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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
71

Caracterização de um Novo Gene da Família F-box Expresso no Pistilo de Nicotiana tabacum L. / Characterization of a New F-box Family Gene Expressed in the Nicotiana tabacum L. Pistil

Abbad, Samantha Vieira 13 August 2012 (has links)
O estudo da reprodução sexual de plantas e uma área de crescente interesse devido a importância de sementes e frutos em nossa dieta diária, ambos resultantes do desenvolvimento de partes do pistilo, apos fertilização. O objetivo deste trabalho foi caracterizar um novo gene F-box expresso no pistilo de N. tabacum. Proteínas F-box atuam na interação proteína-proteína, geralmente direcionando proteínas alvo para degradação pela via ubiquitina-proteassomo. Foram identificados cinco genes de função desconhecida que codificam putativas proteínas F-box, em duas bibliotecas de cDNAs de estigmas/estiletes de N. tabacum (DEPAOLI, 2006; QUIAPIM et al., 2009) previamente construídas em nosso laboratório. A expressão de cada um destes genes foi analisada nos diferentes órgãos de N. tabacum, por qRT-PCR. O clone 085H05 da biblioteca TOBEST (QUIAPIM et al., 2009) apresentou expressão preferencial nos órgãos florais. Este clone foi selecionado para uma caracterização funcional mais detalhada. O padrão de expressão deste gene foi avaliado no estigma/estilete durante os 12 estádios do desenvolvimento floral de N. tabacum (KOLTUNOW et al., 1990). O resultado revelou que sua expressão e regulada durante o desenvolvimento, atingindo o maior nível de expressão na antese (estádio 12). Isto sugere que este gene esteja envolvido no desenvolvimento do estigma/estilete. A sequência codificadora do gene correspondente a 085H05 foi determinada e, apos amplificação e clonagem, este gene foi denominado S/S_F-box (Stigma/Style_F-box). Para compreender a função da proteína de S/S_F-box, plantas transgênicas de superexpressao e de silenciamento (por RNAi) deste gene foram geradas. As plantas de RNAi apresentaram o estilete e o ovário reduzidos quando comparados ao controle SR1. Em concordância, as plantas de superexpressao produziram flores com o estilete mais alongado do que o controle, alem do estigma e do ovário de maior tamanho. Altas concentrações de exudato foram observadas na superfície do estigma destas plantas, a partir do estádio 7 tardio. No controle SR1, concentrações equivalentes apenas são observadas nos estádios finais do desenvolvimento. Os fenótipos observados nas plantas transgênicas sugerem que a proteína codificada por S/S_F-box esteja envolvida com o desenvolvimento do pistilo e com o controle do tamanho deste órgão. Adicionalmente, as plantas de RNAi apresentaram o fenótipo de perda da dominância apical. Os níveis de expressão do gene S/S_F-box foram avaliados em plantas que tiveram aumento na produção de auxina no estigma/estilete (plantas STIG1prom::iaaM), revelando que este gene não e regulado, a nível transcricional, por este hormônio. Experimentos de localização subcelular, realizados por expressão transitória da sequência de S/S_F-box fusionada a sequência dos genes repórteres GFP e YFP (S/S_F-box::GFP; S/S_F-box::YFP), indicaram que a proteína S/S_F-box esta localizada no citoplasma e no núcleo celular. Adicionalmente, foi realizado o screening de uma biblioteca de cDNAs de estigma/estilete, construída no sistema de duplo-hibrido, para investigar proteínas candidatas a interagirem com a proteína de S/S_F-box. Os resultados indicaram interação da proteína S/S_F-box com SKP1, confirmando a participação de S/S_F-box no complexo SCF, que promove a degradação de proteínas alvo pela via ubiquitina-proteassomo. Duas proteínas candidatas a alvo foram