Identificación y caracterización de mutantes de tomate con alteraciones en el desarrollo vegetativo

[ES] Estudiar la función de los genes que participan en procesos clave del desarrollo vegetal y su interacción con el medio aporta información muy importante, tanto básica como aplicada. Una aproximación para ello es la generación de mutantes para estos caracteres y su posterior caracterización. Si para ello se utiliza T-DNA como agente mutágeno, además de producir un mutante, el gen afectado queda etiquetado por el inserto de secuencia conocida, por lo que su clonación es más sencilla y rápida. En este trabajo se han analizado un total de 40 líneas T-DNA de Solanum lycopersicum variedad Money Maker, transformadas con una trampa de intensificadores. Para ello se han empleado tanto técnicas de cultivo in vitro como técnicas de cultivo convencional en el invernadero. El objetivo final era detectar nuevos mutantes con alteraciones en el desarrollo vegetativo. Aplicando este abordaje se han detectado tres nuevos mutantes. Uno de ellos con el desarrollo radicular afectado y los otros dos con un fenotipo compacto en la parte aérea. En todos ellos se ha realizado la caracterización fenotípica in vitro e in vivo y la caracterización genética. Se ha determinado el tipo de mutación, el número de insertos integrados y, combinando ambos análisis, la presencia de cosegregación entre algún inserto T-DNA con el fenotipo mutante. A partir de esta información se podrá iniciar las tareas para poder identificar el gen responsable de cada mutación.[EN] Studying the function of genes that play a key role in vegetative development and its interaction with the environment provides important basic as well as applied information. A good approach for this purpose is the generation of mutants and its subsequent characterization. If T-DNA is used as a mutagenic agent, besides generating a mutant, the affected gene remains tagged by the insert which sequence is known, which makes the cloning process simpler and faster. In this work, a total of 40 T-DNA lines of Solanum lycopersicum variety Money Maker transformed with an enhancer trap have been studied. For this purpose, plant tissue culture and greenhouse techniques have been used. The main aim of this project was to find mutants affected in the vegetative development. Applying this procedure three new mutants have been identified. One affected in the root development, and two showing an aerial compact phenotype. For all of them, in vitro and in vivo phenotypic characterization as well as genetic characterization was carried out. The type of mutation and the number of inserts has been determined for each line. And by combining this two analysis, the determination of cosegregation between a T-DNA insert with the mutant phenotype has also been studied. This information constitutes the foundation for further investigation on the gene involved in each mutation.Quesada Traver, C. (2018). Identificación y caracterización de mutantes de tomate con alteraciones en el desarrollo vegetativo. http://hdl.handle.net/10251/109091TFG

Regulatory circuits involving bud dormancy factor PpeDAM6

DORMANCY-ASSOCIATED MADS-BOX (DAM) genes have recently emerged as key potential regulators of the dormancy cycle and climate adaptation in perennial species. Particularly, PpeDAM6 has been proposed to act as a major repressor of bud dormancy release and bud break in peach (Prunus persica). PpeDAM6 expression is downregulated concomitantly with the perception of a given genotype-dependent accumulation of winter chilling time, and the coincident enrichment in H3K27me3 chromatin modification at a specific genomic region. We have identified three peach BASIC PENTACYSTEINE PROTEINs (PpeBPCs) interacting with two GA-repeat motifs present in this H3K27me3- enriched region. Moreover, PpeBPC1 represses PpeDAM6 promoter activity by transient expression experiments. On the other hand, the heterologous overexpression of PpeDAM6 in European plum (Prunus domestica) alters plant vegetative growth, resulting in dwarf plants tending toward shoot meristem collapse. These alterations in vegetative growth of transgenic lines associate with impaired hormone homeostasis due to the modulation of genes involved in jasmonic acid, cytokinin, abscisic acid, and gibberellin pathways, and the downregulation of shoot meristem factors, specifically in transgenic leaf and apical tissues. The expression of many of these genes is also modified in flower buds of peach concomitantly with PpeDAM6 downregulation, which suggests a role of hormone homeostasis mechanisms in PpeDAM6-dependent maintenance of floral bud dormancy and growth repression

