Laser micromarking technique in studying the negative gravitropism in pea stem

A laser micromarking technique on plant epidermis was developed to study how a plant can reduce the stress in bending behavior by controlling the growth and morphogenesis. The negative gravitropism in a pea seedling (Pisum sativum L.) was discussed based on the time-dependent displacement of laser marking points which were formed by spatially-selective laser ablation of the cuticle layer that covers the outer surface of a plant. The elongation of the stem in the horizontal direction was remarkable in the first half of the gravitropism. The elongation percentages of the stem length between laser-marking points at around upper surface, middle, and bottom surface were evaluated to be 2.57, 4.87, and 7.70%, respectively. The characteristic feature of the stem bending in gravitropism is the elongation even at the upper surface region, that is, inside of the bending. This is a different feature from cantilever beams for structural materials like metals and polymers, where the compression of the upper surface and elongation of the bottom surface are caused by bending. Another laser micromarking technique was developed to improve the resolution of a dot-matrix pattern by fluorescent material transfer to a plant through a masking film with a micro-hole matrix pattern. Similar time-dependent displacement behavior was observed for a fluorescent dot-marked stem showing a feedback control loop in the mechanical optimization. These results suggested that plants solve the problem of the stress in stem bending through growth. The laser micromarking is an effective method for studying the mechanical optimization in plants.     

Biomechanical study of the effect of a controlled bending on tomato stem elongation: global mechanical analysis

An experiment was designed to apply a controlled bending to a tomato stem and simultaneously to measure its effect on stem elongation. Stem elongation was measured over 2 d until steady and equal rates were obtained for the control and the treated plants. Thereafter, the basal part of the stem was submitted to a transient controlled bending at constant displacement rate using a motorized dynamometer. After load removal, stem elongation was again measured for 2 d. The tested plants were mature (height visible internodes) and only the basal part of the stem, which had already finished elongation, was loaded (hypocotyl and the first three internodes). A few minutes after the application of bending, elongation stopped completely for 60 min. Thereafter it took 120-1000 min to recover a rate of elongation similar to the control. The growth response was exclusively due to the bending of the basal part of the stem. It was shown that the side mechanical perturbations on the roots and on the stem tissues interacting directly with the clamp were not significantly involved on the elongation response. These results give evidence for mechanical perception and plant signalling from the basal stem to the upper elongating zone. However, none of the variables characterizing the global mechanical state of the bent part of the stem (i.e. the maximal force, bending moment, inclination, mean curvature of the stem, stored mechanical energy) could quantitatively explain the variability of the growth response. A more local mechanical analysis is therefore needed to elucidate how the mechanical stimulus is perceived.     

Development and Growth Form of the Neotropical Liana <Emphasis Type="Italic">Croton nuntians</Emphasis>: The Effect of Light and Mode of Attachment on the Biomechanics of the Stem

The neotropical liana Croton nuntians (Euphorbiaceae) can occur in a variety of different growth habits. Juvenile freestanding plants are mechanically stable without support and resemble morphologically young trees or shrubs, whereas adult plants are climbers. Ontogenetic variation of bending and torsion properties of different growth phases are analyzed by measurements of flexural stiffness, structural bending modulus, torsional stiffness and structural torsional modulus. Mechanical and anatomical data show two fundamentally different patterns for juvenile freestanding and adult climbing plants. In freestanding plants, mechanical properties and the contribution of cortex, wood, and pith to the stem cross-section vary only little during ontogeny as is typical for semi-self-supporting plants. In contrast, climbing plants become significantly more flexible during ontogeny, as is characteristic for lianas. This is accompanied by a transition to the formation of a less dense wood type with large diameter vessels and an increasing contribution of flexible tissues (less dense wood and cortex) to the cross-sectional area and the axial second moment of area of the stems. Depending on the environmental conditions, freestanding plants can differ considerably in their appearance due to differences in branching system or stem taper. Therefore the influence of light quantity, measured as percentage of canopy opening, on the mechanical properties and the stem anatomy was tested. Freestanding plants grown with strong shade are significantly more stiff in bending compared with plants grown with a moderate light environment.     

