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The enzymatic hydrolysis of cellulose into glucose, referred to as saccharification, is severely hampered by lignins. Here, we analyzed transgenic poplars (Populus tremula × Populus alba) expressing the Brachypodium (Brachypodium distachyon) p-coumaroyl-Coenzyme A monolignol transferase 1 (BdPMT1) gene driven by the Arabidopsis (Arabidopsis thaliana) Cinnamate 4-Hydroxylase (AtC4H) promoter in the wild-type (WT) line and in a line overexpressing the Arabidopsis Ferulate 5-Hydroxylase (AtF5H). BdPMT1 encodes a transferase which catalyzes the acylation of monolignols by p-coumaric acid (pCA). Several BdPMT1-OE/WT and BdPMT1-OE/AtF5H-OE lines were grown in the greenhouse, and BdPMT1 expression in xylem was confirmed by RT-PCR. Analyses of poplar stem cell walls (CWs) and of the corresponding purified dioxan lignins (DLs) revealed that BdPMT1-OE lignins were as p-coumaroylated as lignins from C3 grass straws. For some transformants, pCA levels reached 11 mg·g−1 CW and 66 mg·g−1 DL, exceeding levels in Brachypodium or wheat (Triticum aestivum) samples. This unprecedentedly high lignin p-coumaroylation affected neither poplar growth nor stem lignin content. Interestingly, p-coumaroylation of poplar lignins was not favored in BdPMT1-OE/AtF5H-OE transgenic lines despite their high frequency of syringyl units. However, lignins of all BdPMT1-OE lines were structurally modified, with an increase of terminal unit with free phenolic groups. Relative to controls, this increase argues for a reduced polymerization degree of BdPMT1-OE lignins and makes them more soluble in cold NaOH solution. The p-coumaroylation of poplar samples improved the saccharification yield of alkali-pretreated CW, demonstrating that the genetically driven p-coumaroylation of lignins is a promising strategy to make wood lignins more susceptible to alkaline treatments used during the industrial processing of lignocellulosics., The expression of a grass p-coumaroyl-CoA:monolignol transferase induces high p-coumaroylation of poplar lignins and better saccharification of alkali-pretreated poplar wood without growth penalty.

Trees can generate large mechanical stresses at the stem periphery to control the orientation of their axes. This key factor in the biomechanical design of trees, named “maturation stress”, occurs in wood fibres during cellular maturation when their secondary cell wall thickens. In this study, the spatial and temporal stiffening kinetics of the different cell wall layers were recorded during fibre maturation on a sample of poplar tension wood using atomic force microscopy. The thickening of the different layers was also recorded. The stiffening of the CML, S 1 and S 2 -layers was initially synchronous with the thickening of the S 2 layer and continued a little after the S 2 -layer reached its final thickness as the G-layer begins to develop. In contrast, the global stiffness of the G-layer, which initially increased with its thickening, was almost stable long before it reached its final maximum thickness. A limited radial gradient of stiffness was observed in the G-layer, but it decreased sharply on the lumen side, where the new sub-layers are deposited during cell wall thickening. Although very similar at the ultrastructural and biochemical levels, the stiffening kinetics of the poplar G-layer appears to be very different from that described in maturing bast fibres. Highlight New insights into the changes in mechanical properties within the cell wall of poplar tension wood fibres during maturation have been obtained using atomic force microscopy.