Models for a molecular calendar of bud-break in fruit trees

Fruit tree crops adapt their phenology to seasonality by finely-tuned mechanisms that perceive environmental day-length and chilling to elaborate a diverse and plastic response, with impact on plant survival and fruit production. These plants use the molecular clock and a prolonged succession of epigenetic events, usually involving the participation of MADS-box genes, for respectively sensing changes in photoperiodicity and accumulated chilling time. Both mechanisms overlap and interact in key processes requiring a precise developmental timing, such as flowering and dormancy, and jointly constitute a molecular calendar, with the ability to report about the arrival of the warm season and winter completion. In this review, we address recent progress in the molecular calendar of fruit trees and the molecular models Arabidopsis and Populus, with a focus on epigenetic-related mechanisms, which provide the regulatory plasticity, reversibility and specificity required for seasonal phenological processes

Evolutionary origin and functional specialization of Dormancy‑Associated MADS box (DAM) proteins in perennial crops

Background: Bud dormancy is a phenological adaptation of temperate perennials that ensures survival under winter temperature conditions by ceasing growth and increasing cold hardiness. SHORT VEGETATIVE PHASE (SVP)-like factors, and particularly a subset of them named DORMANCY-ASSOCIATED MADS-BOX (DAM), are master regulators of bud dormancy in perennials, prominently Rosaceae crops widely adapted to varying environmental conditions. Results: SVP-like proteins from recently sequenced Rosaceae genomes were identifed and characterized using sequence, phylogenetic and synteny analysis tools. SVP-like proteins clustered in three clades (SVP1–3), with known DAM proteins located within SVP2 clade, which also included Arabidopsis AGAMOUS-LIKE 24 (AthAGL24). A more detailed study on these protein sequences led to the identifcation of a 15-amino acid long motif specifc to DAM proteins, which afected protein heteromerization properties by yeast two-hybrid system in peach PpeDAM6, and the unexpected fnding of predicted DAM-like genes in loquat, an evergreen species lacking winter dormancy. DAM gene expression in loquat trees was studied by quantitative PCR, associating with inforescence development and growth in varieties with contrasting fowering behaviour. Conclusions: Phylogenetic, synteny analyses and heterologous overexpression in the model plant Arabidopsis thaliana supported three major conclusions: 1) DAM proteins might have emerged from the SVP2 clade in the Amyg‑daloideae subfamily of Rosaceae; 2) a short DAM-specifc motif afects protein heteromerization, with a likely efect on DAM transcriptional targets and other functional features, providing a sequence signature for the DAM group of dormancy factors; 3) in agreement with other recent studies, DAM associates with inforescence development and growth, independently of the dormancy habit

Structure and Expression of Bud Dormancy-Associated MADS-Box Genes (DAM) in European Plum

Bud dormancy in temperate perennials ensures the survival of growing meristems under the harsh environmental conditions of autumn and winter, and facilitates an optimal growth and development resumption in the spring. Although the molecular pathways controlling the dormancy process are still unclear, DORMANCY-ASSOCIATED MADS-BOX genes (DAM) have emerged as key regulators of the dormancy cycle in different species. In the present study, we have characterized the orthologs of DAM genes in European plum (Prunus domestica L.). Their expression patterns together with sequence similarities are consistent with a role of PdoDAMs in dormancy maintenance mechanisms in European plum. Furthermore, other genes related to dormancy, flowering and stress response have been identified in order to obtain a molecular framework of these three different processes taking place within the dormant flower bud in this species. This research provides a set of candidate genes to be genetically modified in future research, in order to better understand dormancy regulation in perennial species.Esta investigación fue financiada por el Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA) -FEDER (RFP2015-00015-00, RTA2017-00003-00, RTA2017-00011-C03-01) y el Gobierno de Aragón – Fondo Social Europeo, Unión Europea (Grupo Consolidado A12_17R). CQ-T fue financiado por una beca cofinanciada por el Fondo Social Europeo y el Instituto Valenciano de Investigaciones Agrarias (IVIA). BG contó con el apoyo de una beca del Consejo Nacional de Ciencia y Tecnología de México (CONACYT, 471839). AL fue financiado por una beca del Ministerio de Ciencia, Innovación y Universidades del Gobierno de España.Publishe