Catherine Coutand

Xylem Wall Collapse in Water-Stressed Pine Needles

Wall reinforcement in xylem conduits is thought to prevent wall implosion by negative pressures, but direct observations of xylem geometry during water stress are still largely lacking. In this study, we have analyzed the changes in xylem geometry during water stress in needles of four pine species (Pinus spp.). Dehydrated needles were frozen with liquid nitrogen, and xylem cross sections were observed, still frozen, with a cryo-scanning electron microscope and an epifluorescent microscope. Decrease in xylem pressure during drought provoked a progressive collapse of tracheids below a specific threshold pressure (Pcollapse) that correlates with the onset of cavitation in the stems. Pcollapse was more negative for species with smaller tracheid diameter and thicker walls, suggesting a tradeoff between xylem efficiency, xylem vulnerability to collapse, and the cost of wall stiffening. Upon severe dehydration, tracheid walls were completely collapsed, but lumens still appeared filled with sap. When dehydration proceeded further, tracheids embolized and walls relaxed. Wall collapse in dehydrated needles was rapidly reversed upon rehydration. We discuss the implications of this novel hydraulic trait on the xylem function and on the understanding of pine water relations.

Posture control and skeletal mechanical acclimation in terrestrial plants : Implications for mechanical modeling of plant architecture

Self-supporting plant stems are slender, erect structures that remain standing while growing in highly variable mechanical environments. Such ability is not merely related to an adapted mechanical design in terms of material-specific stiffness and stem tapering. As many terrestrial standing animals do, plant stems regulate posture through active and coordinated control of motor systems and acclimate their skeletal growth to prevailing loads. This analogy probably results from mechanical challenges on standing organisms in an aerial environment with low buoyancy and high turbulence. But the continuous growth of plants submits them to a greater challenge. In response to these challenges, land plants implemented mixed skeletal and motor functions in the same anatomical elements. There are two types of kinematic design: (1) plants with localized active movement (arthrophytes) and (2) plants with continuously distributed active movements (contortionists). The control of these active supporting systems involves gravi- and mechanoperception, but little is known about their coordination at the whole plant level. This more active view of the control of plant growth and form has been insufficiently considered in the modeling of plant architecture. Progress in our understanding of plant posture and mechanical acclimation will require new biomechanical models of plant architectural development.

In situ FT-IR microscopic study on enzymatic treatment of poplar wood cross-sections

The feasibility of Fourier transform infrared (FT-IR) microscopy to monitor in situ the enzymatic degradation of wood was investigated. Cross-sections of poplar wood were treated with cellulase Onozuka RS within a custom-built fluidic cell. Light-optical micrographs and FT-IR spectra were acquired in situ from normal and tension wood fibers. Light-optical micrographs showed almost complete removal of the gelatinous (G) layer in tension wood. No structural and spectral changes were observed in the lignified cell walls. The accessibility of cellulose within the lignified cell wall was found to be the main limiting factor, whereas the depletion of the enzyme due to lignin adsorption could be ruled out. The fast, selective hydrolysis of the crystalline cellulose in the G-layer, even at room temperature, might be explained by the gel-like structure and the highly porous surface. Young plantation grown hardwood trees with a high proportion of G-fibers thus represent an interesting resource for bioconversion to fermentable sugars in the process to bioethanol.

Stress generation in the tension wood of poplar is based on the lateral swelling power of the G-layer

The mechanism of active stress generation in tension wood is still not fully understood. To characterize the functional interdependency between the G-layer and the secondary cell wall, nanostructural characterization and mechanical tests were performed on native tension wood tissues of poplar (Populus nigra x Populus deltoids) and on tissues in which the G-layer was removed by an enzymatic treatment. In addition to the well-known axial orientation of the cellulose fibrils in the G-layer, it was shown that the microfibril angle of the S2-layer was very large (about 36 degrees). The removal of the G-layer resulted in an axial extension and a tangential contraction of the tissues. The tensile stress-strain curves of native tension wood slices showed a jagged appearance after yield that could not be seen in the enzyme-treated samples. The behaviour of the native tissue was modelled by assuming that cells deform elastically up to a critical strain at which the G-layer slips, causing a drop in stress. The results suggest that tensile stresses in poplar are generated in the living plant by a lateral swelling of the G-layer which forces the surrounding secondary cell wall to contract in the axial direction.

The gravitropic response of poplar trunks: key roles of prestressed wood regulation and the relative kinetics of cambial growth versus wood maturation.

In tree trunks, the motor of gravitropism involves radial growth and differentiation of reaction wood (Archer, 1986). The first aim of this study was to quantify the kinematics of gravitropic response in young poplar (Populus nigra x Populus deltoides, ‘I4551’) by measuring the kinematics of curvature fields along trunks. Three phases were identified, including latency, upward curving, and an anticipative autotropic decurving, which has been overlooked in research on trees. The biological and mechanical bases of these processes were investigated by assessing the biomechanical model of Fournier et al. (1994). Its application at two different time spans of integration made it possible to test hypotheses on maturation, separating the effects of radial growth and cross section size from those of wood prestressing. A significant correlation between trunk curvature and Fourniers model integrated over the growing season was found, but only explained 32% of the total variance. Moreover, over a weeks time period, the model failed due to a clear out phasing of the kinetics of radial growth and curvature that the model does not take into account. This demonstrates a key role of the relative kinetics of radial growth and the maturation process during gravitropism. Moreover, the degree of maturation strains appears to differ in the tension woods produced during the upward curving and decurving phases. Cell wall maturation seems to be regulated to achieve control over the degree of prestressing of tension wood, providing effective control of trunk shape.

