Les puces à ADN : outils pour une nouvelle vision de la diversité et des ressources génétiques

À l'aube du troisième millénaire, on assiste à la fusion de différents domaines du savoir pour créer de nouvelles disciplines de recherche. Ainsi, biologie et informatique se sont mariées pour produire la bio-informatique du côté conceptuel et, entre autres, les biopuces au niveau de l'instrumentation. On assiste à un changement d'échelle au niveau de la quantité de locus et du nombre d'échantillons à analyser, et donc de données à acquérir et à traiter. Plus qu'optimiser les techniques de biologie moléculaire (sélection assistée par marqueurs, par ex.), instruments de la genétique, les technologies à grand débit sur lesquelles est fondée la génomique ont dévoilé de nouvelles perspectives et entraînent un changement de paradigme, ouvrant l'ère de la post-génomique. D'analytique, l'approche scientifique évoluera vers une méthodologie systémique. Notre vision, essentiellement descriptive, de la diversité et des ressources génétiques promet de devenir réellement prédictive. (Résumé d'auteur

Regulation of tomato fruit ripening

Fruit ripening is a sophisticatedly orchestrated developmental process, unique to plants, that results in major physiological and metabolic changes, ultimately leading to fruit decay and seed dispersal. Because of their strong impact on fruit nutritional and sensory qualities, the ripeningassociated changes have been a matter of sustained investigation aiming at unravelling the molecular and genetic basis of fruit ripening. Tomato rapidly emerged as the model of choice for fleshy fruit research and a wealth of genetic resources and genomics tools have been developed, providing new entries into the regulatory mechanisms involved in the triggering and coordination of the ripening process. Some of the key components participating in the control of tomato fruit ripening have been uncovered, but our knowledge of the network of signalling pathways engaged in this complex developmental process remains fragmentary. This review highlights the main advances and emphasizes issues still to be addressed using the rapidly developing ‘omics’ approaches

Silencing Sl-EBF1 and Sl-EBF2 expression causes constitutive ethylene response phenotype, accelerated plant senescence, and fruit ripening in tomato

The hormone ethylene regulates a wide range of plant developmental processes and EBF (EIN3-binding F-box) proteins were shown to negatively regulate the ethylene signalling pathway via mediating the degradation of EIN3/EIL proteins. The present study reports on the identification of two tomato F-box genes, Sl-EBF1 and Sl-EBF2 from the EBF subfamily. The two genes display contrasting expression patterns in reproductive and vegetative tissues and in response to ethylene and auxin treatment. Sl-EBF1 and Sl-EBF2 genes are actively regulated at crucial stages in the development of the reproductive organs. Their dynamic expression in flowers during bud-to-anthesis and anthesis-to-post-anthesis transitions, and at the onset of fruit ripening, suggests their role in situations where ethylene is required for stimulating flower opening and triggering fruit ripening. VIGS-mediated silencing of a single tomato EBF gene uncovered a compensation mechanism that tends to maintain a threshold level of Sl-EBF expression via enhancing the expression of the second Sl-EBF gene. In line with this compensation, tomato plants silenced for either of the Sl-EBF genes were indistinguishable from control plants, indicating functional redundancy among Sl-EBF genes. By contrast, co-silencing of both Sl-EBFs resulted in ethylene-associated phenotypes. While reports on EBF genes to date have focused on their role in modulating ethylene responses in Arabidopsis, the present study uncovered their role in regulating crucial stages of flower and fruit development in tomato. The data support the hypothesis that protein degradation via the ubiquitin/26S proteasome pathway is a control point of fruit ripening and open new leads for engineering fruit quality

Over-expression of a chimeric gene of the transcriptional co-activator MBF1 fused to the EAR repressor motif causes developmental alteration in Arabidopsis and tomato

Transcriptional co-activators of the Multiprotein Bridging Factor1 (MBF1) type belong to a multigenic family that encode key components of the machinery controlling gene expression by communicating between transcription factors and the basal transcription machinery. Knocking-down the expression of one member of the family has proved difficult probably due to functional redundancy. We show here that a fusion of SlER24, an MBF1 type gene of tomato, to the Ethylene-responsive element-binding associated Amphiphilic Repression (EAR) motif is capable of slowing down significantly the expression of the GFP protein driven by a synthetic ethylene-responsive GCC-rich promoter in a single cell transient expression system. A fusion of AtMBF1c of Arabidopsis to EAR, driven by the 35S promoter, caused a reduction of the percentage of seed germination and dwarfism of the plant. Similar fusion with the SlER24 of tomato in the MicroTom cultivar induced a delay of seed germination and no obvious effect on plant growth. Besides giving information on the role of the MBF1 genes in plant development, this study demonstrates that the EAR strategy is efficient not only for regular transcription factors as demonstrated so far, but also in the case of co-activators known to not bind directly to DNA

Functional analysis and binding affinity of tomato ethylene response factors provide insight on the molecular bases of plant differential responses to ethylene

