04070nam a2200205 a 450000100080000000500110000800800410001910000170006024501060007726000160018330000110019950001690021052033720037965000180375165000110376965000190378065000220379965000210382165300220384220669802017-03-14       2016    bl uuuu  m   00u1 u    #d1 aCOTTA, M. G.  aMolecular mechanisms in the first step of aba-mediated response in Coffea ssp.h[electronic resource]  a2016.c2016  a169 p.  aTese (doutorado Biotecnologia Vegetal) - Universidade Federal de Lavras, Lavras, MG. Orientador: Alan Carvalho Andrade. Coorientador: Pierre Roger René Marraccini.  aAbscisic acid (ABA) pathway is a phytohormone universally conserved in land plants which coordinates several aspects of the plant response to water deficit such as root architecture, seed dormancy and regulation of stomatal closure. A mechanism of ABA signal transduction has been proposed, evolving intracellular ABA receptors (PYR/PYL/RCARs) interacting with PP2Cs phosphatases and SnRK2 protein kinases regulating this tripartite protein system. The goal of this study was to identify and characterize for the first time the orthologs genes of this tripartite system in Coffea. For this purpose, protein sequences from Arabidopsis, citrus, rice, grape, tomato and potato were chosen as query to search orthologous genes in the Coffee Genome Hub (http://coffee-genome.org/). Differential expression in tissues as leaves, seeds, roots and floral organs waschecked through in silico analyses. In vivo gene expression analyses were also performed by RT-qPCR in leaves and roots of drought-tolerant (DT 14, 73 and 120) and drought-susceptible (DS 22) C. canephora Conilon clones submitted (or not) to drought. The expression profiles of the tripartite system CcPYL-PP2C-SnRK2 genes were also analyzed in leaves of C. arabica and C. canephora plants growing under hydroponic condition and submitted to ABA exogenous treatment (500 ?M). This approach allowed the identification and characterization of 24 candidate genes (9 PYL/RCARs, 6 PP2Cs and 9 SnRK2s) in C. canephora genome. The protein motifs identified in predict coffee sequences enabled characterize these genes as family’s members of PYL/RCARs receptors, PP2Cs phosphatases or SnRK2 kinases of the ABA response pathway. These families were functionally annotated in the C. canephora genome. In vivo analyses revealed that eight genes are upregulated under drought conditions in both leaves and roots tissues. Among them, three genes coding phosphatases were expressed in all (DT and DS) clones therefore suggesting that they were activated as a general response to cope with drought stress. However, two other phosphatase coding genes were up-regulated only in the DT clones, suggesting that they constituted key-genes for drought tolerance in these clones. The DT clones also showed differential gene expression profiles for five other genes therefore reinforcing the idea that multiple biological mechanisms are involved drought tolerance in C. canephora. In response to exogenous ABA, 17 genes were expressed in leaves of C. canephora and C. arabica plants. Several genes were differentially expressed in the DT clone 14 either in control condition or after 24h ABA treatment. Under control conditions, five genes were higher expressed in the C. canephora and C. arabica DT plants. The kinase CcSnRK2.6 was highlighted as gene specifically expressed in the C. canephora plants (DT and DS) after 72h of ABA treatment. Overall, it was observed that ABA signaling pathway is delayed in the DS C. arabica Rubi. Those molecular evidences corroborated with microscopies analyses which showed that the DT clone 14 was more efficient to control stomatal closure than other coffee plants in response to ABA treatment. All these evidences will help us to identify the genetic determinism of drought tolerance through ABA pathway essential to obtain molecular markers that could be used in coffee breeding programs..  aAbscisic acid  aCoffea  aPlant hormones  aÁcido abscísico  aHormonio vegetal  aDéficit hídrico