Biomechanical study of the effect of a controlled bending on tomato stem elongation: local strain sensing and spatial integration of the signal

In a previous paper it has been demonstrated that tomato stems, submitted to a controlled basal bending, had a reduced terminal primary elongation, indicating mechanosensing and intra plant signalling. The 'intensity' of the growth response, as measured by the time to recover an elongation rate similar to the control, varied hugely between plants. However, no relation was found between the intensity of this response and the mechanical variables characterizing the global mechanical state of the stem. In this paper, a local analysis of mechanical state of each bent stem is performed in the context of beam theory. The spatial distributions of local variables all along the stem (curvature, bending moment, strains and stresses) are established. The validity of hypotheses underlying the mechanical analysis is demonstrated. To investigate the relationships between the mechanical stimulus and the growth response, a novel biomechanical analysis based on spatial integration of the mechanical stimulus is presented. It revealed that the mechanosensing is local and scattered through the stem and that the variability of the growth response is only explained by the integrals of the longitudinal strain field.     

Overexpression of the PtSOS2 gene improves tolerance to salt stress in transgenic poplar plants

In higher plants, the salt overly sensitive (SOS) signalling pathway plays a crucial role in maintaining ion homoeostasis and conferring salt tolerance under salinity condition. Previously, we functionally characterized the conserved SOS pathway in the woody plant Populus trichocarpa. In this study, we demonstrate that overexpression of the constitutively active form of PtSOS2 (PtSOS2TD), one of the key components of this pathway, significantly increased salt tolerance in aspen hybrid clone Shanxin Yang (Populus davidiana × Populus bolleana). Compared to the wild‐type control, transgenic plants constitutively expressing PtSOS2TD exhibited more vigorous growth and produced greater biomass in the presence of high concentrations of NaCl. The improved salt tolerance was associated with a decreased Na⁺ accumulation in the leaves of transgenic plants. Further analyses revealed that plasma membrane Na⁺/H⁺ exchange activity and Na⁺ efflux in transgenic plants were significantly higher than those in the wild‐type plants. Moreover, transgenic plants showed improved capacity in scavenging reactive oxygen species (ROS) generated by salt stress. Taken together, our results suggest that PtSOS2 could serve as an ideal target gene to genetically engineer salt‐tolerant trees.     

Role of Mechanical Loading in Growth of Sunflower (Helianthus annuus) Seedlings

Seedlings of the herbaceous annual Helianthus annuus L. weregrown under three regimens of daily bending. Bending wasadjustedduring ontogeny so that the deflexion of the stem tip dividedby the stem height was constant (elastic similarity). After6weeks, mechanically–treated plants exhibited a significantlygreater ratio of stem diameter to stem height, flexural stiffness,stemgrowth rate, and proportion of collenchymatous tissue inthe stem. Treated plants were also significantly smaller inheight,showed a greater proportion of stem tissue as cortex,and greater initial stem growth rate. No significant differenceswereobserved in the elastic modulus of the tissue composingthe stem, above–ground biomass, and stem diameter. Thedegree ofmechanical loading also had a significant effect onmost parameters. The most highly stressed plants were thicker,shorter, and elastically stiffer with more collenchyma and lesscortex in the stem. Since plants were loaded for only 60 s d^–1it is concludedthat mechanical effects early in life can haveprofound effects on the form and hence ecology of seedlings. Key words: Seedling growth, Helianthus annuus L., stem mechanical properties, thigmomorphogenesis, bending stress     

小麦耐逆基因-TaLEA2转化拟南芥的研究

研究小麦第3组LEA基因中T aLEA2对耐旱和耐盐性能的影响.将小麦第3组LEA基因T aLEA2连接在双元表达载体pB I121 C aM V 35S启动子下游,构建了能在植物中高效表达的载体pB I121-T aLEA2.通过农杆菌介导的真空渗透法,将其转入野生拟南芥中,经抗性筛选及PCR验证,获得T0代转基因植株,并用不同浓度的PEG 4000和N aC l对转基因拟南芥的耐逆性进行检测.结果表明,这些转基因植株可明显改进拟南芥在10%PEG及0.8%N aC l培养基上的生长状态.在实验条件下,转基因拟南芥的耐旱性及耐盐性均有所提高,提示T aLEA2基因在植物水分调节方面有重要作用.     