Background : The phytohormone ethylene is involved in a wide range of developmental processes and in mediating plant responses to biotic and abiotic stresses. Ethylene signalling acts via a linear transduction pathway leading to the activation of Ethylene Response Factor genes (ERF)which represent one of the largest gene families of plant transcription factors. How an apparently simple signalling pathway can account for the complex and widely diverse plant responses to ethylene remains yet an unanswered question. Building on the recent release of the complete tomato genome sequence, the present study aims at gaining better insight on distinctive features among ERF proteins. Results : A set of 28 cDNA clones encoding ERFs in the tomato (Solanum lycopersicon) were isolated and shown to fall into nine distinct subclasses characterised by specific conserved motifs most of which with unknown function. In addition of being able to regulate the transcriptional activity of GCC-box containing promoters, tomato ERFs are also shown to be active on promoters lacking this canonical ethylene-responsive-element. Moreover, the data reveal that ERF affinity to the GCC-box depends on the nucleotide environment surrounding this cis-acting element. Site-directed mutagenesis revealed that the nature of the flanking nucleotides can either enhance or reduce the binding affinity, thus conferring the binding specificity of various ERFs to target promoters. Based on their expression pattern, ERF genes can be clustered in two main clades given their preferential expression in reproductive or vegetative tissues. The regulation of several tomato ERF genes by both ethylene and auxin, suggests their potential contribution to the convergence mechanism between the signalling pathways of the two hormones. Conclusions : The data reveal that regions flanking the core GCC-box sequence are part of the discrimination mechanism by which ERFs selectively bind to their target promoters. ERF tissue-specific expression combined to their responsiveness to both ethylene and auxin bring some insight on the complexity and fine regulation mechanisms involving these transcriptional mediators. All together the data support the hypothesis that ERFs are the main component enabling ethylene to regulate a wide range of physiological processes in a highly specific and coordinated manner

Sl-ERF2, a Tomato Ethylene Response Factor Involved in Ethylene Response and Seed Germination

Ethylene response factors (ERFs) are plant transcriptional regulators mediating ethylene-dependent gene expression via binding to the GCC motif found in the promoter region of ethylene-regulated genes. We report here on the structural and functional characterization of the tomato Sl-ERF2 gene that belongs to a distinct class of the large ERF gene family. Both spliced and unspliced versions of Sl-ERF2 transcripts were amplified from RNA samples and the search in the public tomato expressed sequence tag (EST) database confirmed the existence of the two transcript species in a number of cDNA libraries. The unspliced transcript contains two open reading frames yielding two hypothetical proteins, a small highly truncated version lacking the APETALA2 domain and a bigger protein lacking the N-terminal MCGGAAII/L consensus peptide specific to ERF members from subfamily IV. Nevertheless, functional Sl-ERF2 protein may only derive from spliced transcripts since, depending on the tissue, the level of the spliced transcript is much higher than that of the unspliced transcript. Sl-ERF2 is expressed in all plant tissues tested, though its transcript accumulates preferentially in germinating seeds and ripening fruit. Overexpression of the Sl-ERF2 gene in transgenic tomato lines results in premature seed germination and enhanced hook formation of darkgrown seedlings, which is indicative of increased ethylene sensitivity. The expression of the mannanase2 gene is upregulated in Sl-ERF2-overexpressing seeds, suggesting that Sl-ERF2 stimulates seed germination through the induction of the mannanase2 gene. It is noteworthy that the exaggerated hook phenotype is abolished when ethylene perception is blocked, strongly suggesting that Sl-ERF2 requires other ethylene-dependent components to impact the hook formation process

UV-C radiation modifies the ripening and accumulation of ethylene response factor (ERF) transcripts in tomato fruit

Ultraviolet-C (UV-C) radiation is used as a postharvest treatment to prolong the shelf life of fruit. However, this stressful process may also affect ethylene production and, consequently, the expression of genes encoding ethylene response factors (ERFs). To test this hypothesis, MicroTom tomatoes harvested at the breaker stage were subjected to: 1 – application of 3.7 kJ m-2 UV-C radiation, 2 – application of 2 µLL-1 1-methylcyclopropene (1-MCP) followed by UV-C radiation; and 3 – without 1-MCP or UV-C (control treatment). After treatment all fruit were stored for 12 d at 21 +/- 2C° and 80 +/- 5% relative humidity (RH). Although UV-C radiation increased ACC oxidase transcripts and stimulated ethylene production, the ripening evolution was delayed. Fruit treated with UV-C showed lower accumulation of lycopene, b-carotene, lutein + zeaxanthin and d-tocopherol; but retained higher levels of chlorogenic acid, r-coumaric acid and quercetin after 6 d. Additionally, UV-C treated fruit had higher contents of polyamines (putrescine and spermidine). Among the 14 ERFs studied, 11 (Sl-ERF A.1, Sl-ERF A.3,Sl-ERF B.1, Sl-ERF B.2, Sl-ERF B.3, Sl-ERF C.6, Sl-ERF D.1, Sl-ERF D.3, Sl-ERF E.1, Sl-ERF F.5, Sl-ERF G.2) exhibited increased transcript accumulation, 2 ERFs (Sl-ERF E.2 and Sl-ERF E.4) showed decreased transcript accumulation and only 1 ERF (Sl-ERF E.3) was not significantly affected by UV-C treatment. As expected, the transcript profiles of 1-MCP and/or UV-C-treated tomatoes demonstrate that ethylene plays an important role in the expression of ERFs. The delay in fruit ripening may be caused by the activation of ERFs that could act as regulators of metabolic pathways during ripening. However, this hypothesis needs to be better tested. In conclusion, a relationship has been established between UV-C treatment and ripening delay, correlated to changes in 13 ERF transcripts evaluated during postharvest treatment

