Biosynthesis of Phenylalanine in Plants

El aminoácido fenilalanina (Phe) desempeña en las plantas terrestres una función esencial, al actuar como precursor tanto de las proteínas como de la síntesis de fenilpropanoides, una amplia familia de metabolitos secundarios que cumplen funciones muy diversas y cuya aparición y diversificación está interrelacionada con la propia evolución de las plantas terrestres. La importancia de la síntesis de Phe es tal que se estima que más del 30% del CO2 fijado por las plantas en la fotosíntesis es finalmente derivado hacia la síntesis de este aminoácido, y de ahí hacia la biosíntesis de fenilpropanoides (Boerjan et ál., 2003), más particularmente ligninas, uno de los componentes fundamentales de las paredes celulares secundarias de las plantas. El objetivo general de esta tesis es obtener un mejor conocimiento la biosíntesis de fenilalanina en las plantas y su regulación en relación a la biosíntesis de metabolitos secundarios

The arogenate dehydratase ADT2 is essential for seed development in Arabidopsis

Phenylalanine (Phe) biosynthesis in plants is a key process, as Phe serves as precursor of proteins and phenylpropanoids. The prephenate pathway connects chorismate, final product of the shikimate pathway, with the biosynthesis of Phe and Tyr. Two alternative routes of Phe biosynthesis have been reported: one depending of arogenate, and the other of phenylpyruvate. Whereas the arogenate pathway is considered the main route, the role of the phenylpyruvate pathway remains unclear. Here, we report that the deficiency in ADT2, a bifunctional arogenate dehydratase (ADT)/ prephenate dehydratase (PDT) enzyme, causes embryo arrest and seed abortion. This result makes a clear distinction between the essential role of ADT2 and the five remaining ADTs from Arabidopsis, which display mostly overlapping functions. We have found that PHA2, a monofunctional PDT from yeast, restores the adt2 phenotype when is targeted within the plastids, but not when is expressed in the cytosol. Similar results can be obtained by expressing ADT3, a monofunctional ADT. These results suggest that Phe can be synthesized from phenylpyruvate or arogenate when the bifunctional ADT2 is replaced by other ADT or PDT enzymes during seed formation, highlighting the importance of Phe for embryo development, and providing further insights into the plasticity of Phe biosynthesis.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Multidisciplinary teaching of Biotechnology and Omics sciences

In the last years, there was a great boom in the Omics fields that have developed as multidisciplinary sciences. They use laboratory techniques related to Biology and Chemistry but also Bioinformatics tools. However, the developmental progress of these disciplines has led that much of undergraduate studies related to Biology have curricula that become outdated. From this point of view, it is necessary to focus the students to the fundamentals and techniques of complementary disciplines that will be essentials for the understanding of the Omics sciences. In the present work, we have developed a new teaching approach for Biochemistry, Biology and Bioinformatics students. They formed interdisciplinary working groups. These groups have prepared and presented communications about different techniques or methods in Molecular Biology, Omics or Bioinformatics participating in a technical meeting. This learning strategy “I do and I learn” has enabled to the students a first contact with the scientific communication including the approach to the scientific literature to acquire technical knowledge. The cooperation between students from different disciplines has enriched their point of view and even has been used in some practical master’s works.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Maritime pine PpMYB8 directly co-regulates secondary cell wall architecture and the associated Phe-biosynthesis pathway

Plants rely on the biosynthesis of L-Phenylalanine as building block for the synthesis of proteins but also as precursor for a tremendous range of plant-derived compounds essential for its grown, development and defense. Polymerization of secondary cell wall in trees involves the massive biosynthesis, among others, of the Phe-derived compound lignin. Thus, these plants require an accurate metabolic coordination between Phe and lignin biosynthesis to ensure its normal development. We have here identified that the pine arogenate dehydratase, whose enzyme activity limits the biosynthesis of Phe in plants, is transcriptionally regulated through direct interaction with PpMyb8. We have also shown that this transcription factor is directly involve in secondary cell wall biogenesis and cell death processes. Together these results indicate that a single transcription factor coordinates lignin accumulation and the proper biosynthesis of its essential precursor L-Phe.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Metabolic channeling of phe for lignin biosynthesis in maritime pine