identificadas: os fatores de transcrição VOZ1 e SIP1, ambos envolvidos com a proliferação celular. Em suma, e possível propor que a proteína codificada por S/S_F-box tenha função relacionada a proliferação celular e ao desenvolvimento dos órgãos vegetais, incluindo o pistilo. / The study of sexual reproduction in plants is an area of increasing interest due to the importance of seeds and fruits in our daily diet, both resulting from the development of parts of the pistil, after fertilization. The aim of this study was to characterize a new F-box gene expressed in the N. tabacum pistil. F-box proteins act in protein-protein interactions, generally directing target proteins to degradation via ubiquitin-proteasome. Five genes of unknown function coding for putative F-box proteins were identified at two cDNAs libraries from N. tabacum stigmas/styles (DEPAOLI, 2006; QUIAPIM et al., 2009), previously constructed in our laboratory. The expression of each of these genes was analyzed in the different N. tabacum organs, by qRT-PCR. The 085H05 clone from the TOBEST library (QUIAPIM et al., 2009) showed preferential expression in floral organs. This clone was select for a more detailed functional characterization. The expression pattern of this gene was evaluated in the stigma/style during the 12 N. tabacum flower developmental stages (KOLTUNOW et al., 1990). The result revealed that its expression is regulated during development, reaching the highest expression level at anthesis (stage 12). It suggests that this gene is involved in the stigma/style development. The coding sequence of the gene corresponding to 085H05 was determined and, after amplification and cloning, the gene was named S/S_F-box (Stigma/Style_F-box). To understand the S/S_F-box protein function, transgenic plants either overexpressing or silencing (by RNAi) the S/S_F-box gene were generated. The RNAi plants showed reduced style and ovary when compared to the control SR1. In accordance, the overexpressing plants produced flowers with a style more elongated than the control, besides an ovary and a stigma of larger size. High concentrations of exudate were observed on the stigma surface of these plants, since the later stage 7. In the control SR1, equivalent concentrations are only observed at the later stages of development. The phenotypes observed in the transgenic plants suggest that the protein encoded by S/S_F-box is involved with pistil development and with the control of pistil size. Additionally, the RNAi plants showed the phenotype of loss of apical dominance. The expression levels of the S/S_F-box gene were evaluated in plants with increased auxin production in the stigma/style (plants STIG1prom::iaaM), showing that this gene is not transcriptionally regulated by this hormone. Subcellular localization experiments, carried out by transient expression of the S/S_F-box sequence fused to the reporter genes GFP and YFP V (S/S_F-box::GFP; S/S_F-box::YFP), showed that the S/S_F-box protein is localized in the cytoplasm and in the nucleus. Additionally, the screening of a stigma/style cDNA library constructed on the yeast two hybrid system was performed, to investigate candidate proteins for S/S_F-box protein interaction. The results indicated interaction between S/S_Fbox and the SKP1 protein, confirming the involvement of the S/S_F-box protein in the SCF complex, which promotes degradation of target proteins via ubiquitin-proteasome. Two candidates for target proteins were identified: the transcription factors VOZ1 and SIP1, both involved in cell proliferation. In summary, it is possible to propose that the protein encoded by S/S_F-box has functions related to cell proliferation and organ development, including the pistil.