Transgenic poplars (Populus tremula x Populus alba, clone INRA 717-1B4) were produced by introducing the Brachypodium distachyon Bradi2g36910 (BdPMT1) gene driven by the Arabidopsis (Arabidopsis thaliana) Cinnamate 4-Hydroxylase (AtC4H) promoter in the wild-type (WT) line and in a line overexpressing the Arabidopsis Ferulate 5-Hydroxylase (AtF5H). BdPMT1 encodes a transferase which catalyzes the acylation of monolignols by p-coumaric acid (CA). Several BdPMT1- OE/WT and BdPMT1-OE/AtF5H-OE transgenic lines were grown in the greenhouse and BdPMT1 expression in xylem was confirmed by RT-PCR. The analysis of the cell walls (CW) of poplar stems and of corresponding purified dioxan lignins (DL) revealed that the BdPMT1-OE lignins were as p-coumaroylated as the lignins of C3 grass straws. For some transformants, CA levels even reached about 11 mg/g CW and 66 mg/g DL, which by far exceeds those of Brachypodium or wheat samples. This unprecedentedly high p-coumaroylation of poplar lignins affected neither the poplar growth, nor the stem lignin content. By contrast, the transgenic lignins were structurally modified, with an increase of terminal units with free phenolic groups. Relative to controls, this increase argues for a reduced polymerization degree of BdPMT1-OE lignins and makes them more soluble in cold NaOH solution. The p-coumaroylation of poplar samples, up to the levels of C3 grasses, improved the saccharification yield of alkali-pretreated poplar CW. These results establish that the genetically-driven p-coumaroylation of lignins is a promising strategy to make wood lignins more susceptible to the alkaline treatments that can be used during the industrial processing of lignocellulosics.One-sentence summaryThe expression of a grass p-coumaroyl-CoA:monolignol transferase induces a high p-coumaroylation of poplar lignins and a better saccharification of alkali-pretreated poplar wood without growth penalty

International audience; Les arbres atteignent des hauteurs et des durées de vie considérables grâce aux propriétés remarquables de leur bois. En effet, le bois remplit trois fonctions principales : (1) la conduction de l’eau de la racine au houppier, (2) le soutien mécanique de la masse toujours en augmentation de l’arbre en croissance et (3) le stockage de réserves temporaires, capitales pour la pérennité de l’arbre. Chez les angiospermes, les vaisseaux, les fibres et les rayons parenchymateux sont, respectivement, affiliés à ces fonctions [1]. La composition et la structure des parois de ces cellules varient considérablement en fonction des stades de développement et des conditions environnementales. Par exemple, en réponse à des contraintes mécaniques, les angiospermes produisent un bois dit de tension qui se caractérise par la présence d’une couche surnuméraire dans les parois des fibres, appelée couche G, et composée exclusivement de cellulose et de polysaccharides non-cellulosiques [2-3]. Le bois est donc un assemblage complexe des parois secondaires des fibres et vaisseaux, interconnectés aux cellules vivantes de rayons. Afin de s’affranchir de cette grande complexité, les mécanismes moléculaires de la formation du bois doivent être préférentiellement étudiés au niveau cellulaire.Ici, nous présentons le développement d’une méthode de microphénotypage non destructrice basée sur la microspectroscopie ATR-FTIR. Nous avons appliqué cette technique sur du bois de peupliers inclinés pour induire la formation de bois de tension. Grâce aux analyses d’images hyperspectrales obtenues, nous avons montré que(i) les couches G sont principalement composées de cellulose et de polysaccharides non cellulosiques,(ii) les lignines des parois cellulaires des vaisseaux sont plus riches en unités G alors que celles des fibres sont plus riches en unités S.Ces résultats sont conformes aux études antérieures [3-6], avec une résolution spatiale cinq fois plus élevée. Nous avons enfin pu étudier avec précision la composition des parois de fibres de bois de tension hors couche G et de rayons, jusqu’alors peu étudiées.En résumé, la microspectroscopie ATR-FTIR offre de nouvelles possibilités pour l’étude de la composition de la paroi cellulaire à l’échelle de la cellule.

