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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.
11

Molecular Mechanisms of p63-Derived Ectodermal Dysplasia

Lustig, Daniel 20 March 2012 (has links)
Molecular defects in the p63 gene give rise to severe physiological abnormalities in patients with ectodermal dysplasia, however the mechanisms by which p63 mutations disrupt p63 function are unknown. In this study we examined four ΔNp63α mutants; Ectrodactyly-Ectodermal Dysplasia with Clefting (EEC) R204W, R304W and Ankyloblepharon-Ectodermal Dysplasia with Clefting (AEC) mutants, L514F and G530V, and characterized DNA binding, transcription factor activity, oligomerization with wild-type p63 and changes in protein stability/nuclear localization. We also investigated the putative OD-SAM interaction in p63 and p73. We demonstrated that both the EEC and AEC mutants cannot transcriptionally activate the PERP promoter and can hetero-oligomerize forming dominant negative complexes with wild-type p63. We show that both EEC mutants and AEC L514F mutants are more stable which is not due to aberrant degradation by the E3 ligase Itch. Finally, we discovered that a novel interaction between the p73 OD and SAM domain is absent in p63.
12

Molecular Mechanisms of p63-Derived Ectodermal Dysplasia

Lustig, Daniel 20 March 2012 (has links)
Molecular defects in the p63 gene give rise to severe physiological abnormalities in patients with ectodermal dysplasia, however the mechanisms by which p63 mutations disrupt p63 function are unknown. In this study we examined four ΔNp63α mutants; Ectrodactyly-Ectodermal Dysplasia with Clefting (EEC) R204W, R304W and Ankyloblepharon-Ectodermal Dysplasia with Clefting (AEC) mutants, L514F and G530V, and characterized DNA binding, transcription factor activity, oligomerization with wild-type p63 and changes in protein stability/nuclear localization. We also investigated the putative OD-SAM interaction in p63 and p73. We demonstrated that both the EEC and AEC mutants cannot transcriptionally activate the PERP promoter and can hetero-oligomerize forming dominant negative complexes with wild-type p63. We show that both EEC mutants and AEC L514F mutants are more stable which is not due to aberrant degradation by the E3 ligase Itch. Finally, we discovered that a novel interaction between the p73 OD and SAM domain is absent in p63.
13

Modulation of allergic airway inflammation by glucocorticoids

Karabinskaya, Anna 19 September 2013 (has links)
No description available.
14

La 7β-hydroxy-épiandrostérone dans des modèles in vitro de cancer du sein : effets anti-estrogéniques et rôle des récepteurs des estrogènes

Niro, Sandra 29 May 2012 (has links) (PDF)
La 7β-hydroxy-épiandrostérone, stéroïde endogène dérivant de la DHEA, présente des propriétés anti-inflammatoires. En effet, elle module la voie des prostaglandines (PGs) en inhibant la production de la PGE2 pro-inflammatoires et en augmentant la production de la 15-Deoxy-∆12,14-PGJ2 cyto-protectrice in vivo et in vitro. Les faibles doses de 7β-hydroxy-épiandrostérone (1nM, 10nM, 100nM) pour lesquelles ces effets sont observés, suggèrent une liaison à un récepteur spécifique. L'inflammation et la production des PGs jouent un rôle important dans le développement et la prolifération des tumeurs mammaires estrogéno-dépendantes. Le 17β-estradiol (E2), en se fixant sur les récepteurs des estrogènes (REs), induit la production de PGE2 et la prolifération cellulaire dans ces cellules tumorales. De ce fait, notre objectif était de tester les effets de la 7β-hydroxy-épiandrostérone sur la prolifération (comptage avec exclusion au bleu trypan), le cycle cellulaire et l'apoptose (cytométrie de flux) dans les lignées cellulaires de cancer du sein MCF-7 (REα+, REβ+, GPR30+) et MDA-MB-231 (REα-, REβ+, GPR30+) et d'identifier une(des) cible(s) potentielle(s) dans ces cellules (transactivation) et dans des cellules négatives pour les REs nucléaires SKBr3 (GPR30+) (études de prolifération). Cette étude a montré que la 7β-hydroxy-épiandrostérone exerce des effets anti-estrogéniques dans les cellules MCF-7 et MDA-MB-231 associés à une inhibition de la prolifération et un arrêt du cycle cellulaire. Les études de transactivation et de prolifération avec les agonistes spécifiques des REs ont montré une interaction avec le REβ. De plus, les résultats des études de proliférations sur les trois lignées cellulaires suggèrent que la 7β-hydroxy-épiandrostérone pourrait également interagir avec le GPR30. Ces résultats indiquent que ce stéroïde androgène agit comme un anti-estrogène. De plus, c'est la première fois qu'un stéroïde androgène à faible dose montre une action anti-proliférative dans des lignées de cancers du sein. Des études ultérieures restent à réaliser afin de mieux comprendre ces effets observés.
15