72

In silico analysis of mitochondrial proteins

Shen, Yaoqing 10 1900 (has links)
Le rôle important joué par la mitochondrie dans la cellule eucaryote est admis depuis longtemps. Cependant, la composition exacte des mitochondries, ainsi que les processus biologiques qui sy déroulent restent encore largement inconnus. Deux facteurs principaux permettent dexpliquer pourquoi létude des mitochondries progresse si lentement : le manque defficacité des méthodes didentification des protéines mitochondriales et le manque de précision dans lannotation de ces protéines. En conséquence, nous avons développé un nouvel outil informatique, YimLoc, qui permet de prédire avec succès les protéines mitochondriales à partir des séquences génomiques. Cet outil intègre plusieurs indicateurs existants, et sa performance est supérieure à celle des indicateurs considérés individuellement. Nous avons analysé environ 60 génomes fongiques avec YimLoc afin de lever la controverse concernant la localisation de la bêta-oxydation dans ces organismes. Contrairement à ce qui était généralement admis, nos résultats montrent que la plupart des groupes de Fungi possèdent une bêta-oxydation mitochondriale. Ce travail met également en évidence la diversité des processus de bêta-oxydation chez les champignons, en corrélation avec leur utilisation des acides gras comme source dénergie et de carbone. De plus, nous avons étudié le composant clef de la voie de bêta-oxydation mitochondriale, lacyl-CoA déshydrogénase (ACAD), dans 250 espèces, couvrant les 3 domaines de la vie, en combinant la prédiction de la localisation subcellulaire avec la classification en sous-familles et linférence phylogénétique. Notre étude suggère que les gènes ACAD font partie dune ancienne famille qui a adopté des stratégies évolutionnaires innovatrices afin de générer un large ensemble denzymes susceptibles dutiliser la plupart des acides gras et des acides aminés. Finalement, afin de permettre la prédiction de protéines mitochondriales à partir de données autres que les séquences génomiques, nous avons développé le logiciel TESTLoc qui utilise comme données des Expressed Sequence Tags (ESTs). La performance de TESTLoc est significativement supérieure à celle de tout autre outil de prédiction connu. En plus de fournir deux nouveaux outils de prédiction de la localisation subcellulaire utilisant différents types de données, nos travaux démontrent comment lassociation de la prédiction de la localisation subcellulaire à dautres méthodes danalyse in silico permet daméliorer la connaissance des protéines mitochondriales. De plus, ces travaux proposent des hypothèses claires et faciles à vérifier par des expériences, ce qui présente un grand potentiel pour faire progresser nos connaissances des métabolismes mitochondriaux. / The important role of mitochondria in the eukaryotic cell has long been appreciated, but their exact composition and the biological processes taking place in mitochondria are not yet fully understood. The two main factors that slow down the progress in this field are inefficient recognition and imprecise annotation of mitochondrial proteins. Therefore, we developed a new computational tool, YimLoc, which effectively predicts mitochondrial proteins from genomic sequences. This tool integrates the strengths of existing predictors and yields higher performance than any individual predictor. We applied YimLoc to ~60 fungal genomes in order to address the controversy about the localization of beta oxidation in these organisms. Our results show that in contrast to previous studies, most fungal groups do possess mitochondrial beta oxidation. This work also revealed the diversity of beta oxidation in fungi, which correlates with their utilization of fatty acids as energy and carbon sources. Further, we conducted an investigation of the key component of the mitochondrial beta oxidation pathway, the acyl-CoA dehydrogenase (ACAD). We combined subcellular localization prediction with subfamily classification and phylogenetic inference of ACAD enzymes from 250 species covering all three domains of life. Our study suggests that ACAD genes are an ancient family with innovative evolutionary strategies to generate a large enzyme toolset for utilizing most diverse fatty acids and amino acids. Finally, to enable the prediction of mitochondrial proteins from data beyond genome sequences, we designed the tool TESTLoc that uses expressed sequence tags (ESTs) as input. TESTLoc performs significantly better than known tools. In addition to providing two new tools for subcellular localization designed for different data, our studies demonstrate the power of combining subcellular localization prediction with other in silico analyses to gain insights into the function of mitochondrial proteins. Most importantly, this work proposes clear hypotheses that are easily testable, with great potential for advancing our knowledge of mitochondrial metabolism.