Comparison of mechanical properties of tension and opposite wood in Populus

In this paper we focused on the differences of mechanical properties of tension and normal wood of 1-year-old poplar trees, artificially tilted. Elastic and fracture properties have been measured and linked to the anatomy. Tension wood is well known because it prevents good surface finishing and leads to difficulties with sawing. We studied three main mechanical properties: young modulus, energy of cutting and longitudinal residual strain of maturation (with strain gauges) because of their importance in wood technology. Moreover, this work takes place in a larger project of study, the phenomena of axes re-orientation in trees (allowing by the production of reaction wood), where these data are required for biomechanical modelling. The results show that tension wood has a higher young modulus, needs a higher energy to be cut and exhibited a higher level of longitudinal residual strain of maturation than those of normal wood. The results suggest that these differences require deeper analysis of the wood than anatomy: measurement of microfibril orientation in the S2 layer and also the lignin composition in monomeric units.

Characterization and expression analysis under bending and other abiotic factors of PtaZFP2, a poplar gene encoding a Cys2/His2 zinc finger protein

In plants, mechanoperception and transduction of mechanical signals have been studied essentially in Arabidopsis thaliana L. and Lycopersicon esculentum L. plants, i.e., in nonwoody plants. Here, we have described the isolation of both the full-length cDNA and the regulatory region of PtaZFP2, encoding a member of Cys2/His2 zinc finger protein (ZFP) family in Populus tremula L. x Populus alba L. Time course analysis of expression demonstrated that PtaZFP2 mRNA accumulated as early as 5 min in response to a controlled stem bending and is restricted to the organ where the mechanical stimulus is applied. The real-time quantitative Reverse Transcriptase Polymerase Chain Reaction experiments showed that PtaZFP2 was also rapidly up-regulated in poplar stems in response to gravitropism suggesting that PtaZFP2 is induced by different mechanical signals. Abundance of PtaZFP2 transcripts also increased highly in response to wounding and to a weaker extent to salt treatment and cold, which is consistent with the numerous putative cis-elements found in its regulatory region. As in other species, these data suggest that Cys2/His2 ZFPs could function in poplar as key transcriptional regulators in the acclimation response to different environmental factors.

How to determine sapling buckling risk with only a few measurements

Tree buckling risk (actual height/critical buckling height) is an important biomechanical trait of plant growth strategies, and one that contributes to species coexistence. To estimate the diversity of this trait among wide samples, a method that minimizes damage to the plants is necessary. On the basis of the rarely used, complete version of Greenhills model (1881, Proceedings of the Cambridge Philosophical Society 4(2): 65-73), we precisely measured all the necessary parameters on a sample of 236 saplings of 16 species. Then, using sensitivity (variance) analysis, regressions between successive models for risk factors and species ranks and the use of these models on samples of self- and nonself-supporting saplings, we tested different degrees of simplification up to the most simple and widely used formula that assumes that the tree is a cylindrical homogeneous pole. The size factor had the greatest effect on buckling risk, followed by the form factor and the modulus of elasticity of the wood. Therefore, estimates of buckling risk must consider not only the wood properties but especially the form factor. Finally, we proposed a simple but accurate method of assessing tree buckling risk that is applicable to a wide range of samples and that requires mostly nondestructive measurements.

Acclimation kinetics of physiological and molecular responses of plants to multiple mechanical loadings

During their development, plants are subjected to repeated and fluctuating wind loads, an environmental factor predicted to increase in importance by scenarios of global climatic change. Notwithstanding the importance of wind stress on plant growth and development, little is known about plant acclimation to the bending stresses imposed by repeated winds. The time-course of acclimation of young poplars (Populus tremula L.xP. alba L.) to multiple stem bendings is studied here by following diameter growth and the expression of four genes PtaZFP2, PtaTCH2, PtaTCH4, and PtaACS6, previously described to be involved in the mechanical signalling transduction pathway. Young trees were submitted either to one transient bending per day for several days or to two bendings, 1-14 days apart. A diminution of molecular responses to subsequent bending was observed as soon as a second bending was applied. The minimum rest periods between two successive loadings necessary to recover a response similar to that observed after a single bending, were 7 days and 5 days for growth and molecular responses, respectively. Taken together, our results show a desensitization period of a few days after a single transitory bending, indicating a day-scale acclimation of sensitivity to the type of wind conditions plants experience in their specific environment. This work establishes the basic kinetics of acclimation to low bending frequency and these kinetic analyses will serve as the basis of ongoing work to investigate the molecular mechanisms involved. Future research will also concern plant acclimation to higher wind frequencies.

Proteome analysis of apical and basal regions of poplar stems under gravitropic stimulation

Gravity is a constant force guiding the direction of plant growth. In young poplar stem, reorientation of the apical region is mainly obtained by differential growth of elongating primary tissues. At the base, where elongation is achieved but where the cambium is active, reorientation is due to asymmetrical formation of reaction wood. After 45 min of gravistimulation, the stem showed no reorientation, but 1 week later, reaction wood was observed at the base of the stem. To determine the molecular mechanisms taking place at the top and base of the stem, after 45 min or 1 week of inclination, the changes induced in protein accumulation were studied by two-dimensional polyacrylamide gel electrophoresis and quantitatively analyzed using image analysis software. Around 300 protein spots were reproducibly detected and analyzed. Forty percent of these proteins showed significant changes after inclination. Mass spectrometry analysis of 135 spots led to the identification of 60 proteins involved in a wide range of activities and metabolisms. Very different patterns of protein expression were obtained according to conditions tested, highlighting the complexity of gravitropic responses. Our results suggest that primary and secondary tissues present specific mechanisms to sense reorientation and to respond to inclination. Some selected proteins are discussed.