A cotton group C MAP kinase gene, GhMPK2, positively regulates salt and drought tolerance in tobacco

Mitogen-activated protein kinase (MAPK) cascades play important roles in mediating biotic and abiotic stress responses. In plants, MAPKs are classified into four major groups (A-D) according to their sequence homology and conserved phosphorylation motifs. Compared with well-studied MAPKs in groups A and B, little is known about group C. In this study, we functionally characterised a stress-responsive group C MAPK gene (GhMPK2) from cotton (Gossypium hirsutum). Northern blot analysis indicated that GhMPK2 was induced by abscisic acid (ABA) and abiotic stresses, such as NaCl, PEG, and dehydration. Subcellular localization analysis suggested that GhMPK2 may activate its specific targets in the nucleus. Constitutive overexpression of GhMPK2 in tobacco (Nicotiana tabacum) conferred reduced sensitivity to ABA during both seed germination and vegetative growth. Interestingly, transgenic plants had a decreased rate of water loss and exhibited enhanced drought and salt tolerance. Additionally, transgenic plants showed improved osmotic adjustment capacity, elevated proline accumulation and up-regulated expression of several stress-related genes, including DIN1, Osmotin and NtLEA5. β-glucuronidase (GUS) expression driven by the GhMPK2 promoter was clearly enhanced by treatment with NaCl, PEG, and ABA. These results strongly suggest that GhMPK2 positively regulates salt and drought tolerance in transgenic plants.     

L-Myo-inositol 1-phosphate synthase in the aquatic fern Azolla filiculoides.

L-Myo-inositol 1-phosphate synthase (INPS EC 5.5.1.4) catalyzes the conversion of D-glucose 6-phosphate to L-myo-inositol 1-phosphate. INPS is a key enzyme involved in the biosynthesis of phytate which is a common form of stored phosphates in higher plants. The present study monitored the increase of INPS expression in Azolla filiculoides resulting from exposure to inorganic phosphates, metals and salt stress. The expression of INPS was significantly higher in Azolla plants that were grown in rich mineral growth medium than those maintained on nutritional growth medium. The expression of INPS protein and corresponding mRNA increased in plants cultured in minimal nutritional growth medium when phosphate or Zn2+, Cd2+ and NaCl were added to the growth medium. When employing rich mineral growth medium, INPS protein content increased with the addition of Zn2+, but decreased in the presence of Cd2+ and NaCl. These results indicated that accumulation of phytate in Azolla is a result of the intensified expression of INPS protein and mRNA, and its regulation may be primarily derived by the uptake of inorganic phosphate, and Zn2+, Cd2+ or NaCl.     

Growth and molecular responses to long‐distance stimuli in poplars: bending vs flame wounding

Inter‐organ communication is essential for plants to coordinate development and acclimate to mechanical environmental fluctuations. The aim of this study was to investigate long‐distance signaling in trees. We compared on young poplars the short‐term effects of local flame wounding and of local stem bending for two distal responses: (1) stem primary growth and (2) the expression of mechanoresponsive genes in stem apices. We developed a non‐contact measurement method based on the analysis of apex images in order to measure the primary growth of poplars. The results showed a phased stem elongation with alternating nocturnal circumnutation phases and diurnal growth arrest phases in Populus tremula × alba clone INRA 717‐1B4. We applied real‐time polymerase chain reaction (RT‐PCR) amplifications in order to evaluate the PtaZFP2, PtaTCH2, PtaTCH4, PtaACS6 and PtaJAZ5 expressions. The flame wounding inhibited primary growth and triggered remote molecular responses. Flame wounding induced significant changes in stem elongation phases, coupled with inhibition of circumnutation. However, the circadian rhythm of phases remained unaltered and the treated plants were always phased with control plants during the days following the stress. For bent plants, the stimulated region of the stem showed an increased PtaJAZ5 expression, suggesting the jasmonates may be involved in local responses to bending. No significant remote responses to bending were observed.     

