Project description:The intermediate seed category was defined in the early 1990s using coffee (Coffea arabica) as a model. In contrast to orthodox seeds, intermediate seeds cannot survive complete drying, which is a major constraint for seed storage, for both biodiversity conservation and agricultural purposes. However, intermediate seeds are considerably more tolerant to drying than recalcitrant seeds, which are highly sensitive to desiccation. To gain insight into the mechanisms governing such differences, changes in desiccation tolerance (DT), hormone content and the transcriptome were analysed in developing coffee seeds. Acquisition of DT coincided with a dramatic transcriptional switch characterised by the repression of primary metabolism, photosynthesis and respiration, and the upregulation of genes coding for late embryogenesis abundant (LEA) proteins, heat shock proteins (HSP) and antioxidant enzymes. Analysis of heat-stable proteome in the mature coffee seed confirmed the accumulation of LEA proteins identified at the transcript level. Transcriptome analysis also suggests a major role for ABA and for the transcription factors CaHSFA9, CaDREB2G, CaANAC029, CaPLATZ and CaDOG-like in DT acquisition. The ability of CaHSFA9 and CaDREB2G to trigger HSP gene transcription was validated by Agrobacterium-mediated transformation of coffee somatic embryos.
Project description:In contrast to the desiccation tolerant (DT) ‘orthodox’ seeds, the so-called ‘intermediate’ seeds cannot survive complete drying and are short-lived. All species of the genus Coffea produce intermediate seeds but show a considerable variability for the seed DT level, which may help to decipher the molecular basis of seed DT in plants. We thus led a comparative transcriptome analysis of developing seeds in three coffee species with contrasting seed DT levels. Seeds of all species shared a major transcriptional switch during late maturation that governs a general slow-down of metabolism. However, numerous key stress-related genes, including those coding for the late embryogenesis abundant protein EM6 and the osmosensitive calcium channel ERD4, were upregulated during DT acquisition in the two species with high seed DT, C. arabica and C. eugenioides. By contrast, an upregulation of numerous players of the metabolism, transport and perception of auxin was observed in C. canephora seeds with low DT. Moreover, species with high DT showed a stronger down-regulation of the mitochondrial machinery dedicated to the tricarboxylic acid cycle and oxidative phosphorylation. Accordingly, respiration measurements during seed dehydration demonstrated that intermediate seeds with the highest DT levels are better prepared to cease respiration and avoid oxidative stresses.
Project description:Desiccation tolerance (DT) allowed seed plants to conquer ecosystems with long periods of limited water availability. This adaptive features allows seeds to remain dried for very long times without losing their ability to germinate. There is little information about all the signaling components required to achieve DT and on how transcription factors (TFs) modulate global DT processes. We performed RNA-seq experiment and carbohydrates profiles of lec1, lec2, fus3 and abi3, as well as their corresponding wild types, at three stages of seed development 15, 17 and 21 DAF (day after open flower) belonging to the seed desiccation period. A complex experimental design approach and regulatory networks prediction were used to identify differentially expressed genes specifically involved in DT process. In order to identify mechanisms involved in the acquisition of DT during seed development, we designed a comparative transcriptomic analysis between the seed desiccation intolerant (DI) mutants lec1-1, abi3-5 and fus3-3, the desiccation tolerant mutant lec2-1 and the desiccation tolerant weak allele of abi3 (abi3-1) with their respective wild type controls. This analysis should allow to identify genes that are differentially expressed in the desiccation intolerant mutants respect to tolerant mutants and WT controls.
Project description:Desiccation tolerance (DT) allowed seed plants to conquer ecosystems with long periods of limited water availability. This adaptive features allows seeds to remain dried for very long times without losing their ability to germinate. There is little information about all the signaling components required to achieve DT and on how transcription factors (TFs) modulate global DT processes. We performed RNA-seq experiment and carbohydrates profiles of lec1, lec2, fus3 and abi3, as well as their corresponding wild types, at three stages of seed development 15, 17 and 21 DAF (day after open flower) belonging to the seed desiccation period. A complex experimental design approach and regulatory networks prediction were used to identify differentially expressed genes specifically involved in DT process.