73

In silico analysis of mitochondrial proteins

Shen, Yaoqing 10 1900 (has links)
Le rôle important joué par la mitochondrie dans la cellule eucaryote est admis depuis longtemps. Cependant, la composition exacte des mitochondries, ainsi que les processus biologiques qui sy déroulent restent encore largement inconnus. Deux facteurs principaux permettent dexpliquer pourquoi létude des mitochondries progresse si lentement : le manque defficacité des méthodes didentification des protéines mitochondriales et le manque de précision dans lannotation de ces protéines. En conséquence, nous avons développé un nouvel outil informatique, YimLoc, qui permet de prédire avec succès les protéines mitochondriales à partir des séquences génomiques. Cet outil intègre plusieurs indicateurs existants, et sa performance est supérieure à celle des indicateurs considérés individuellement. Nous avons analysé environ 60 génomes fongiques avec YimLoc afin de lever la controverse concernant la localisation de la bêta-oxydation dans ces organismes. Contrairement à ce qui était généralement admis, nos résultats montrent que la plupart des groupes de Fungi possèdent une bêta-oxydation mitochondriale. Ce travail met également en évidence la diversité des processus de bêta-oxydation chez les champignons, en corrélation avec leur utilisation des acides gras comme source dénergie et de carbone. De plus, nous avons étudié le composant clef de la voie de bêta-oxydation mitochondriale, lacyl-CoA déshydrogénase (ACAD), dans 250 espèces, couvrant les 3 domaines de la vie, en combinant la prédiction de la localisation subcellulaire avec la classification en sous-familles et linférence phylogénétique. Notre étude suggère que les gènes ACAD font partie dune ancienne famille qui a adopté des stratégies évolutionnaires innovatrices afin de générer un large ensemble denzymes susceptibles dutiliser la plupart des acides gras et des acides aminés. Finalement, afin de permettre la prédiction de protéines mitochondriales à partir de données autres que les séquences génomiques, nous avons développé le logiciel TESTLoc qui utilise comme données des Expressed Sequence Tags (ESTs). La performance de TESTLoc est significativement supérieure à celle de tout autre outil de prédiction connu. En plus de fournir deux nouveaux outils de prédiction de la localisation subcellulaire utilisant différents types de données, nos travaux démontrent comment lassociation de la prédiction de la localisation subcellulaire à dautres méthodes danalyse in silico permet daméliorer la connaissance des protéines mitochondriales. De plus, ces travaux proposent des hypothèses claires et faciles à vérifier par des expériences, ce qui présente un grand potentiel pour faire progresser nos connaissances des métabolismes mitochondriaux. / The important role of mitochondria in the eukaryotic cell has long been appreciated, but their exact composition and the biological processes taking place in mitochondria are not yet fully understood. The two main factors that slow down the progress in this field are inefficient recognition and imprecise annotation of mitochondrial proteins. Therefore, we developed a new computational tool, YimLoc, which effectively predicts mitochondrial proteins from genomic sequences. This tool integrates the strengths of existing predictors and yields higher performance than any individual predictor. We applied YimLoc to ~60 fungal genomes in order to address the controversy about the localization of beta oxidation in these organisms. Our results show that in contrast to previous studies, most fungal groups do possess mitochondrial beta oxidation. This work also revealed the diversity of beta oxidation in fungi, which correlates with their utilization of fatty acids as energy and carbon sources. Further, we conducted an investigation of the key component of the mitochondrial beta oxidation pathway, the acyl-CoA dehydrogenase (ACAD). We combined subcellular localization prediction with subfamily classification and phylogenetic inference of ACAD enzymes from 250 species covering all three domains of life. Our study suggests that ACAD genes are an ancient family with innovative evolutionary strategies to generate a large enzyme toolset for utilizing most diverse fatty acids and amino acids. Finally, to enable the prediction of mitochondrial proteins from data beyond genome sequences, we designed the tool TESTLoc that uses expressed sequence tags (ESTs) as input. TESTLoc performs significantly better than known tools. In addition to providing two new tools for subcellular localization designed for different data, our studies demonstrate the power of combining subcellular localization prediction with other in silico analyses to gain insights into the function of mitochondrial proteins. Most importantly, this work proposes clear hypotheses that are easily testable, with great potential for advancing our knowledge of mitochondrial metabolism.