Peptides vasoactifs endogènes dans la prolifération accrue des cellules musculaires lisses vasculaires de rats spontanément hypertendus: rôle des facteurs de croissance.

Lévesque, Louis-Olivier 06 1900 (has links)
Contribuant à la pathophysiologie des maladies vasculaires comme dans le cas de l’hypertension, le remodelage vasculaire est associé à une altération de la croissance des cellules musculaires lisses vasculaires (CMLV) (prolifération, taille, etc.). Or la prolifération des CMLV est augmentée par les peptides vasoactifs tels que l’angiotensine II (AngII) et l’endothéline-1 (ET-1). Ces peptides étant surexprimés lors de l’hypertension, cette étude fut entreprise pour déterminer leur contribution endogène ainsi que celles du facteur de croissance épidermique (EGF), du facteur de croissance insulinique (IGF-1) et du facteur de croissance dérivé des plaquettes (PDGF) à la prolifération accrue des CMLV et aux mécanismes sous-jacents. Des CMLV A-10 et des CMLV de rats WKY et SHR âgés de 12 semaines ont été utilisées pour cette étude. La prolifération cellulaire fut déterminée par incorporation de [3H]thymidine. La phosphorylation de ERK 1/2 et du récepteur de EGF fut déterminée par immunobuvardage. Les CMLV de SHR, comparées à celles de WKY, ont montré une prolifération accrue qui fut atténuée par le losartan, un antagoniste du récepteur AT1 de l’AngII et par le BQ-123 et le BQ-788, antagonistes des récepteurs ETA et ETB de l’ET-1. La prolifération accrue des CMLV de SHR fut ramenée à celle des WKY par les inhibiteurs des récepteurs au PDGF (AG-1295), au IGF-1 (AG-1024) et au EGF (AG-1478). La phosphorylation du récepteur au EGF, accrue dans les CMLV de rats SHR comparée à celle des WKY, fut atténuée par le losartan, le BQ-123, le BQ-788 et l’AG-1478, mais ne fut pas atténuée par l’AG-1295 et l’AG-1024. De plus, la phosphorylation accrue de ERK 1/2 dans les CMLV de rats SHR fut atténuée par le losartan, le BQ-123, le BQ-788 et les inhibiteurs des récepteurs aux facteurs de croissance. Parallèlement, le rôle de la transactivation de EGF-R dans la prolifération accrue induite par AngII et ET-1 fut aussi examiné dans les CMLV A-10. L’augmentation, induite par AngII et ET-1, de la prolifération et de la phosphorylation de ERK 1/2 dans les CMLV A-10 fut ramenée au niveau contrôle par AG-1478. Ces données suggèrent que les peptides vasoactifs endogènes induisent la prolifération accrue des CMLV par la signalisation des MAP kinases résultant de la transactivation de EGF-R. / Vascular remodelling that contributes to the pathophysiology of vascular diseases, including hypertension, is associated with alteration in vascular smooth muscle cell (VSMC) growth, hypertrophy, etc. We have recently shown that vasoactive peptides such as angiotensin II (AngII) and endothelin-1 (ET-1) increased the proliferation of VSMC. Since the levels of AngII, ET-1 and growth factors are increased in hypertension, the present studies were undertaken to examine if these endogenous vasoactive peptides and growth factors contribute to the enhanced proliferation of VSMC in spontaneously hypertensive rats (SHR) and to further investigate the underlying mechanisms responsible for enhanced proliferation. A10 VSMC and aortic VSMC from 12 week old SHR and age-matched WKY rats were used for these studies. Cell proliferation was determined by [3H]thymidine incorporation and ERK ½ and growth factor receptor phosphorylation was determined by Western blotting. VSMC from SHR exhibited enhanced cell proliferation as compared to WKY as determined by enhanced [3H]thymidine incorporation which was attenuated by AngII AT1 receptor antagonist losartan, as well as by endothelin receptor ETA and ETB antagonists BQ-123 and BQ-788, respectively. The inhibitors of platelet derived growth factor receptor (PDGF-R); AG-1295, epidermal growth factor receptor (EGF-R); AG-1478, and insulin-like growth factor receptor (IGF-R); AG-1024 also attenuated the enhanced proliferation of VSMC from SHR to WKY control levels. In addition, VSMC from SHR exhibited enhanced phosphorylation of EGF-R as compared to WKY, which was attenuated by losartan, BQ-123, BQ-788 and AG-1478, and not by AG-1295 and AG-1024. Furthermore, the enhanced phosphorylation of ERK ½ in VSMC from SHR was also attenuated by losartan, BQ-123 and BQ-788 as well as by growth factor receptor inhibitors, AG-1478, AG-1024 and AG-1295. The implication of growth factor receptor transactivation in AngII and ET-1 induced enhanced cell proliferation was also examined in A10 VSMC. Ang II or ET-1 induced enhanced proliferation of A-10 VSMC and enhanced ERK ½ phosphorylation was also restored to control levels by EGF-R inhibitor. These data suggest that vasoactive peptide-induced growth factor receptor transactivation through MAP kinase signaling may contribute to the enhanced proliferation of VSMC from SHR.
16