Phenylalanine (Phe) is the main precursor of phenylpropanoids biosynthesis in plants. This vast family of Phe-derived compounds can represent more than 30% of captured photosynthetic carbon, playing essential roles in plants such as cell wall components, defense molecules, pigments and flavors. In addition to its physiological importance, phenylpropanoids and particularly lignin, a component of wood, are targets in plant biotechnology. The arogenate pathway has been proposed as the main pathway for Phe biosynthesis in plants (Maeda et al., 2010). The final step in Phe biosynthesis, catalyzed by the enzyme arogenate dehydratase (ADT), has been considered as a key regulatory point in Phe biosynthesis, due to its key branch position in the pathway, the multiple isoenzymes identified in plants and the existence of a feedback inhibition mechanism by Phe. So far, the regulatory mechanisms underlying ADT genes expression have been poorly characterized, although a strong regulation of the Phe metabolic flux should be expected depending on its alternative use for protein biosynthesis versus phenylpropanoid biosynthesis. This second fate involves a massive carbon flux compared to the first one. Here we report our current research activities in the transcriptional regulation of ADT genes by MYB transcription factors in Pinus pinaster. The conifers channels massive amounts of photosynthetic carbon for phenylpropanoid biosynthesis during wood formation. We have identified the complete ADT gene family in maritime pine (El-Azaz et al., 2016) and a set of ADT isoforms specifically related with the lignification process. The potential control of transcription factors previously reported as key regulators in pine wood formation (Craven-Bartle et al., 2013) will be presented.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Estudio comparativo de las enzimas implicadas en la biosíntesis de arginina a partir de ornitina en plantas

La arginina es un aminoácido presente en todos los organismos cuya función principal es formar parte de la estructura primaria de las proteínas, pero también tiene otros destinos, como la síntesis de poliaminas y óxido nítrico (ON), participa en el metabolismo de la prolina. En vegetales, además, debido a su alto contenido en nitrógeno (4 átomos de nitrógeno frente a 6 de carbono) sirve como almacén transitorio y a largo plazo de nitrógeno, en forma de proteínas de reserva. Dada la limitación que supone la falta de nitrógeno para el desarrollo de las plantas, muchas han desarrollado estrategias para la rápida captación y acumulación de cualquier especie de nitrógeno que haya disponible. Una de estas estrategias es la acumulación transitoria de arginina, en su forma libre o como parte de las proteínas de reserva, que, acumuladas en la semilla, también proveen del nitrógeno necesario para el desarrollo del embrión durante la germinación. Todas las plantas pueden sintetizar poliaminas a partir de arginina, a través de la ruta de la enzima arginina descarboxilasa (ADC) localizada en el cloroplasto. Sin embargo, existe otra ruta de síntesis de poliaminas, a partir de ornitina, mediante la enzima ornitina descarboxilasa (ODC). En las plantas que no poseen esta ruta alternativa, todo el flujo de nitrógeno para la síntesis de poliaminas, arginina y ON debe pasar por la síntesis de arginina, por lo tanto, la ausencia de ODC supone una mayor eficiencia de la ruta para producir arginina. Se comparan desde el punto de vista cinético las enzimas ornitina transcarbamoilasa (OTC), argininosuccinato sintetasa (ASSY) y argininosuccinato liasa (ASL) de cuatro especies vegetales usadas como modelo de investigación, como son Pinus pinaster, Arabidopsis thaliana, Marchantia polymorpha y Physcomitrella patens, que además cuentan con dos particularidades: sólo P. pinaster y A. thaliana poseen una semilla con reservas nutricionales y tanto A. thaliana como P. patens carecen de ODC.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech