La régulation transcriptionnelle dépendant de l'éthylène. Caractérisation fonctionnelle d'un cofacteur transcriptionnel du type MBF1 et d'un facteur de transcription de la famille des ERF chez la tomate.

La phytohormone éthylène contrôle des étapes importantes de la vie de la plante comme la germination, la maturation des fruits et la réponse aux stress biotiques et abiotiques. L'éthylène exerce ses effets physiologiques en modulant l'expression de gènes cibles par l'intermédiaire des facteurs de transcription. De ce fait, la caractérisation des facteurs de transcription associés à la réponse à l'éthylène est une étape fondamentale pour la compréhension des processus régulés physiologiques régulés par cette hormone. Ce travail décrit l'isolement et la caractérisation fonctionnelle de la famille de coactivateurs transcriptionnels de type MBF1 (Multiproteine Bridging Factor 1) et d'un facteur de transcription de la famille des ERF (Ethylene Response Factor) chez la tomate. Notre étude montre que la famille des MBF1 chez la tomate comprend quatre gènes (LeMBF1a, LeMBF1b, LeMBF1c et LeMBF1/ER24) qui sont tous capables de complémenter un mutant de la levure indiquant qu'il y a une parfaite conservation fonctionnelle de ces gènes de la levure aux plantes supérieures. Les études d'expression révèlent un profil spécifique pour ER24 qui est fortement exprimé en réponse à l'éthylène, au cours de la maturation des fruits et en réponse à différents stress abiotiques. Le criblage par double-hybride de la levure a permis d'identifier plusieurs partenaires des protéines MBF1 chez la tomate parmi lesquels des protéines associées à la maturation et aux stress abiotiques. La surexpression du gène LeMBF1/ER24 chez Arabidopsis confère une tolérance au stress thermique. Par ailleurs, nous avons isolé et caractérisé le gène LeERF2 de la tomate qui appartient à la grande famille des ERF (Ethylene Response Factor) codant pour des facteurs de transcription spécifiques aux plantes. L'expression du gène LeERF2 est induite durant la maturation des fruits et en réponse à la blessure. La surexpression de ce gène dans la tomate induit une germination précoce des graines et affecte la formation du crochet apical, un processus de développement régulé par l'éthylène. L'expression du gène de la mannanase2, un marqueur de la germination, est fortement induite dans les graines transgéniques sur-exprimant le gène LeERF2 suggèrant que LeERF2 stimule la germination à travers l'induction de ce gène. Ce travail a permis d'identifier de nouveaux acteurs de la régulation transcriptionnelle associée à l'action de l'éthylène et d'apporter un nouvel éclairage sur le mode d'action de cette hormone

The mitochondrial elongation factor LeEF-Tsmt is regulated during tomato fruit ripening and upon wounding and ethylene treatment

A gene encoding an elongation factor LeEF-Tsmt that participates in the protein synthesis process in mitochondria shows strong expression in ripening fruit as compared to other organs. It is strongly up-regulated during the first stages of the ripening process in parallel with the climacteric rise in respiration. LeEF-Tsmt expression is stimulated by ethylene, wounding and high temperature but ethylene-insensitive mutants exhibit normal expression. Transgenic fruit have been generated in which LeEF-Tsmt has been constitutively up- and down-regulated. Surprisingly, altering the expression of the gene by genetic transformation with antisense and sense LeEF-Tsmt constructs did not affect the pattern of respiration and ethylene production during ripening and upon wounding. In addition, expression of the alternative oxidase gene which is known to play an important role in respiratory climacteric was not affected. Possible reasons for the absence of effect on respiration of variations of LeEF-Tsmt gene expression are discussed

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

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

The MADS transcription factor XAL2/AGL14 modulates auxin transport during Arabidopsis root development by regulating PIN expression

Elucidating molecular links between cell-fate regulatory networks and dynamic patterning modules is a key for understanding development. Auxin is important for plant patterning, particularly in roots, where it establishes positional information for cell-fate decisions. PIN genes encode plasma membrane proteins that serve as auxin efflux transporters; mutations in members of this gene family exhibit smaller roots with altered root meristems and stem-cell patterning. Direct regulators of PIN transcription have remained elusive. Here, we establish that a MADS-box gene (XAANTAL2, XAL2/AGL14) controls auxin transport via PIN transcriptional regulation during Arabidopsis root development; mutations in this gene exhibit altered stem-cell patterning, root meristem size, and root growth. XAL2 is necessary for normal shootward and rootward auxin transport, as well as for maintaining normal auxin distribution within the root. Furthermore, this MADS-domain transcription factor upregulates PIN1 and PIN4 by direct binding to regulatory regions and it is required for PIN4-dependent auxin response. In turn, XAL2 expression is regulated by auxin levels thus establishing a positive feedback loop between auxin levels and PIN regulation that is likely to be important for robust root patterning.This work was supported by grants from CONACYT, México: Red Tematica de Investigacion: ‘Complejidad, Ciencia y Sociedad’ (124909; ERAB; BGP; AGA) and 81542 and 105678 (ERAB), 167705 (AGA), 152649 (MPS), 81433 (BGP), 177739 (SF) and 127957 (JGD), from PAPIIT-UNAM, IN204011-3 (BGP), IN229009-3 (ERAB), IN226510-3 (AGA), IB201212 (MPS), and IN204312 (JGD), from the Spanish Government BFU2012-33746 (SP) and from the National Science Foundation (NSF-IOS) 0820648 (ASM). ERAB acknowledges the support of the Miller Institute for Basic Research in Science, University of California, Berkeley while spending a sabbatical leave in the laboratory of Chelsea Specht at UC-B.Peer reviewe

Flower Development

Flowers are the most complex structures of plants. Studies of Arabidopsis thaliana, which has typical eudicot flowers, have been fundamental in advancing the structural and molecular understanding of flower development. The main processes and stages of Arabidopsis flower development are summarized to provide a framework in which to interpret the detailed molecular genetic studies of genes assigned functions during flower development and is extended to recent genomics studies uncovering the key regulatory modules involved. Computational models have been used to study the concerted action and dynamics of the gene regulatory module that underlies patterning of the Arabidopsis inflorescence meristem and specification of the primordial cell types during early stages of flower development. This includes the gene combinations that specify sepal, petal, stamen and carpel identity, and genes that interact with them. As a dynamic gene regulatory network this module has been shown to converge to stable multigenic profiles that depend upon the overall network topology and are thus robust, which can explain the canalization of flower organ determination and the overall conservation of the basic flower plan among eudicots. Comparative and evolutionary approaches derived from Arabidopsis studies pave the way to studying the molecular basis of diverse floral morphologies