Transactivation of platelet-derived growth factor receptor type ??: Mechanisms and potential relevance in neurobiology

Kruk, Jeffrey Stephen January 2013 (has links)
In the absence of ligand, certain growth factor receptors can be activated via G protein-coupled receptor (GPCR) activation in a process termed transactivation. Serotonin (5-HT) receptors can transactivate the receptor tyrosine kinase (RTK) platelet-derived growth factor (PDGF) ?? receptors in smooth muscle cells, but it is not known if similar pathways occur in neuronal cells. Here, it is shown that 5-HT can transiently increase the phosphorylation of PDGF?? receptors in a time- and concentration-dependent manner in SH-SY5Y neuroblastoma cells. This transactivation pathway was pertussis-toxin sensitive, and was dependent on phospholipase C activity, intracellular calcium signaling and subsequent protein kinase C activation. Exogenous application of non-lethal concentrations of H2O2 induced the phosphorylation of PDGF?? receptors in a concentration-dependent fashion, similar to that observed with 5-HT. Further investigation revealed reactive oxygen species (ROS) production as a necessary component in the transactivation pathway, as scavenging ROS eliminated PDGF?? receptor phosphorylation. NADPH oxidase was determined to be the likely source of ROS given that the NADPH oxidase inhibitors diphenyleneiodonium chloride and apocynin abrogated PDGF?? receptor transactivation. The role of Src tyrosine kinase was also investigated, and its location in this signaling cascade was determined to be downstream of calcium signaling, but upstream of NADPH oxidase activity. In addition, the activation of ERK1/2 in this system was elucidated to be independent of PDGF?? receptor transactivation. Interestingly, 5-HT also transactivated TrkB receptors, another RTK whose function is implicated in clinical depression. Expectedly, the enzymes in this mechanism were consistent with those revealed in 5-HT-to-PDGF?? receptor signaling. This cross-talk between 5-HT and RTKs such as TrkB and PDGF?? receptors identifies a potentially important signaling link between the serotonergic system and neurotrophic factor signaling in neurons that could have implications in mental health disorders including depression. Furthermore, although transactivation pathways are commonly initiated by a GPCR, recent reports have demonstrated that selective serotonin reuptake inhibitors (SSRIs) were able to block 5-HT-induced transactivation of PDGF?? receptors, suggesting that in addition to GPCRs, monoamine transporters may also be involved in RTK transactivation. SH-SY5Y cells pretreated with the SSRI fluoxetine blocked 5-HT-induced transactivation of the PDGF?? receptors, but not PDGF-induced PDGF?? receptor activation. Upon further examination, it was discovered that during the pretreatment period, fluoxetine itself was transiently transactivating the PDGF?? receptor via 5-HT2 receptors. By the end of the pretreatment period, the effects of fluoxetine on PDGF?? receptor phosphorylation had returned to baseline, and a subsequent transactivating stimulus (5-HT) failed to ???re-transactivate??? the PDGF?? receptor. Additional investigations demonstrated that 5-HT pretreatment can block dopamine-induced PDGF?? receptor transactivation, but not PDGF-induced PDGF?? receptor activation. This is the first demonstration of the heterologous desensitization of an RTK via a transactivation pathway, and this phenomenon is specific for transactivation pathways because in all cases the PDGF?? receptor ligand PDGF-BB was able to directly stimulate receptor activity in spite of GPCR agonist pretreatment. Heterologous desensitization in transactivation signaling reveals a previously unknown short-term ???blackout??? period wherein no further transactivation signaling can occur to potentially exploit the mitogenic effects of RTK activation.
17