In insects, the gall-inducing life-style has evolved independently many times. Several evolutionary pathways leading to this lifestyle have been proposed. While there is compelling evidence supporting surface-feeders and stem-borers as ancestral states of insect gall-inducers, an evolutionary pathway from leaf-miners remains hypothetical. Here we explored this question by comparing the developmental processes of two micromoths, a gall-inducer Caloptilia cecidophora (Lep., Gracillariidae), and its non-gall-inducing relative C. ryukyuensis. Like other Caloptilia, the first and second instars of C. cecidophora are leaf-miners and the gall is initiated inside the leaf mine by the third instar, thus suggesting leaf-mining as an ancestral, plesiomorphic state in this case. This is the first example of an insect species switching from leaf-mining to gall-inducing during larval development. The first two leaf-mining instars of C. cecidophora exhibit an absence of growth and a reduced time duration compared to C. ryukyuensis. The shortening of the duration of leaf-mining stages is apparently compensated in C. cecidophora by a larger egg size than C. ryukyuensis, and an additional larval instar during the gall phase.

National audience; Trees are able to grow high and to live old thanks to the remarkable properties of their wood. As a matter of a fact, wood delivers three major functions: (1) water conduction from roots to crown, (2) support of the ever-increasing mass of the growing tree and (3) storage of temporary reserves, important for tree growth over the years. In angiosperm trees, different wood cell types are affected to each of these functions. Fibers are involved in tree mechanical support, vessels in water conduction and parenchyma rays in starch and/or lipid storage during the resting period. In addition, these cell types have distinct developmental programs. While fibers and vessels are early-dying cells, parenchyma rays stay alive longer. Therefore wood is a complex patchwork of cells and its structure results from the three-dimensional assembly of the cell walls of dead fibers and vessels, interconnected with still living parenchyma rays. This great complexity stands as an obstacle when studying wood formation and the construction of wood properties. However, this can be circumvented thanks to the development of cell-specific approaches. We developed a non-destructive method based on ATR-FTIR imaging on poplar wood sections. This technology enables to collect IR-absorbance spectra from small areas of cross-sections, which makes possible to differentiate between wood cell-types or even between the different cell wall layers from a single fiber. We first demonstrated that spectra taken from fiber cell walls on cross-sections differed from spectra obtained from wood powder. We also showed that ATR-FTIR imaging is able to discriminate the cell walls of fibers, vessels and rays. These findings are in accordance with other studies [1], but with an improved spatial resolution. ATR-FTIR microspectroscopy is thus a promising tool to finely characterize the cell wall of different wood cell types. This work has been partly supported by the OPeNSPeNU project (funded by the Centre Val de Loire Region)

REALLY INTERESTING NEW GENE (RING) proteins play important roles in the regulation of many processes by recognizing target proteins for ubiquitination. Previously, we have shown that the expression of PtaRHE1, encoding a Populus tremula × Populus alba RING-H2 protein with E3 ubiquitin ligase activity, is associated with tissues undergoing secondary growth. To further elucidate the role of PtaRHE1 in vascular tissues, we have undertaken a reverse genetic analysis in poplar. Within stem secondary vascular tissues, PtaRHE1 and its corresponding protein are expressed predominantly in the phloem. The downregulation of PtaRHE1 in poplar by artificial miRNA triggers alterations in phloem fibre patterning, characterized by an increased portion of secondary phloem fibres that have a reduced cell wall thickness and a change in lignin composition, with lower levels of syringyl units as compared with wild-type plants. Following an RNA-seq analysis, a biological network involving hormone stress signalling, as well as developmental processes, could be delineated. Several candidate genes possibly associated with the altered phloem fibre phenotype observed in amiRPtaRHE1 poplar were identified. Altogether, our data suggest a regulatory role for PtaRHE1 in secondary phloem fibre development.

article en open access; International audience; In the search for renewable energy sources, genetic engineering is a promising strategy to improve plant cell wall composition for biofuel and bioproducts generation. Lignin is a major factor determining saccharification efficiency and, therefore, is a prime target to engineer. Here, lignin content and composition were modified in poplar (Populus tremula 3 Populus alba) by specifically down-regulating CINNAMYL ALCOHOL DEHYDROGENASE1 (CAD1) by a hairpin-RNA-mediated silencing approach, which resulted in only 5% residual CAD1 transcript abundance. These transgenic lines showed no biomass penalty despite a 10% reduction in Klason lignin content and severe shifts in lignin composition. Nuclear magnetic resonance spectroscopy and thioacidolysis revealed a strong increase (up to 20-fold) in sinapaldehyde incorporation into lignin, whereas coniferaldehyde was not increased markedly. Accordingly, ultra-high-performance liquid chromatography-mass spectrometry-based phenolic profiling revealed a more than 24,000-fold accumulation of a newly identified compound made from 8-8 coupling of two sinapaldehyde radicals. However, no additional cinnamaldehyde coupling products could be detected in the CAD1-deficient poplars. Instead, the transgenic lines accumulated a range of hydroxycinnamate-derived metabolites, of which the most prominent accumulation (over 8,500-fold) was observed for a compound that was identified by purification and nuclear magnetic resonance as syringyl lactic acid hexoside. Our data suggest that, upon down-regulation of CAD1, coniferaldehyde is converted into ferulic acid and derivatives, whereas sinapaldehyde is either oxidatively coupled into S'(8-8) S' and lignin or converted to sinapic acid and derivatives. The most prominent sink of the increased flux to hydroxycinnamates is syringyl lactic acid hexoside. Furthermore, low-extent saccharification assays, under different pretreatment conditions, showed strongly increased glucose (up to +81%) and xylose (up to +153%) release, suggesting that down-regulating CAD1 is a promising strategy for improving lignocellulosic biomass for the sugar platform industry.

The lignification process in poplar tension wood lignified cell wall layers, specifically the S(1) and S(2) layers and the compound middle lamella (CML), was analysed using ultraviolet (UV) and transmission electron microscopy (TEM). Variations in the thickness of the gelatinous layer (G-layer) were also measured to clarify whether the lignified cell wall layers had completed their lignification before the deposition of G-layers, or, on the contrary, if lignification of these layers was still active during G-layer formation. Observations using UV microscopy and TEM indicated that both UV absorbance and the degree of potassium permanganate staining increased in the CML and S(1) and S(2) layers during G-layer formation, suggesting that the lignification of these lignified layers is still in progress during G-layer formation. In the context of the cell-autonomous monolignol synthesis hypothesis, our observations suggest that monolignols must go through the developing G-layer during the lignification of CML and the S(1) and S(2) layers. The alternative hypothesis of external synthesis (in the rays) does not require that monolignols go through the G-layer before being deposited in the CML, or the S(1) and S(2) layers. Interestingly, the previous observation of lignin in the poplar G-layer was not confirmed with the microscopy techniques used in the present study.

Wood formation is a complex biological process, involving five major developmental steps, including (1) cell division from a secondary meristem called the vascular cambium, (2) cell expansion (cell elongation and radial enlargement), (3) secondary cell wall deposition, (4) programmed cell death, and (5) heartwood formation. Thanks to the development of genomic studies in woody species, as well as genetic engineering, recent progress has been made in the understanding of the molecular mechanisms underlying wood formation. In this review, we will focus on two different aspects, the lignification process and the control ofmicrofibril angle in the cell wall of wood fibres, as they are both key features of wood material properties., La formation du bois est un processus de développement complexe, impliquant cinq étapes majeures : (1) la division des cellules à partir d’un méristème secondaire, appelé le cambium; (2) l’élongation des cellules ; (3) le dépôt d’une paroi secondaire épaisse ; (4) la mort programmée des cellules ; et (5) leur imprégnation par des composés phénoliques conduisant à la formation du bois de coeur. Grâce au développement des techniques de génomique et aux travaux menés sur la caractérisation d’arbres transgéniques, certains mécanismes moléculaires impliqués dans la formation du bois commencent à être mieux connus. Dans cette revue, deux aspects importants pour la qualité du bois seront approfondis : la lignification et le contrôle de l’angle des microfibrilles de cellulose dans la paroi des fibres de bois.

International audience; Six Populus deltoides Bartr. ex Marsh. × P. nigra L. genotypes were selected to investigate whether stem xylem anatomy correlated with gas exchange rates, water-use efficiency (WUE) and growth performance. Clonal copies of the genotypes were grown in a two-plot common garden test under contrasting water regimes, with one plot maintained irrigated and the other one subjected to moderate summer water deficit. The six genotypes displayed a large range of xylem anatomy, mean vessel and fibre diameter varying from about 40 to 60 μm and from 7.5 to 10.5 μm, respectively. Decreased water availability resulted in a reduced cell size and an important rise in vessel density, but the extent of xylem plasticity was both genotype and trait dependent. Vessel diameter and theoretical xylem-specific hydraulic conductivity correlated positively with stomatal conductance, carbon isotope discrimination and growth performance-related traits and negatively with intrinsic WUE, especially under water deficit conditions. Vessel diameter and vessel density measured under water deficit conditions correlated with the relative losses in biomass production in response to water deprivation; this resulted from the fact that a more plastic xylem structure was generally accompanied by a larger loss in biomass production.

Article en open access; International audience; Xylem resistance to drought-induced cavitation is a key trait of plant water relations. This study assesses the genetic variation expressed for stem cavitation resistance within a population of a riparian species, the European black poplar (Populus nigra L.), and explores its relationships with xylem anatomy, water-use efficiency (WUE), and growth. Sixteen structural and physiological traits related to cavitation resistance, xylem anatomy, growth, bud phenology, and WUE were measured on 33 P. nigra genotypes grown under optimal irrigation in a 2-year-old clonal experiment in a nursery. Significant genetic variation was expressed for the xylem tension inducing 50% loss of hydraulic conductivity (Ψ50) within the studied population, as attested by the high value of broad-sense heritability estimated for this trait (H 2 ind = 0.72). Stem cavitation resistance was associated with xylem structure: the more cavitation-resistant genotypes exhibited lower hydraulic efficiency and higher mechanical reinforcement as assessed from stem xylem cross sections. By contrast, Ψ50 was not significantly related to shoot height increment, total above-ground dry mass, or bulk leaf carbon isotope discrimination, a proxy for intrinsic WUE. These findings indicate that the trade-offs between xylem resistance to cavitation, hydraulic efficiency, and mechanical reinforcement can occur at the within-population level. Given that the studied genotypes were exposed to the same environmental conditions and evolutionary drivers in situ, the trade-offs detected at this scale are expected to reflect true functional relationships.

International audience; Wood is a complex and highly variable tissue, the formation of which is developmentally and environmentally regulated. In reaction to gravitropic stimuli, angiosperm trees differentiate tension wood, a wood with specific anatomical, chemical and mechanical features. In poplar the most significant of these features is an additional layer that forms in the secondary wall of tension wood fibres. This layer is mainly constituted of cellulose microfibrils oriented nearly parallel to the fibre axis. Tension wood formation can be induced easily and strongly by bending the stem of a tree. Located at the upper side of the bent stem, tension wood can be compared with the wood located on its lower side. Therefore tension wood represents an excellent model for studying the formation of xylem cell walls. This review summarizes results recently obtained in the field of genomics on tension wood. In addition, we present an example of how the application of functional genomics to tension wood can help decipher the molecular mechanisms responsible for cell wall characteristics such as the orientation of cellulose microfibrils.

Autorisation de diffusion par l'éditeur; Le puceron lanigère du peuplier. Phloeomyzus passerinii est un ravageur important des peupleraies du sud de l'Europe, du Proche-Orient et d'Afrique du Nord. Après une insertion de ses stylets, à la fois intercellulaire et intracellulaire, il se nourrit exclusivement dans le parenchyme cortical du tronc des peupliers. Durant les pullulations, ce puceron Cause des nécroses corticales étendues et un dessèchement des rameaux, pouvant conduire à la mort de l'arbre. Nous avons comparé la réaction corticale après blessure mécanique et après piqure du puceron entre deux génotypes hybrides Populus x canadensis avec des niveaux de résistance contrastés ('I214' qui est sensible et 'I45/51' qui affecte les paramètres démographiques du puceron). Chaque semaine durant un mois, nous avons réalisé des coupes histologiques autour des zones d'insertion des stylets et des blessures mécaniques pour comparer les modifications anatomiques et biochimiques des tissus de l'écorce. Les blessures mécanique ont induit des péridermes de blessures très localisés alors que l'alimentation du puceron a entraîné des réactions élendues, présentant des similarités avec un bourrelet cicatriciel, mais avec également des cellules hypertrophiées à parois minces, une localisation particulière des dépôts pariétaux de lignine et des accumulations périphériques de tanins et flavanols. Les dépôts de lignine étaient plus étendus et importants chez le génotype résistant. Notre hypothèse est que P. passerinzrii, aveC son mode destructif de pénétration des tissus de son hôte, déclenche les mécanismes de défense du peuplier, puis les manipule pour induire la formation d'une pseudogalle.

International audience; In hardwood trees, tension wood formation is a remarkable adaptive mechanism that makes possible for the tree to reorient its axis. Tension wood differs from normal wood by anatomical, chemical and physical characteristics: it is less lignified, and contains more cellulose of higher crystallinity, with longitudinally oriented microfibrils. In poplar, tension wood fibers harbor an extra cell wall layer, named G-layer, responsible for the peculiar mechanical properties of tension wood (1). We propose that differences between tension and normal wood mechanical properties are linked to the polysaccharide composition of the G-layer. In order to investigate the nature of these links, we realized immunolocalisation studies with light microscope observation. 158 monoclonal antibodies (from CCRC, LM, JIM and MAC series) raised against various epitopes specific to different families of plant cell wall polysaccharides were tested. For each antibody tested, we analyzed where labeling was present, not only between tension and opposite wood (that is completely devoid of G-fibres), but also according to cell type, cell wall layer, and this along the gradient of xylem cell differentiation. Most antibodies directed against xyloglucans, fucosylated or not, specifically labeled the primary wall / middle lamella in both tension and opposite wood fibers. This was also the case for several antibodies specific to rhamnogalacturonans I. Some other antibodies labeled only the S2 layer, such as most antibodies directed against xylans, whereas, unexpectedly, a few other xylan-specific antibodies exhibited a wider range of labeling. None of the antibody tested was able to specifically label opposite wood cell walls. On the contrary, among all the antibodies directed against rhamnogalacturonan I or arabinogalactan epitopes that were tested, eighteen of them specifically labeled the G-layer in tension wood fibers and, for some of them, the labeling was restricted to newly formed G-layers. Overall, from the signals obtained with each of the antibody tested, differences between tension and opposite wood fibers when observed were only detected at the level of the G-layer whose composition appears to evolve throughout its maturation. Furthermore, our study strongly suggests the involvement of specific arabinogalactan-proteins and pectins during G-layer construction. Finally, we optimized a procedure for G-layer isolation in order to analyze the polysaccharide composition of purified G-layers by mass spectrometry. In the presentation, we will discuss more in details all the results obtained using these two complementary approaches.

International audience; The course of the infection and development of the biotrophic fungus Melampsora larici-populina on leaf tissue from the hybrid poplar Populus deltoides × P. nigra ‘Ogy’ was monitored at the histological level. Leaf disks were inoculated with one of two rust physiological races (E1 and E2), resulting in interactions that were either incompatible (race E1) or compatible (race E2). In the compatible interaction, the fungus rapidly colonized the leaf without inducing any apparent host response. Symptoms appeared on the leaf several days after inoculation just prior to spore dissemination. The incompatible interaction was characterized by the early collapse and disorganization of cytoplasm of infected cells 17 h after inoculation and within 2 h after the appearance of the first haustoria. Resistance to M. larici-populina was mediated through a hypersensitive response, since it was extremely localized and involved only the few cells that were in the immediate vicinity of each infected cell.

Des plants de Populus tremula Michx. × Populus alba L. clone 717-1B4 ont été cultivés en système de culture hydroponique pour obtenir une activité cambiale dans un milieu nutritif variable. La croissance secondaire de la tige a été caractérisée par une mesure automatique de la croissance radiale et des observations histologiques. Les résultats montrent le premier enregistrement de l'activité cambiale en hydroponie chez le peuplier. En outre, nous démontrons que les carences nutritives peuvent être étudiées en privant progressivement les plants de solution nutritive. Le système se prête à la mesure de la conductance stomatique, de la croissance primaire de la tige, de la croissance foliaire et de la croissance radiale. Dans cette étude nous observons que les croissances primaire et secondaire sont touchées par les carences nutritives. Ce système de culture en hydroponie est exploitable pour l'élucidation de l'impact des facteurs environnementaux, physiologiques et moléculaires sur l'activité cambiale et la formation du bois., Populus tremula Michx. × Populus alba L. clone 717-1B4 plants were grown in a hydroponic system in an effort to detect cambial activity in a changing nutrient environment. The secondary growth of the stem was determined by automated measurement of radial growth, as well as by histological study. This is the first time poplar cambial activity has been recorded in a hydroponic system. Further, we demonstrate that nutrient limitations can be tested with progressive deprivation of liquid medium. The system lends itself to measurements of stomatal conductance, primary stem growth, leaf growth, and radial stem growth. In this study we found that primary and secondary growth were affected by nutrient solution limitations. This hydroponic system will be valuable in elucidating the impact of environmental, physiological, and molecular factors on cambial activity and wood formation.

International audience; • Fifteen poplar cDNA encoding fasciclin-like arabinogalactan proteins (PopFLAs) were finely characterized, whereas the presence of arabinogalactan proteins (AGPs) was globally assessed during wood formation. • PopFLAs transcript accumulation was analysed through EST distribution in cDNA libraries, semi-quantitative RT-PCR, microarray experiment and Northern blot analysis. Similarly, AGPs contents were globally quantified by rocket electrophoresis. AGPs accumulation was further examined by Western blotting and immunocytolocalization. • Ten PopFLAs were specifically expressed in tension wood (TW) and not expressed in the cambial zone. Rocket electrophoresis revealed important AGPs accumulation in TW xylem. An anti-AGPs specific antibody recognized two proteins preferentially present in the cell wall-bound fraction from TW. Immunocytochemistry revealed a strong labelling close to the inner part of the G-layer of TW fibres. • PopFLAs are expressed in xylem and many are up-regulated in TW. It is suggested that some PopFLAs accumulating at the inner side of the G-layer may have a specific function in the building of this layer. PopFLAs expression may therefore be linked to the specific mechanical properties of TW.

We describe in this study punchless , a nonpathogenic mutant from the rice blast fungus M. grisea, obtained by plasmid-mediated insertional mutagenesis. As do most fungal plant pathogens, M. grisea differentiates an infection structure specialized for host penetration called the appressorium. We show that punchless differentiates appressoria that fail to breach either the leaf epidermis or artificial membranes such as cellophane. Cytological analysis of punchless appressoria shows that they have a cellular structure, turgor, and glycogen content similar to those of wild type before penetration, but that they are unable to differentiate penetration pegs. The inactivated gene, PLS1 , encodes a putative integral membrane protein of 225 aa (Pls1p). A functional Pls1p-green fluorescent protein fusion protein was detected only in appressoria and was localized in plasma membranes and vacuoles. Pls1p is structurally related to the tetraspanin family. In animals, these proteins are components of membrane signaling complexes controlling cell differentiation, motility, and adhesion. We conclude that PLS1 controls an appressorial function essential for the penetration of the fungus into host leaves.

Background Due to its peculiar properties, tension wood formation constitutes a remarkable adaptation mechanism, that makes possible for the tree to reorientate its axes (stem and branches) in response to environnemental cues. In poplar, tension wood fibres harbour an extra cell wall layer, the G-layer, responsible for the peculiar mechanical properties of tension wood. This G-layer is very thick, most likely devoid of lignins and strongly enriched in highly cristalline cellulose. In addition, cellulose microfibril orientation is almost parallel to the fibre axis. We aim to identify molecular actors responsible for the tensioning of cellulose microfibrils and we choose as candidate, molecules containing complex carbohydrates, such as pectin and the glycosylated part of arabinogalactan proteins. Indeed, a wide array of different carbohydrates has been recently evidenced in the G-layer, suggesting the occurrence of complex polysaccharides other than cellulose within this layer (1, 2).

Le bois de tension (BT) est produit par les arbres feuillus en réponse à des contraintes mécaniques. Il présente des caractéristiques anatomiques, physico-chimiques et mécaniques particulières qui confèrent aux arbres un potentiel d’adaptation vis-à-vis de la contrainte subie. Traditionnellement dans la production de bois d’oeuvre, sa présence en trop grande proportion dans les grumes est une caractéristique indésirable car elle peut induire des déformations ou des fentes importantes des sciages. Néanmoins, comme le bois de tension se caractérise par une très faible teneur en lignine et une forte teneur en cellulose, il présente un intérêt pour une valorisation du bois en bioéthanol de 2ème génération. Quelle que soit la valorisation envisagée, il apparait crucial de pouvoir quantifier la teneur en bois de tension dans le bois. Cependant, les techniques actuelles, principalement basées sur de la coloration différentielle, de l’analyse histologique ou une inspection visuelle (Badia et al., 2005), sont couteuses et laborieuses, et de fait elles ne permettent pas d’évaluer cette propriété dans un nombre important d’échantillons. La spectrométrie infrarouge est une technique à haut-débit prometteuse pour évaluer les propriétés physico-chimiques du bois. Sa mise en oeuvre nécessite la construction préalable d’un modèle de calibration pour la propriété ciblée. Comme le bois de tension présente des propriétés physico-chimiques particulières, l’évaluation de sa teneur dans un échantillon de bois doit être possible par spectrométrie infrarouge. L’objectif de cette étude est de tester cette hypothèse sur des échantillons de peuplier et à partir d’une gamme de pourcentage de BT artificiellement créée.

This work is part of an ANR - Genoplante 2010 research project entitled: “Molecular bases of acclimation and adaptation to water deficit in poplar” (POPSEC, 2007-2009).; International audience; The numerous molecular studies on drought response in plants were mostly focused on short-term responses without considering dose-response effects. Moreover, despite the importance of xylem in tree water relations, assessments of drought-induced gene expression in wood are very scarce. In this work, we investigated xylem correlations between histological characteristics induced by water shortage and drought-induced gene expression. The experimental plan consisted of two Populus deltoides x Populus nigra hybrid clones (Soligo and Carpaccio) contrasted for their tolerance to water shortage and four water regimes: one short-term response to water deficit and two long-term responses either to mild and moderate water deficit and the well-watered control. For each condition tested, six trees were dedicated to molecular studies and three for ecophysiological and growth measurements. After histological staining of stem cross sections, image analysis was carried out on selected areas corresponding to stressed xylem. Total vessel lumen area was consistently higher in Carpaccio as compared to Soligo, whatever the stress condition. Interestingly, under the most stringent water shortage, Soligo exhibited a significant increase of the cell wall/ray cell fraction and an increase of water use efficiency. Spatial statistics are underway to better describe the variability of these histological variables. In addition, complete debarked stem samples and cambial zones only plus very young differentiating scraped xylem were analyzed on whole-genome Affymetrix poplar arrays. We identified modules of co-expressed genes especially using a new clustering algorithm developed by Maere et al. (BMC Systems Biology 2008, 2:33). A limited number of modules are representative of clone-specific responses but some others are potentially correlated to genotype-treatment interactions. Prospective models to integrate these phenotypic, ecophysiological and gene expression data will be discussed.

National audience