinas y antígenos carbohidratos estudiados siguiendo la secuencia: mucosa normal, mucosa normal adyacente al TP y TP, pudiendo comprobarse que la mucosa normal adyacente al tumor es una zona de transición entre mucosa normal y TP. Conclusión: Las mucinas y los antígenos carbohidratos asociados constituyen parte de los cambios ocurridos tanto durante la carcinogénesis como en los procesos de invasión del CCR. Asimismo, constituirían un elemento a considerar para comprender las recaídas locales.Facultad de Ciencias Médica

Plant Terpenoids: Applications and Future Potentials

The importance of terpenes in both nature and human application is difficult to overstate. Basic knowledge of terpene and isoprene biosynthesis and chemistry has accelerated the pace at which scientists have come to understand many plant biochemical and metabolic processes. The abundance and diversity of terpene compounds in nature can have ecosystem-wide influences. Although terpenes have permeated human civilization since the Egyptians, terpene synthesis pathways are only now being understood in great detail. The use of bioinformatics and molecular databases has largely contributed to analyzing exactly how and when terpenes are synthesized. Additionally, terpene synthesis is beginning to be understood in respect to the various stages of plant development. Much of this knowledge has been contributed by the plant model, Arabidopsis thaliana. Considering the advances in plant terpene knowledge and potential uses, it is conceivable that they may soon be used in agrobiotechnology

Partial Characterization of Two Moderately Halophilic Bacteria from a Kansas Salt Marsh

Two bacterial species were isolated from a salt marsh located on privately owned land in Russell County, Kansas. Water samples from the saIt marsh were streaked for isolation on tryptic soy agar supplemented with 12 % NaCI. Visual scanning of the plates revealed two prominent colony types. The two colony types were subcultured repeatedly until axenic cultures were obtained. 80th of these organisms were shown to be moderately halophilic. The organisms were characterized partially by fatty acid methyl ester analysis, 16S rRNA sequencing, and scanning electron microscopy. These studies revealed that the bacteria previously were unreported members of genera Marinococcus and Halomonas

Potential therapeutic targets for growth arrest of colorectal cancer cells exposed to PTHrP

Although PTHrP is implicated in several cancers, its role in chemoresistance is not fully elucidated. We found that in CRC cells, PTHrP exerts proliferative and protective effects and induces cell migration. The aim of this work was to further study the effects of PTHrP in CRC cells. Herein we evidenced, for the first time, that PTHrP induces resistance to CPT-11 in Caco-2 and HCT116 cells; although both cell lines responded to the drug through different molecular mechanisms, the chemoresistance by PTHrP in these models is mediated through ERK, which in turn is activated by PCK, Src and Akt. Moreover, continue administration of PTHrP in nude mice xenografts increased the protein levels of this MAPK and of other markers related to tumorigenic events. The understanding of the molecular mechanisms leading to ERK 1/2 activation and the study of ERK targets may facilitate the development of new therapeutic strategies for CRC treatment.Facultad de Ciencias Veterinaria

Potential therapeutic targets for growth arrest of colorectal cancer cells exposed to PTHrP

Although PTHrP is implicated in several cancers, its role in chemoresistance is not fully elucidated. We found that in CRC cells, PTHrP exerts proliferative and protective effects and induces cell migration. The aim of this work was to further study the effects of PTHrP in CRC cells. Herein we evidenced, for the first time, that PTHrP induces resistance to CPT-11 in Caco-2 and HCT116 cells; although both cell lines responded to the drug through different molecular mechanisms, the chemoresistance by PTHrP in these models is mediated through ERK, which in turn is activated by PCK, Src and Akt. Moreover, continue administration of PTHrP in nude mice xenografts increased the protein levels of this MAPK and of other markers related to tumorigenic events. The understanding of the molecular mechanisms leading to ERK 1/2 activation and the study of ERK targets may facilitate the development of new therapeutic strategies for CRC treatment.Facultad de Ciencias Veterinaria

Potential therapeutic targets for growth arrest of colorectal cancer cells exposed to PTHrP

Although PTHrP is implicated in several cancers, its role in chemoresistance is not fully elucidated. We found that in CRC cells, PTHrP exerts proliferative and protective effects and induces cell migration. The aim of this work was to further study the effects of PTHrP in CRC cells. Herein we evidenced, for the first time, that PTHrP induces resistance to CPT-11 in Caco-2 and HCT116 cells; although both cell lines responded to the drug through different molecular mechanisms, the chemoresistance by PTHrP in these models is mediated through ERK, which in turn is activated by PCK, Src and Akt. Moreover, continue administration of PTHrP in nude mice xenografts increased the protein levels of this MAPK and of other markers related to tumorigenic events. The understanding of the molecular mechanisms leading to ERK 1/2 activation and the study of ERK targets may facilitate the development of new therapeutic strategies for CRC treatment.Fil: Martín, María Julia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Ciencias Biológicas y Biomédicas del Sur. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia. Instituto de Ciencias Biológicas y Biomédicas del Sur; ArgentinaFil: Gigola, Graciela. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia; ArgentinaFil: Zwenger, Ariel. Hospital Provincial de Neuquén; ArgentinaFil: Carriquiriborde, Martin. Universidad Nacional de La Plata; ArgentinaFil: Gentil, Florencia. Universidad Nacional de La Plata; ArgentinaFil: Gentili, Claudia Rosana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Ciencias Biológicas y Biomédicas del Sur. Universidad Nacional del Sur. Departamento de Biología, Bioquímica y Farmacia. Instituto de Ciencias Biológicas y Biomédicas del Sur; Argentin