Peptides vasoactifs endogènes dans la prolifération accrue des cellules musculaires lisses vasculaires de rats spontanément hypertendus: rôle des facteurs de croissance

Lévesque, Louis-Olivier 06 1900 (has links)
No description available.
18

Identificação de fatores reguladores da expressão de genes ASR(ABA, Stress and Ripening) de arroz (Oryza sativa)

Schünemann, Mariana January 2015 (has links)
Os estresses abióticos impostos à planta no campo, tais como o estresse salino, toxidez por alumínio, frio, seca, entre outros, afetam seu crescimento, desenvolvimento e produtividade. Dentre as gramíneas cultivadas, o arroz (Oryza sativa) é uma das culturas de maior importância no Brasil, cuja oscilação na produção acarreta prejuízos consideráveis à economia brasileira. Dessa forma, o estudo das interações entre os estresses abióticos e as respostas dos vegetais frente a esses estímulos ambientais é fundamental para um conhecimento detalhado desses mecanismos. O arroz é um dos cereais mais tolerantes ao alumínio (Al), sendo um ótimo modelo para o estudo de mecanismos de tolerância. Neste trabalho, foi estudado o papel das proteínas ASR na tolerância ao Al. A expressão dos genes ASR (ABA, Stress and Ripening) é induzida por ABA, estresses e amadurecimento do fruto. Essas proteínas foram caracterizadas como chaperonas e fatores de transcrição. Os genes da família OsASR também têm o nível de transcritos aumentado em resposta ao Al, sendo OsASR1 e OsASR5 os genes com expressão mais abundante em arroz. No entanto, as regiões promotoras responsivas ao Al e os fatores de transcrição reguladores da expressão desses genes ainda não foram descritos. Assim, este trabalho tem como objetivo geral a identificação dos fatores que regulam a transcrição dos genes OsASR1 e OsASR5. Para tanto, construções contendo fragmentos dos promotores desses genes dirigindo a expressão do gene repórter GUS foram obtidas, visando ensaio de expressão transiente em protoplastos de arroz submetidos ao tratamento com Al. Os fragmentos das regiões promotoras de ambos os genes OsASR1 e OsASR5 foram todos responsivos ao Al. Também foi realizado ensaio de transativação em protoplastos de Arabidopsis thaliana para verificar a existência de auto-regulação nesses genes. Tanto ASR1 quanto ASR5 foram capazes de transativar seus próprios promotores. Além disso, foi realizada uma triagem de biblioteca de cDNAs por mono-híbrido em levedura com fragmento da região promotora de OsASR5. Com essa abordagem, foram identificados sete genes candidatos a codificadores de fatores capazes de interação DNA-proteína. / The abiotic stress that plants in the field are subjected to, such as salt, aluminum, cold, drought, among others, affect their growth, development and productivity. Among cultivated grasses, rice (Oryza sativa) is one of the most important crops in Brazil, whose oscillation in production causes considerable costs to the Brazilian economy. Thus, the study of interactions between abiotic stresses and plant responses to these environmental stimuli is essential to a detailed knowledge of these mechanisms. Rice is one of the most Al-tolerant crops, being a great model for studying Al-tolerance mechanisms. In this work, the role of ASR proteins in Al tolerance was studied. The expression of ASR (ABA, Stress and Ripening) genes is induced by ABA, stresses and fruit ripening. These proteins were characterized as chaperones and transcription factors. The OsASR genes also have increased transcript accumulation in response to Al, and OsASR1 and OsASR5 have the most abundant expression in rice. However, the Al-responsive promoter regions and the transcription factors that regulate these genes have not yet been described. Therefore, the goal of this work is to identify regulating factors of OsASR1 and OsASR5 gene transcription. For this, vectors containing promoter fragments of these genes driving the expression of the GUS gene were constructed and transient expression assays were performed in rice protoplasts subjected to Al treatment. All of the promoter fragments were Al-responsive for both OsASR1 and OsASR5 genes. Transactivation assays in Arabidopsis thaliana protoplasts were also conducted in order to verify the existence of auto-regulation in these genes. Both ASR1 and ASR5 were able to transactivate its own promoters. Furthermore, a library screening was performed by yeast one-hybrid using a promoter fragment from OsASR5. With this approach, seven candidate genes encoding transcription factors capable of DNA-protein interactions were identified.
19