Modulation of polyamine balance in Lotus glaber by salinity and arbuscular mycorrhiza.

In this work we investigated the involvement of Glomus intraradices in the regulation of plant growth, polyamines and proline levels of two Lotus glaber genotypes differing in salt tolerance, after longterm exposure to saline stress. The experiment consisted of a randomized block design with three factors: (1) mycorrhizal treatments (with or without AM fungus); (2) two salinity levels of 0 and 200mM NaCl; and (3) L. glaber genotype. Experiments were performed using stem cuttings derived from L. glaber individuals representing a natural population from saline lowlands. One of the most relevant results was the higher content of total free polyamines in mycorrhized plants compared to non-AM ones. Since polyamines have been proposed as candidates for the regulation of root development under saline situations, it is possible that AM plants (which contained higher polyamine levels and showed improved root growth) were better shaped to cope with salt stress. Colonization by G. intraradices also increased (Spd+Spm)/Put ratio in L. glaber roots. Interestingly, such increment in salt stressed AM plants of the sensitive genotype, was even higher than that produced by salinization or AM symbiosis separately. On the other hand, salinity but not mycorrhizal colonization influenced proline levels in both L. glaber genotypes since high proline accumulation was observed in both genotypes under salt stress conditions. Our results suggest that modulation of polyamine pools can be one of the mechanisms used by AM fungi to improve L. glaber adaptation to saline soils. Proline accumulation in response to salt stress is a good indicator of stress perception and our results suggest that it could be used as such among L. glaber genotypes differing in salt stress tolerance.     

Effect of mechanical perturbation on the biomechanics, primary growth and secondary tissue development of inflorescence stems of Arabidopsis thaliana

Background and Aims

Mechanical perturbation is known to inhibit elongation of the inflorescence stem of Arabidopsis thaliana. The phenomenon has been reported widely for both herbaceous and woody plants, and has implications for how plants adjust their size and form to survive in mechanically perturbed environments. While this response is an important aspect of the plant''s architecture, little is known about how mechanical properties of the inflorescence stem are modified or how its primary and secondary tissues respond to mechanical perturbation.

Methods

Plants of the Columbia-0 ecotype were exposed to controlled brushing treatments and then submitted to three-point bending tests to determine stem rigidity and stiffness. Contributions of different tissues to the inflorescence stem geometry were analysed.

Key Results

Perturbed plants showed little difference in stem diameter, were 50 % shorter, 75 % less rigid and 70 % less stiff than controls. Changes in mechanical properties were linked to significant changes in tissue geometry – size and position of the pith, lignified interfascicular tissue and cortex – as well as a reduction in density of lignified cells. Stem mechanical properties were modified by changes in primary tissues and thus differ from changes observed in most woody plants tested with indeterminate growth – even though a vascular cambium is present in the inflorescence axis.

Conclusions

The study suggests that delayed development of key primary developmental features of the stem in this ecotype of Arabidopsis results in a ‘short and flexible’ rather than a ‘short and rigid’ strategy for maintaining upright axes in conditions of severe mechanical perturbation. The mechanism is comparable with more general phenomena in plants where changes in developmental rate can significantly affect the overall growth form of the plant in both ecological and evolutionary contexts.     

MS2 coat proteins bound to yeast mRNAs block 5′ to 3′ degradation and trap mRNA decay products: implications for the localization of mRNAs by MS2-MCP system

Arrays of MS2 binding sites are placed into mRNAs and are commonly used to visualize the localization of mRNAs in vivo by the expression of an MS2-GFP fusion protein. In Saccharomyces cerevisiae, we observed that arrays of MS2 binding sites inhibit 5′ to 3′ degradation of the mRNA in yeast cells and lead to the accumulation of a 3′ mRNA fragment containing the MS2 binding sites. This accumulation can be dependent on the binding of the MS2 stem loops (MS2-SL) by MS2 coat proteins (MCPs). Since such decay fragments can still bind MCP-GFP, the localization of such mRNA fragments can complicate the interpretation of the localization of full-length mRNA in vivo.     

