Blade Expansion Research Articles

The STF/WOX1 MD is required for physical interaction with MtWOX9 and leaf blade outgrowth in Medicago truncatula.

Plant-specific WUSCHEL-related homeobox (WOX) family transcription factors play critical roles in maintaining meristems and lateral organ development. The WUS clade member STF/LAM1 physically interacts with the intermediate clade member WOX9. This interaction contributes to their antagonistical functions on leaf blade outgrowth by competing for the same cis-elements in the promoter of their common target in M. truncatula and N. sylvestris. Here, we identified the main interaction domains of STF and MtWOX9 in Medicago, shedding light on the mechanism of WOX gene function. The middle domain of STF and MtWOX9 are both critical for the interaction, while the conserved motif of STF in the C-terminal domain is also required. Deletion of the middle domain of STF partially rescued the leaf blade phenotypes of the stf null mutant, indicating that the middle domain plays an essential role during leaf blade expansion. This finding provides a new insight that the versatility of WOX function is not only caused by the conserved DNA binding and repression domains but also by the middle domain that recruits different partners.

Light and temperature regulation of leaf morphogenesis in Arabidopsis.

Leaves are the main photosynthetic organs in plants, and their anatomy is optimized for light interception and gas exchange. Although each species has a characteristic leaf anatomy, which depends on the genotype, leaves also show a large degree of developmental plasticity. Light and temperature regulate leaf development from primordia differentiation to late stages of blade expansion. While the molecular mechanisms of light and temperature signaling have been mostly studied in seedlings, in the latest years, research has focused on leaf development. Here, I will describe the latest work carried out in the environmental regulation of Arabidopsis leaf development, comparing signaling mechanisms between leaves and seedlings, highlighting the new discoveries, and pointing out the most exciting open questions.

Direct observation of the conformational states of PIEZO1

PIEZOs are mechanosensitive ion channels that convert force into chemoelectric signals1,2 and have essential roles in diverse physiological settings3. In vitro studies have proposed that PIEZO channels transduce mechanical force through the deformation of extensive blades of transmembrane domains emanating from a central ion-conducting pore4–8. However, little is known about how these channels interact with their native environment and which molecular movements underlie activation. Here we directly observe the conformational dynamics of the blades of individual PIEZO1 molecules in a cell using nanoscopic fluorescence imaging. Compared with previous structural models of PIEZO1, we show that the blades are significantly expanded at rest by the bending stress exerted by the plasma membrane. The degree of expansion varies dramatically along the length of the blade, where decreased binding strength between subdomains can explain increased flexibility of the distal blade. Using chemical and mechanical modulators of PIEZO1, we show that blade expansion and channel activation are correlated. Our findings begin to uncover how PIEZO1 is activated in a native environment. More generally, as we reliably detect conformational shifts of single nanometres from populations of channels, we expect that this approach will serve as a framework for the structural analysis of membrane proteins through nanoscopic imaging.

Biomechanical evaluation of a novel tri-blade titanium implantable vertebral augmentation device

BACKGROUND CONTEXTTitanium implantable vertebral augmentation device (TIVAD) are regarded as having potential in the treatment of vertebral compression fractures (VCFs). However, improper design in current TIVADs results in the inability to effectively restore VCF height and maintain stability. There is still an unmet clinical need for improvement. PURPOSEThe authors tested a newly developed a TIVAD (Tri-blade fixed system) that can provide enough endplate collapse support to restore the vertebral body height in a safe retraction mechanism for VCFs using minimally invasive surgery (MIS). STUDY DESIGNThe performed biomechanical tests included blade expansion force, lifetime of cement embedded and vertebral height restoration efficiency of porcine osteoporosis VCFs for its feasibility. METHODSA cylinder with 3 surface cuts that form blades that can be expanded into a conical space was designed (Tri-blade fixed system). The 3 blades can be expanded outward with angles between blades as 105°/ 105°/150° for lower left/lower right/upper arms, respectively that reach 15mm in height and 14.8 mm in width. A frame was specifically designed to measure the contact force using force sensing resistors during blade expansion. The Tri-blade fixed system was embedded into a cement block to perform fatigue testing under 2000N pressure (5*106 cycles) for understanding the device lifetime limitation. The Tri-blade system was then inserted into porcine osteoporosis VCFs to examine the vertebral height restoration efficiency. RESULTSThe average maximum contact force for the top, bottom left and right blades were 299.0N, 283.5N and 279.3N, respectively with uniformly outward expansion forces. The fatigue test found that there were no obvious cracks or damage to the cement block. The porcine osteoporosis vertebral body at the anterior, middle, and posterior heights can be restored to 21.9%, 12.6% and 6.4%, respectively. CONCLUSIONSThis study developed a novel TIVAD with conical shape that can provide a more stable structure with sufficient/uniform expansion force, passing the fatigue test with bone cement and high effective in vertebral height restoration tests for porcine osteoporosis VCFs. CLINICAL SIGNIFICANCEThe new 3D Tri-blade TIVAD may offer a new treatment option for VCFs.