Identificação de fatores reguladores da expressão de genes ASR(ABA, Stress and Ripening) de arroz (Oryza sativa)

Schünemann, Mariana January 2015 (has links)
Os estresses abióticos impostos à planta no campo, tais como o estresse salino, toxidez por alumínio, frio, seca, entre outros, afetam seu crescimento, desenvolvimento e produtividade. Dentre as gramíneas cultivadas, o arroz (Oryza sativa) é uma das culturas de maior importância no Brasil, cuja oscilação na produção acarreta prejuízos consideráveis à economia brasileira. Dessa forma, o estudo das interações entre os estresses abióticos e as respostas dos vegetais frente a esses estímulos ambientais é fundamental para um conhecimento detalhado desses mecanismos. O arroz é um dos cereais mais tolerantes ao alumínio (Al), sendo um ótimo modelo para o estudo de mecanismos de tolerância. Neste trabalho, foi estudado o papel das proteínas ASR na tolerância ao Al. A expressão dos genes ASR (ABA, Stress and Ripening) é induzida por ABA, estresses e amadurecimento do fruto. Essas proteínas foram caracterizadas como chaperonas e fatores de transcrição. Os genes da família OsASR também têm o nível de transcritos aumentado em resposta ao Al, sendo OsASR1 e OsASR5 os genes com expressão mais abundante em arroz. No entanto, as regiões promotoras responsivas ao Al e os fatores de transcrição reguladores da expressão desses genes ainda não foram descritos. Assim, este trabalho tem como objetivo geral a identificação dos fatores que regulam a transcrição dos genes OsASR1 e OsASR5. Para tanto, construções contendo fragmentos dos promotores desses genes dirigindo a expressão do gene repórter GUS foram obtidas, visando ensaio de expressão transiente em protoplastos de arroz submetidos ao tratamento com Al. Os fragmentos das regiões promotoras de ambos os genes OsASR1 e OsASR5 foram todos responsivos ao Al. Também foi realizado ensaio de transativação em protoplastos de Arabidopsis thaliana para verificar a existência de auto-regulação nesses genes. Tanto ASR1 quanto ASR5 foram capazes de transativar seus próprios promotores. Além disso, foi realizada uma triagem de biblioteca de cDNAs por mono-híbrido em levedura com fragmento da região promotora de OsASR5. Com essa abordagem, foram identificados sete genes candidatos a codificadores de fatores capazes de interação DNA-proteína. / The abiotic stress that plants in the field are subjected to, such as salt, aluminum, cold, drought, among others, affect their growth, development and productivity. Among cultivated grasses, rice (Oryza sativa) is one of the most important crops in Brazil, whose oscillation in production causes considerable costs to the Brazilian economy. Thus, the study of interactions between abiotic stresses and plant responses to these environmental stimuli is essential to a detailed knowledge of these mechanisms. Rice is one of the most Al-tolerant crops, being a great model for studying Al-tolerance mechanisms. In this work, the role of ASR proteins in Al tolerance was studied. The expression of ASR (ABA, Stress and Ripening) genes is induced by ABA, stresses and fruit ripening. These proteins were characterized as chaperones and transcription factors. The OsASR genes also have increased transcript accumulation in response to Al, and OsASR1 and OsASR5 have the most abundant expression in rice. However, the Al-responsive promoter regions and the transcription factors that regulate these genes have not yet been described. Therefore, the goal of this work is to identify regulating factors of OsASR1 and OsASR5 gene transcription. For this, vectors containing promoter fragments of these genes driving the expression of the GUS gene were constructed and transient expression assays were performed in rice protoplasts subjected to Al treatment. All of the promoter fragments were Al-responsive for both OsASR1 and OsASR5 genes. Transactivation assays in Arabidopsis thaliana protoplasts were also conducted in order to verify the existence of auto-regulation in these genes. Both ASR1 and ASR5 were able to transactivate its own promoters. Furthermore, a library screening was performed by yeast one-hybrid using a promoter fragment from OsASR5. With this approach, seven candidate genes encoding transcription factors capable of DNA-protein interactions were identified.
20

