Ureide synthesis, accumulation and transport in Arabidopsis plants under salt and osmotic stress

The ureides allantoin and allantoic acid play a central role in nitrogen transport in nodulating tropical legumes. However, the complete enzyme set for ureide synthesis and a family of ureide permeases are widely distributed in the plant kingdom suggesting their participation in physiological processes not properly characterized yet. In Arabidopsis, microarrays studies showed an upregulation of ureides synthesis genes (xanthine dehydrogenase, uricase) during abiotic stresses. On the contrary, allantoinase gene expression is strongly reduced after stress suggesting that allantoin may accumulate in the cells. We observed accumulation of allantoin in Arabidopsis plants under osmotic and salt stresses. This effect was exacerbated in plants grown with ammonium as nitrogen source and suppressed in the presence of sucrose as carbon source. The analysis of two independent T-DNA insertion lines, causing knockout of allantoinase (alla-1 and alla-2) showed constitutively elevated concentrations of allantoin, but a noticeable morphological phenotype remained elusive. To determine the physiological relevance of allantoinase gene repression on resistance to stress, transgenic lines were generated on the genotype alla-1, in which the coding sequence of allantoinase was introduced under the control of the stress inducible promoter RD29A. As expected, alla-1 pRD29A::Aln plants were not able to accumulate allantoin under stress conditions. The phenotype of KO and RD29A:ALN plants was analysed under salt and osmotic stress conditions. In addition, the phenotype of a KO mutant of AtUPS5 (ups5) was analysed. AtUPS5 transports allantoin, is expressed in the root cortex and endodermis and its expression increases during salt and osmotic stress, suggesting a rol in long-distance transport of allantoin during stress.Fil: Lescano, Ignacio. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales; Argentina.Fil: Lescano, Ignacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto Multidisciplinario de Biología Vegetal; Argentina.Fil: Martini, Carolina. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales; Argentina.Fil: Martini, Carolina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto Multidisciplinario de Biología Vegetal; Argentina.Fil: Tessi, Tomás. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales; Argentina.Fil: Tessi, Tomás. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto Multidisciplinario de Biología Vegetal; Argentina.Fil: González, Claudio. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales; Argentina.Fil: González, Claudio. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto Multidisciplinario de Biología Vegetal; Argentina.Fil: Desimone, Marcelo. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales; Argentina.Fil: Desimone, Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto Multidisciplinario de Biología Vegetal; Argentina.Bioquímica y Biología Molecular (ídem 3.1.10

Allantoin accumulation mediated by allantoinase downregulation and transport by Ureide Permease 5 confers salt stress tolerance to Arabidopsis plants

Allantoin, a metabolite generated in the purine degradation pathway, was primarily considered an intermediate for recycling of the abundant nitrogen assimilated in plant purines. More specifically, tropical legumes utilize allantoin and allantoic acid as major nodule-to-shoot nitrogen transport compounds. In other species, an increase in allantoin content was observed under different stress conditions, but the underlying molecular mechanisms remain poorly understood. In this work, Arabidopsis thaliana was used as a model system to investigate the effects of salt stress on allantoin metabolism and to know whether its accumulation results in plant protection. Plant seedlings treated with NaCl at different concentrations showed higher allantoin and lower allantoic acid contents. Treatments with NaCl favored the expression of genes involved in allantoin synthesis, but strongly repressed the unique gene encoding allantoinase (AtALN). Due to the potential regulatory role of this gene for allantoin accumulation, AtALN promoter activity was studied using a reporter system. GUS mediated coloration was found in specific plant tissues and was diminished with increasing salt concentrations. Phenotypic analysis of knockout, knockdown and stress-inducible mutants for AtALN revealed that allantoin accumulation is essential for salt stress tolerance. In addition, the possible role of allantoin transport was investigated. The Ureide Permease 5 (UPS5) is expressed in the cortex and endodermis of roots and its transcription is enhanced by salt treatment. Ups5 knockout plants under salt stress presented a susceptible phenotype and altered allantoin root-to-shoot content ratios. Possible roles of allantoin as a protectant compound in oxidative events or signaling are discussed.Fil: Lescano López, Ignacio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: Martini, Carolina María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: González, Claudio Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: Desimone, Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; Argentin

Expression pattern of a nuclear encoded mitochondrial arginine-ornithine translocator gene from Arabidopsis