74

IDENTIFICATION OF TARGETS AND AUXILIARY PROTEINS OF PYR/PYL/RCAR ABA RECEPTORS: PROTEIN PHOSPHATASES TYPE 2C (PP2Cs) AND C2-DOMAIN ABA-RELATED PROTEINS (CARs)

Rodríguez Solovey, Leisa Natacha 16 December 2015 (has links)
[EN] ABSTRACT Abscisic acid (ABA) signaling plays a critical role in regulating root growth and root system architecture. ABA-mediated growth promotion and root tropic response under water stress are key responses for plant survival under limiting water conditions. In this work, we have explored the role of Arabidopsis (Arabidopsis thaliana) PYR/PYL/RCAR receptors (PYRABACTIN RESISTANCE1 (PYR1)/PYR1 LIKE (PYL)/REGULATORY COMPONENTS OF ABA RECEPTORS) for root ABA signaling. As a result, we discovered that PYL8 plays a nonredundant role for the regulation of root ABA sensitivity. Unexpectedly, given the multigenic nature and partial functional redundancy observed in the PYR/PYL family, the single pyl8 mutant showed reduced sensitivity to ABA-mediated root growth inhibition. This effect was due to the lack of PYL8-mediated inhibition of several clade A phosphatases type 2C (PP2Cs), since PYL8 interacted in vivo with at least five PP2Cs, namely HYPERSENSITIVE TO ABA1 (HAB1), HAB2, ABAINSENSITIVE1 (ABI1), ABI2, and PP2CA/ABA-HYPERSENSITIVE GERMINATION3 as revealed by tandem affinity purification and mass spectrometry proteomic approaches. Membrane-delimited abscisic acid (ABA) signal transduction plays a critical role in early ABA signaling, but the molecular mechanisms linking core signaling components to the plasma membrane are unclear. We show that transient calciumdependent interactions of PYR/PYL/RCAR ABA receptors with membranes are mediated through a 10-member family of C2-domain ABA-related (CAR) proteins in Arabidopsis thaliana. Specifically, we found that PYL4 interacted in an ABA-independent manner with CAR1 in both the plasma membrane and nucleus of plant cells. CAR1 belongs to a plant-specific gene family encoding CAR1 to CAR10 proteins, and bimolecular fluorescence complementation and coimmunoprecipitation assays showed that PYL4-CAR1 as well as other PYR/PYL-CAR pairs interacted in plant cells. The crystal structure of CAR4 was solved, which revealed that, in addition to a classical calcium-dependent lipid binding C2 domain, a specific CAR signature is likely responsible for the interaction with PYR/PYL/RCAR receptors and their recruitment to phospholipid vesicles. This interaction is relevant for PYR/PYL/RCAR function and ABA signaling, since different car triple mutants affected in CAR1, CAR4, CAR5, and CAR9 genes showed reduced sensitivity to ABA in seedling establishment and root growth assays. In summary, we identified PYR/PYL/RCAR-interacting partners that mediate a transient Ca2+-dependent interaction with phospholipid vesicles, which affects PYR/PYL/RCAR subcellular localization and positively regulates ABA signaling. / [ES] RESUMEN La señalización por la hormona vegetal ácido abscísico (ABA) desempeña un papel crítico en la regulación del crecimiento de la raíz y en la arquitectura del sistema radical. La promoción de crecimiento de la raíz en condiciones de estrés hídrico mediada por ABA es clave para la supervivencia de las plantas bajo condiciones limitantes de agua. En este trabajo, hemos explorado el papel de los receptores PYR/PYL/RCAR (PYRABACTIN RESISTANCE1 (PYR1)/PYR1 LIKE (PYL)/ REGULATORY COMPONENTS OF ABA RECEPTORS) de Arabidopsis (Arabidopsis thaliana) en la ruta de señalización de ABA en raíz. Así, hemos descubierto que