Identificação de fatores reguladores da expressão de genes ASR(ABA, Stress and Ripening) de arroz (Oryza sativa)

Schünemann, Mariana January 2015 (has links)
Os estresses abióticos impostos à planta no campo, tais como o estresse salino, toxidez por alumínio, frio, seca, entre outros, afetam seu crescimento, desenvolvimento e produtividade. Dentre as gramíneas cultivadas, o arroz (Oryza sativa) é uma das culturas de maior importância no Brasil, cuja oscilação na produção acarreta prejuízos consideráveis à economia brasileira. Dessa forma, o estudo das interações entre os estresses abióticos e as respostas dos vegetais frente a esses estímulos ambientais é fundamental para um conhecimento detalhado desses mecanismos. O arroz é um dos cereais mais tolerantes ao alumínio (Al), sendo um ótimo modelo para o estudo de mecanismos de tolerância. Neste trabalho, foi estudado o papel das proteínas ASR na tolerância ao Al. A expressão dos genes ASR (ABA, Stress and Ripening) é induzida por ABA, estresses e amadurecimento do fruto. Essas proteínas foram caracterizadas como chaperonas e fatores de transcrição. Os genes da família OsASR também têm o nível de transcritos aumentado em resposta ao Al, sendo OsASR1 e OsASR5 os genes com expressão mais abundante em arroz. No entanto, as regiões promotoras responsivas ao Al e os fatores de transcrição reguladores da expressão desses genes ainda não foram descritos. Assim, este trabalho tem como objetivo geral a identificação dos fatores que regulam a transcrição dos genes OsASR1 e OsASR5. Para tanto, construções contendo fragmentos dos promotores desses genes dirigindo a expressão do gene repórter GUS foram obtidas, visando ensaio de expressão transiente em protoplastos de arroz submetidos ao tratamento com Al. Os fragmentos das regiões promotoras de ambos os genes OsASR1 e OsASR5 foram todos responsivos ao Al. Também foi realizado ensaio de transativação em protoplastos de Arabidopsis thaliana para verificar a existência de auto-regulação nesses genes. Tanto ASR1 quanto ASR5 foram capazes de transativar seus próprios promotores. Além disso, foi realizada uma triagem de biblioteca de cDNAs por mono-híbrido em levedura com fragmento da região promotora de OsASR5. Com essa abordagem, foram identificados sete genes candidatos a codificadores de fatores capazes de interação DNA-proteína. / The abiotic stress that plants in the field are subjected to, such as salt, aluminum, cold, drought, among others, affect their growth, development and productivity. Among cultivated grasses, rice (Oryza sativa) is one of the most important crops in Brazil, whose oscillation in production causes considerable costs to the Brazilian economy. Thus, the study of interactions between abiotic stresses and plant responses to these environmental stimuli is essential to a detailed knowledge of these mechanisms. Rice is one of the most Al-tolerant crops, being a great model for studying Al-tolerance mechanisms. In this work, the role of ASR proteins in Al tolerance was studied. The expression of ASR (ABA, Stress and Ripening) genes is induced by ABA, stresses and fruit ripening. These proteins were characterized as chaperones and transcription factors. The OsASR genes also have increased transcript accumulation in response to Al, and OsASR1 and OsASR5 have the most abundant expression in rice. However, the Al-responsive promoter regions and the transcription factors that regulate these genes have not yet been described. Therefore, the goal of this work is to identify regulating factors of OsASR1 and OsASR5 gene transcription. For this, vectors containing promoter fragments of these genes driving the expression of the GUS gene were constructed and transient expression assays were performed in rice protoplasts subjected to Al treatment. All of the promoter fragments were Al-responsive for both OsASR1 and OsASR5 genes. Transactivation assays in Arabidopsis thaliana protoplasts were also conducted in order to verify the existence of auto-regulation in these genes. Both ASR1 and ASR5 were able to transactivate its own promoters. Furthermore, a library screening was performed by yeast one-hybrid using a promoter fragment from OsASR5. With this approach, seven candidate genes encoding transcription factors capable of DNA-protein interactions were identified.

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