Project description:The combination of robust physiological models with “omics” studies holds promise for the discovery of genes and pathways linked to how organisms deal with drying. Here we used a transcriptomics approach in combination with an in vivo physiological model of re-establishment of desiccation tolerance (DT) in Arabidopsis thaliana seeds. We show that the incubation of desiccation-sensitive (DS) germinated Arabidopsis seeds in a polyethylene glycol (PEG) solution re-induces the mechanisms necessary for expression of DT. Based on a SNP-tile array gene expression profile, our data indicates that the re-establishment of DT, in this system, is related to a programmed reversion from a metabolic active to a quiescent state similar to prior to germination. Our findings show that transcripts of germinated seeds after the PEG treatment are dominated by those encoding LEA, seed storage and dormancy-related proteins. On the other hand, a massive repression of genes belonging to many other classes such as photosynthesis, cell wall modification and energy metabolism occurs in parallel. Furthermore, comparison with a similar system for Medicago truncatula reveals a significant overlap between the two transcriptomes. Such overlap may highlight core mechanisms and key regulators of the trait DT. Taking into account the availability of the many genetic and molecular resources for Arabidopsis, the described system may prove useful for unraveling DT in higher plants. Desiccation-sensitive seeds vs. desiccation-tolerant seeds in the same developmental stage in triplicate.
Project description:The combination of robust physiological models with “omics” studies holds promise for the discovery of genes and pathways linked to how organisms deal with drying. Here we used a transcriptomics approach in combination with an in vivo physiological model of re-establishment of desiccation tolerance (DT) in Arabidopsis thaliana seeds. We show that the incubation of desiccation-sensitive (DS) germinated Arabidopsis seeds in a polyethylene glycol (PEG) solution re-induces the mechanisms necessary for expression of DT. Based on a SNP-tile array gene expression profile, our data indicates that the re-establishment of DT, in this system, is related to a programmed reversion from a metabolic active to a quiescent state similar to prior to germination. Our findings show that transcripts of germinated seeds after the PEG treatment are dominated by those encoding LEA, seed storage and dormancy-related proteins. On the other hand, a massive repression of genes belonging to many other classes such as photosynthesis, cell wall modification and energy metabolism occurs in parallel. Furthermore, comparison with a similar system for Medicago truncatula reveals a significant overlap between the two transcriptomes. Such overlap may highlight core mechanisms and key regulators of the trait DT. Taking into account the availability of the many genetic and molecular resources for Arabidopsis, the described system may prove useful for unraveling DT in higher plants.
Project description:Desiccation tolerance (DT) is the capacity to withstand total loss of cellular water. This property is acquired during seed filling and lost just after germination. However, in many species, a germinated seed can regain DT under adverse conditions such as osmotic stress. We discovered that the germinated seeds of the abscisic acid insensitive 5 (Mtabi5) mutant of Medicago truncatula lost their ability to re-establish DT during osmotic stress. To characterize the molecular processes that are influenced by MtABI5 during the re-establishment of DT tolerance, a transcriptome analysis was performed on the protruded radicles of germinated Mtabi5 mutants and wilt type before and after an osmotic treatment.
Project description:Mature seeds of Arabidopsis thaliana are desiccation tolerant, but they lose DT while progressing to germination. Yet, there is a small developmental window during which DT can be rescued by treatment with polyethylene glycol (PEG). We used microarrays to identify relevant genes in the re-establishment of desiccation tolerance by PEG.
Project description:Coffee leaf miner is an important plague in coffee crops. Using subtracted cDNA libraries and nylon filter arrays, we analyzed the expression profile of 1536 expressed sequence tags (ESTs) of coffee plants from an hybrid progeny (C. arabica x C. racemosa), containg resistant (R) and susceptible plants (S) to the infestation of coffee leaf miner. Leaf discs were collected from non-infested plants (R control - RC; S control - SC), infested plants after moth oviposition (R oviposition - Ro; S oviposition - So) and infested after larvar eclosion (R eclosion - Re; S eclosion - Se). Isolation and characterization of Coffea genes induced during coffee leaf miner (Leucoptera coffeella) infestation. Plant Science 169(2):351-360 Keywords: ordered