el receptor de ABA PYL8 juega un papel no redundante en la regulación de la percepción de ABA en raíz. Inesperadamente, dada la naturaleza multigénica y la redundancia funcional parcial observada en la familia PYR/PYL/RCAR, el mutante pyl8 fue el único mutante sencillo de pérdida de función de los receptores PYR/PYL/RCAR que mostraba una sensibilidad reducida a la inhibición del crecimiento mediada por ABA en raíz. Este efecto se debe a la falta de inhibición mediada por PYL8 de varias fosfatasas del grupo A tipo 2C (PP2Cs), ya que PYL8 es capaz de interactuar in vivo con al menos cinco PP2Cs, denominadas HYPERSENSITIVE TO ABA1 (HAB1), HAB2, ABAINSENSITIVE1 (ABI1), ABI2, and PP2CA/ABA-HYPERSENSITIVE GERMINATION3 según lo han revelado la purificación por afinidad en tándem (TAP por sus siglas en inglés) y estudios proteómicos de espectrometría de masas. La transducción de la señal del ABA localizada en la membrana plasmática celular juega un papel crucial en los pasos iniciales de la señalización de la fitohormona, pero los mecanismos moleculares que unen los componentes básicos de la señalización y la membrana plasmática no están claros. Estudiando las interacciones de los receptores del ABA PYR/PYL/RCAR con la membrana plasmática hemos encontrado que éstos pueden interaccionar transitoriamente con ella de forma dependiente de calcio gracias a una familia de proteínas con dominios C2 relacionadas con la ruta de señalización de ABA (denominadas C2-domain ABA-related (CAR) proteins). Específicamente, se encontró que PYL4 interacciona de manera independiente de ABA con CAR1 tanto en la membrana plasmática como en el núcleo de las células vegetales. La proteína CAR1 pertenece a una familia multigénica constituida por 10 miembros en Arabidopsis thaliana, desde CAR1 hasta CAR10, y que solo se encuentra en plantas. Los ensayos de complementación bi-molecular de fluorescencia y de co-immunoprecipitación confirmaron la interacción en células vegetales tanto de PYL4-CAR1 como de otras parejas de PYR/PYL-CAR. La cristalización de la proteína CAR4 reveló que, además de un dominio C2 clásico de unión a lípidos dependiente de calcio, las proteínas de la familia CAR presentan un dominio específico que probablemente es responsable de la interacción con los receptores PYR/PYL/RCAR y de su posterior reclutamiento a las vesículas de fosfolípidos. Esta interacción es relevante para la función de los receptores PYR/PYL/RCAR en la señalización del ABA, ya que diferentes mutantes triples car de pérdida de función, que tienen afectados los genes CAR1, CAR4, CAR5, y CAR9, demostraron una reducción de la sensibilidad al ABA en ensayos de establecimiento de plántula y crecimiento de la raíz. En resumen, hemos identificado nueva familia de proteínas que son capaces mediar las interacciones transitorias dependientes de Ca2+ con vesículas de fosfolípidos, lo que a su vez afecta localización de PYR/PYL/RCAR y regula positivamente la señalización de ABA. / [CAT] RESUM La senyalització per l'hormona vegetal àcid abcíssic (ABA) exerceix un paper crític en la regulació del creixement de l'arrel i també en l'arquitectura del sistema radical. La promoció del creixement de l'arrel en condicions d'estrés hídric, regulada per ABA és clau per la supervivència de les plantes sota condicions limitants d'aigua. Amb aquest treball, hem investigat el paper dels receptors PYR/PYL/RCAR (PYRABACTIN RESISTANCE1 (PYR1)/PYR1 LIKE (PYL)/ REGULATORY COMPONENTS OF ABA RECEPTORS) d'Arabidopsis (Arabidopsis thaliana) en el camí de