Genome-Wide Identification, Functional Analysis and Expression Profiling of the Aux/IAA Gene Family in Tomato

Auxin is a central hormone that exerts pleiotropic effects on plant growth including the development of roots, shoots, flowers and fruit. The perception and signaling of the plant hormone auxin rely on the cooperative action of several components,among which auxin/indole-3-acetic acid (Aux/IAA) proteins play a pivotal role. In this study, we identified and comprehensively analyzed the entire Aux/IAA gene family in tomato (Solanum lycopersicum), a reference species for Solanaceae plants, and the model plant for fleshy fruit development. Functional characterization using a dedicated single cell system revealed that tomato Aux/IAA proteins function as active repressors of auxin-dependent gene transcription, with, however, different Aux/IAA members displaying varying levels of repression. Phylogenetic analysis indicated that the Aux/IAA gene family is slightly contracted in tomato compared with Arabidopsis, with a lower representation of non-canonical proteins. Sl-IAA genes display distinctive expression pattern in different tomato organs and tissues, and some of them display differential responses to auxin and ethylene, suggesting that Aux/IAAs may play a role in linking both hormone signaling pathways. The data presented here shed more light on Sl-IAA genes and provides new leads towards the elucidation of their function during plant development and in mediating hormone cross-talk

Numerical study of the smoke plume of the Jerada thermal power plant Morocco

Abstract The main goal of this study is firstly to model the phenomenon of advection and diffusion of the gaseous residues of coal combustion. On the other hand, to study the dispersion of this pollution by varying the parameters like the wind and the resources of pollution, where there are four emissions: SO2, CO, NO2 and PM10. The present model will compare estimating concentrations with the results found by the contribution of the thermal power plant of Jerada city, where the American Environmental Regulatory Model is used. The results suggest that SO2, PM10, NO2 and CO concentration simulated by turbulent k-ε model in combination with the transport model of diluted species, at the 3h and 24h intervals, greater than their respective observed concentrations compared with the American Environmental Regulatory Model. The concentrations of the model found are very high and exceed the limit values under the study conditions. Indeed, the simulated model was used in 2D with a very low wind speed (10−4 m/s) and in the absence of gravity, hence the total neglect of altitude because the two sources of pollution are at the height of 120 m

Functional characterization of a melon alcohol acyl-transferase gene family involved in the biosynthesis of ester volatiles. Identification of the crucial role of a threonine residue for enzyme activity

Volatile esters, a major class of compounds contributing to the aroma of many fruit, are synthesized by alcohol acyl-transferases (AAT). We demonstrate here that, in Charentais melon (Cucumis melo var. cantalupensis), AAT are encoded by a gene family of at least four members with amino acid identity ranging from 84% (Cm-AAT1/Cm-AAT2) and 58% (Cm-AAT1/Cm-AAT3) to only 22% (Cm-AAT1/Cm-AAT4). All encoded proteins, except Cm-AAT2, were enzymatically active upon expression in yeast and show differential substrate preferences. Cm-AAT1 protein produces a wide range of short and long-chain acyl esters but has strong preference for the formation of E-2-hexenyl acetate and hexyl hexanoate. Cm-AAT3 also accepts a wide range of substrates but with very strong preference for producing benzyl acetate. Cm-AAT4 is almost exclusively devoted to the formation of acetates, with strong preference for cinnamoyl acetate. Site directed mutagenesis demonstrated that the failure of Cm-AAT2 to produce volatile esters is related to the presence of a 268-alanine residue instead of threonine as in all active AAT proteins. Mutating 268-A into 268-T of Cm-AAT2 restored enzyme activity, while mutating 268-T into 268-A abolished activity of Cm-AAT1. Activities of all three proteins measured with the prefered substrates sharply increase during fruit ripening. The expression of all Cm-AAT genes is up-regulated during ripening and inhibited in antisense ACC oxidase melons and in fruit treated with the ethylene antagonist 1-methylcyclopropene (1-MCP), indicating a positive regulation by ethylene. The data presented in this work suggest that the multiplicity of AAT genes accounts for the great diversity of esters formed in melon