Deciphering the developmental plasticity of walnut saplings in relation to climatic factors and light environment

Developmental plasticity, the acclimation of plants to their local environment, is known to be crucial for the fitness of perennial organisms such as trees. However, deciphering the many possible developmental and environmental influences involved in such plasticity in natural conditions requires dedicated statistical models integrating developmental phases, environmental factors, and interindividual heterogeneity. These models should be able to analyse retrospective data (number of leaves or length of annual shoots along the main stem in the present case). In this study Markov switching linear mixed models were applied to the analysis of the developmental plasticity of walnut saplings during the establishment phase in a mixed Mediterranean forest. In the Markov switching linear mixed models estimated from walnut data sets, the underlying Markov chain represents both the succession and lengths of growth phases, while the linear mixed models represent both the influence of climatic factors and interindividual heterogeneity within each growth phase. On the basis of these integrative statistical models, it is shown that walnut saplings have an opportunistic mode of development that is primarily driven by the changing light environment. In particular, light availability explains the ability of a tree to reach a phase of strong growth where the first branches can appear. It is also shown that growth fluctuation amplitudes in response to climatic factors increased while interindividual heterogeneity decreased during tree development.     

Ectopic expression of wheat expansin gene TaEXPA2 improved the salt tolerance of transgenic tobacco by regulating Na+/K+ and antioxidant competence

High salinity is one of the most serious environmental stresses that limit crop growth. Expansins are cell wall proteins that regulate plant development and abiotic stress tolerance by mediating cell wall expansion. We studied the function of a wheat expansin gene, TaEXPA2, in salt stress tolerance by overexpressing it in tobacco. Overexpression of TaEXPA2 enhanced the salt stress tolerance of transgenic tobacco plants as indicated by the presence of higher germination rates, longer root length, more lateral roots, higher survival rates and more green leaves under salt stress than in the wild type (WT). Further, when leaf disks of WT plants were incubated in cell wall protein extracts from the transgenic tobacco plants, their chlorophyll content was higher under salt stress, and this improvement from TaEXPA2 overexpression in transgenic tobacco was inhibited by TaEXPA2 protein antibody. The water status of transgenic tobacco plants was improved, perhaps by the accumulation of osmolytes such as proline and soluble sugar. TaEXPA2‐overexpressing tobacco lines exhibited lower Na⁺ but higher K⁺ accumulation than WT plants. Antioxidant competence increased in the transgenic plants because of the increased activity of antioxidant enzymes. TaEXPA2 protein abundance in wheat was induced by NaCl, and ABA signaling was involved. Gene expression regulation was involved in the enhanced salt stress tolerance of the TaEXPA2 transgenic plants. Our results suggest that TaEXPA2 overexpression confers salt stress tolerance on the transgenic plants, and this is associated with improved water status, Na⁺/K⁺ homeostasis, and antioxidant competence. ABA signaling participates in TaEXPA2‐regulated salt stress tolerance.     

Ethylene sensitivity affects changes in growth patterns, but not stem properties, in response to mechanical stress in tobacco

Plant responses to mechanical stress (e.g. wind or touch) involve a suite of physiologic and developmental changes, collectively known as thigmomorphogenesis, including reductions in height increment, Young's modulus of stems, shoot growth, and seed production, and increased stem girth and root growth. A role of the phytohormone ethylene in thigmomorphogenesis has been proposed but the extent of this involvement is not entirely clear. To address this issue, wild-type (WT) and ethylene-insensitive transgenic (Tetr) tobacco ( Nicotianum tabacum ) plants were subjected to three levels of mechanical stress: 0, 25 and 75 daily flexures. Flexed plants produced shorter, thicker stems with a lower Young's modulus than non-flexed ones, and these responses occurred independently of genotype. This suggests that ethylene does not play a role in thigmomorphogenesis-related changes in stem characteristics in tobacco. The effect of mechanical stress on dry mass increment (growth), on the other hand, differed between the genotypes: in the WT plants, shoot growth but not root growth was reduced under mechanical stress, resulting in reduced total growth and increased root mass fractions. In the Tetr plants, neither shoot nor root growth were affected. This suggests that ethylene is involved in the inhibition of tobacco shoot growth under mechanical stress.