senyalització d'ABA en arrel. Així, hem descobert que el receptor d'ABA PYL8 exerceix un paper no redundant en la regulació de la percepció d'ABA en arrel. Inesperadament, donada la naturalesa multigènica i la redundància funcional parcial que s'observa en la família PYR/PYL/RCAR, el mutant pyl8 va ser l'únic mutant senzill de pèrdua de funció dels receptors PYR/PYL/RCAR que mostrava una sensibilitat reduïda a la inhibició del creixement mitjançada per l'ABA en l'arrel. Doncs aquest efecte es deu a la falta d'inhibició regulada per PYL8 de diverses fosfatases del grup A tipus 2C (PP2Cs), ja que PYL8 té la capacitat d'interactuar in vivo almenys amb cinc PP2Cs, anomenades HYPERSENSITIVE TO ABA1 (HAB1), HAB2, ABAINSENSITIVE1 (ABI1), ABI2, and PP2CA/ABAHYPERSENSITIVE GERMINATION3 segons ho han revelat per una banda la purificació per afinitat en tàndem (TAP són les seues sigles en anglés) i per altra banda, estudis proteòmics d'espectrometria de masses. Pel que fa a la transducció del senyal del l'ABA, la qual es localitza en la membrana plasmàtica cel¿lular, juga un paper molt important en els primers instants de la senyalització de la fitohormona, no obstant això els mecanismes moleculars que uneixen els components bàsics d'aquesta senyalització amb la membrana plasmàtica, no es troben del tot clars. Per tant, s'han estudiat les interaccions que tenen els receptors del ABA PYR/PYL/RCAR amb la membrana plasmàtica, i hem trobat que aquests tenen la capacitat d'interaccionar transitòriament amb la membrana de forma dependent al calci, gràcies a una família de proteïnes amb domini C2, les quals es troben relacionades amb la ruta de senyalització d'ABA(anomenades C2domain ABArelated (CAR) proteins).Específicament, es va trobar que PYL4 interacciona d'una manera independent al ABA amb CAR1, tant en la membrana plasmàtica, com en el nucli de les cèl¿lules vegetals. La proteïna CAR1 pertany a la família multigènica constituïda per 10 components en Arabidopsis thaliana, des de CAR1 fins CAR10, que tan sols es troba en plantes. Els assajos de complementació bimolecular de fluorescència i de co-immunoprecipitació, van confirmar la interacció en cèl¿lules vegetals, tant de PYL4CAR1 com d'altres parelles de PYR/PYL-CAR. La cristal¿lització de la proteïna CAR4 va revelar que, a més d'un domini C2 clàssic de unió a lípids dependent del calci, les proteïnes de la família CAR presenten un domini PYR/PYL/RCAR, i del seu posterior reclutament a les vesícules fosfolipídiques. Doncs, aquesta interacció és rellevant en la funció dels receptors PYR/PYL/RCAR, ja que participa en la senyalització del l'ABA. Aquesta interacció es clau per a la funció dels receptors, ja que diferents mutants triples car de pèrdua de funció, els quals posseïxen afectats els gens CAR1, CAR4, CAR5 i CAR9, van mostrar una reducció de la sensibilitat a l'ABA en assajos d'establiment de plàntula i creixement de l'arrel. En conclusió, hem identificat una nova família de proteïnes amb la capacitat d'organitzar les interaccions transitòries dependents del calci amb vesícules de fosfolípids, fet que al seu torn afecta la localització de PYR/PYL/RCAR i regula positivament la senyalització d'ABA. / Rodríguez Solovey, LN. (2015). IDENTIFICATION OF TARGETS AND AUXILIARY PROTEINS OF PYR/PYL/RCAR ABA RECEPTORS: PROTEIN PHOSPHATASES TYPE 2C (PP2Cs) AND C2-DOMAIN ABA-RELATED PROTEINS (CARs) [Tesis doctoral no publicada]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/58862 / TESIS

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