BACKGROUND: Arginine and citrulline serve as nitrogen storage forms, but are also involved in biosynthetic and catabolic pathways. Metabolism of arginine, citrulline and ornithine is distributed between mitochondria and cytosol. For the shuttle of intermediates between cytosol and mitochondria transporters present on the inner mitochondrial membrane are required. Yeast contains a mitochondrial translocator for ornithine and arginine, Ort1p/Arg11p. Ort1p/Arg11p is a member of the mitochondrial carrier family (MCF) essential for ornithine export from mitochondria. The yeast arg11 mutant, which is deficient in Ort1p/Arg11p grows poorly on media lacking arginine. RESULTS: High-level expression of a nuclear encoded Arabidopsis thaliana homolog (AtmBAC2) of Ort1p/Arg11p was able to suppress the growth deficiency of arg11. RT-PCR analysis demonstrated expression of AtmBAC2 in all tissues with highest levels in flowers. Promoter-GUS fusions showed preferential expression in flowers, i.e. pollen, in the vasculature of siliques and in aborted seeds. Variable expression was observed in leaf vasculature. Induction of the promoter was not observed during the first two weeks in seedlings grown on media containing NH(4)NO(3), arginine or ornithine as sole nitrogen sources. CONCLUSION: AtmBAC2 was isolated as a mitochondrial transporter for arginine in Arabidopsis. The absence of expression in developing seeds and in cotyledons of seedlings indicates that other transporters are responsible for storage and mobilization of arginine in seeds

Identification of an Arabidopsis mitochondrial succinate–fumarate translocator

AbstractComplementation of a yeast acr1 mutant carrying a deletion of the succinate/fumarate carrier gene enabled functional identification of a mitochondrial succinate translocator from Arabidopsis thaliana (AtmSFC1). Thus complementation of yeast mutants is applicable also for identification and characterization of organellar transporters. Reverse transcription polymerase chain reaction and promoter-GUS fusion showed expression of AtmSFC1 in 2 day old dark grown seedlings, which declined in cotyledons during further development, consistent with a role in export of fumarate for gluconeogenesis during lipid mobilization at early germination of Arabidopsis seeds. In mature plants, expression was found in developing and germinating pollen, suggesting a role in ethanolic fermentation

Caracterización morfo-fisiológica de una solanaceae halófila extrema nativa de los Andes

Lycium humile Phil. (Solanaceae) es un arbusto halófilo endémico de los Andes argentino-chilenos que crece en los salares puneños. De los aspectos morfo-fisiológicos estudiados hasta el momento, se destaca el reporte de glándulas de sal en hojas de una población chilena y su mesofilo suculento que podría estar relacionado con metabolismo CAM. El objetivo de este trabajo es corroborar la existencia de glándulas de sal y evaluar variación diaria de la acidez tisular, prolina y relaciones Na+/K+/Ca+2/Mg+2 utilizando hojas de plantas del Salar de Antofalla (Catamarca, Argentina). El análisis anatómico determinó la ausencia de glándulas de sal mientras que, tampoco hubo cambios diarios en la acidez tisular, por lo que se presume no presentan metabolismo fotosintético tipo CAM. Se detectaron niveles de prolina comparables a los reportados para otras especies halófilas, y una alta proporción de sodio respecto al resto de los iones. Dado que se ha demostrado que la acumulación de ureidos ocurre frente a situaciones de salinidad, se determinaron las concentraciones de alantoína y ácido alantoico, encontrándose bajos niveles de ambos compuestos. De los resultados obtenidos podemos inferir que la estrategia de L. humile para afrontar la salinidad extrema estaría relacionada con la acumulación de prolina e iones.Fil: Palchetti, Maria Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: Martini, Carolina María. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales; ArgentinaFil: Barboza, Gloria Estela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: Grunberg, Karina Alejandra. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Farmacia; Argentina. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigaciones Agropecuarias. Unidad de Estudios Agropecuarios - Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Unidad de Estudios Agropecuarios; ArgentinaFil: Desimone, Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: Cantero, Juan Jose. Instituto Nacional de Tecnología Agropecuaria; ArgentinaXXXII Reunión Argentina, XVI Congreso Latinoamericano de Fisiología VegetalCórdobaArgentinaSociedad Argentina de Fisiología Vegeta

PvUPS1, an allantoin transporter in nodulated roots of French bean

Nodulated legumes receive their nitrogen via nitrogen-fixing rhizobia, which exist in a symbiotic relationship with the root system. In tropical legumes like French bean (Phaseolus vulgaris) or soybean (Glycine max), most of the fixed nitrogen is used for synthesis of the ureides allantoin and allantoic acid, the major long-distance transport forms of organic nitrogen in these species. The purpose of this investigation was to identify a ureide transporter that would allow us to further characterize the mechanisms regulating ureide partitioning in legume roots. A putative allantoin transporter (PvUPS1) was isolated from nodulated roots of French bean and was functionally characterized in an allantoin transport-deficient yeast mutant showing that PvUPS1 transports allantoin but also binds its precursors xanthine and uric acid. In beans, PvUPS1 was expressed throughout the plant body, with strongest expression in nodulated roots, source leaves, pods, and seed coats. In roots, PvUPS1 expression was dependent on the status of nodulation, with highest expression in nodules and roots of nodulated plants compared with non-nodulated roots supplied with ammonium nitrate or allantoin. In situ RNA hybridization localized PvUPS1 to the nodule endodermis and the endodermis and phloem of the nodule vasculature. These results strengthen our prediction that in bean nodules, PvUPS1 is involved in delivery of allantoin to the vascular bundle and loading into the nodule phloem

Cytokinin transporter azg2 modulates cell wall remodeling during lateral root emergence in Arabidopsis thaliana

Tissue remodeling is essential for lateral root (LR) emergence. Many studies have focused on the role of auxin in this process but little is known about cytokinin (CK) relevance. AZG2 is an auxin regulated CK transporter, involved in negative regulation of LR development in Arabidopsis. AZG2 is expressed in a small group of cortical and epidermal cells (OLT) surrounding lateral root primordia (LRP), resembling some cell wall remodeling enzymes (CWREs) involved in lateral root emergence (LRE). Here, we study LRE and cell wall remodeling (CWR) in Arabidopsis roots of WT and azg2-1 genotypes through two independent experiments. LRP/LR index was calculated and values were 2,93 for azg2-1 and 3,48 for WT roots, confirming that AZG2 has a regulatory effect on LRE. Additionally, a lower propidium iodide fluorescence intensity was detected in the OLTs of azg2 seedlings compared to WT, suggesting an increased CWR activity in the mutant genotype. Taken together, these results indicate that CK transport mediated by AZG2 could modulate CWR during LRE. In order to get further insight, we are currently working on pCWRE:GUS:GFP/WT and pCWRE:GUS:GFP/azg2 reporter lines to study CK effect on the expression of two different CWREs.Fil: Pettinari, Georgina Lucía. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Departamento de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Tessi, Tomás María. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: González, Claudio Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaFil: Desimone, Marcelo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto Multidisciplinario de Biología Vegetal. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas Físicas y Naturales. Instituto Multidisciplinario de Biología Vegetal; ArgentinaXXXII Reunión argentina XVI Congreso Latinoamericano de Fisiología VegetalCórdobaArgentinaSociedad Argentina de Fisiología Vegeta

Transcription Analysis of Arabidopsis Membrane Transporters and Hormone Pathways during Developmental and Induced Leaf Senescence

A comparative transcriptome analysis for successive stages of Arabidopsis (Arabidopsis thaliana) developmental leaf senescence (NS), darkening-induced senescence of individual leaves attached to the plant (DIS), and senescence in dark-incubated detached leaves (DET) revealed many novel senescence-associated genes with distinct expression profiles. The three senescence processes share a high number of regulated genes, although the overall number of regulated genes during DIS and DET is about 2 times lower than during NS. Consequently, the number of NS-specific genes is much higher than the number of DIS- or DET-specific genes. The expression profiles of transporters (TPs), receptor-like kinases, autophagy genes, and hormone pathways were analyzed in detail. The Arabidopsis TPs and other integral membrane proteins were systematically reclassified based on the Transporter Classification system. Coordinate activation or inactivation of several genes is observed in some TP families in all three or only in individual senescence types, indicating differences in the genetic programs for remobilization of catabolites. Characteristic senescence type-specific differences were also apparent in the expression profiles of (putative) signaling kinases. For eight hormones, the expression of biosynthesis, metabolism, signaling, and (partially) response genes was investigated. In most pathways, novel senescence-associated genes were identified. The expression profiles of hormone homeostasis and signaling genes reveal additional players in the senescence regulatory network