DEFECTIVE EMBRYO AND MERISTEMS1 (DEM1) Is Essential for Cell Proliferation and Cell Differentiation in Tomato.

Most flowering plant species contain at least two copies of the DEFECTIVE EMBRYO AND MERISTEMS (DEM) gene with the encoded DEM proteins lacking homology to proteins of known biochemical function. In tomato (Sl; Solanum lycopersicum), stable mutations in the SlDEM1 locus result in shoot and root meristem defects with the dem1 mutant failing to progress past the cotyledon stage of seedling development. Generation of a Somatic Mutagenesis of DEM1 (SMD) transformant line in tomato allowed for the characterization of SlDEM1 gene function past the seedling stage of vegetative development with SMD plants displaying a range of leaf development abnormalities. Further, the sectored or stable in planta expression of specific regions of the SlDEM1 coding sequence also resulted in the generation of tomato transformants that displayed a range of vegetative development defects, which when considered together with the dem1 mutant seedling and SMD transformant line phenotypic data, allowed for the assignment of SlDEM1 gene function to early embryo development, adaxial epidermis cell development, lateral leaf blade expansion, and mesophyll cell proliferation and differentiation.

WOX family transcriptional regulators modulate cytokinin homeostasis during leaf blade development in Medicago truncatula and Nicotiana sylvestris.

The plant-specific family of WUSCHEL (WUS)-related homeobox (WOX) transcription factors is key regulators of embryogenesis, meristem maintenance, and lateral organ development in flowering plants. The modern/WUS clade transcriptional repressor STENOFOLIA/LAMINA1(LAM1), and the intermediate/WOX9 clade transcriptional activator MtWOX9/NsWOX9 antagonistically regulate leaf blade expansion, but the molecular mechanism is unknown. Using transcriptome profiling and biochemical methods, we determined that NsCKX3 is the common target of LAM1 and NsWOX9 in Nicotiana sylvestris. LAM1 and NsWOX9 directly recognize and bind to the same cis-elements in the NsCKX3 promoter to repress and activate its expression, respectively, thus controlling the levels of active cytokinins in vivo. Disruption of NsCKX3 in the lam1 background yielded a phenotype similar to the knockdown of NsWOX9 in lam1, while overexpressing NsCKX3 resulted in narrower and shorter lam1 leaf blades reminiscent of NsWOX9 overexpression in the lam1 mutant. Moreover, we established that LAM1 physically interacts with NsWOX9, and this interaction is required to regulate NsCKX3 transcription. Taken together, our results indicate that repressor and activator WOX members oppositely regulate a common downstream target to function in leaf blade outgrowth, offering a novel insight into the role of local cytokinins in balancing cell proliferation and differentiation during lateral organ development.

The WOX family transcriptional regulator SlLAM1 controls compound leaf and floral organ development in Solanum lycopersicum.

Plant-specific WOX family transcription factors play important roles ranging from embryogenesis to lateral organ development. The WOX1 transcription factors, which belong to the modern clade of the WOX family, are known to regulate outgrowth of the leaf blade specifically in the mediolateral axis; however, the role of WOX1 in compound leaf development remains unknown. Phylogenetic analysis of the whole WOX family in tomato (Solanum lycopersicum) indicates that there are 10 members that represent the modern, intermediate, and ancient clades. Using phylogenetic analysis and a reverse genetic approach, in this study we identified SlLAM1 in the modern clade and examined its function and tissue-specific expression pattern. We found that knocking out SlLAM1 via CRISPR/Cas9-mediated genome editing led to narrow leaves and a reduced number of secondary leaflets. Overexpression of tomato SlLAM1 could rescue the defects of the tobacco lam1 mutant. Anatomical and transcriptomic analyses demonstrated that floral organ development, fruit size, secondary leaflet initiation, and leaf complexity were altered due to loss-of-function of SlLAM1. These findings demonstrate that tomato SlLAM1 plays an important role in the regulation of secondary leaflet initiation, in addition to its conserved function in blade expansion.

WOX9 functions antagonistic to STF and LAM1 to regulate leaf blade expansion in Medicago truncatula and Nicotiana sylvestris.

WOX family transcription factors regulate multiple developmental programs. The intermediate clade transcriptional activator WOX9 functions together with the modern clade transcriptional repressor WOX genes in embryogenesis and meristems maintenance, but the mechanism of this interaction is unclear. STF and LAM1 are WOX1 orthologs required for leaf blade outgrowth in Medicago truncatula and Nicotiana sylvestris, respectively. Using biochemical methods and genome editing technology, here we show that WOX9 is an abaxial factor and functions antagonistically to STF and LAM1 to regulate leaf blade development. While NsWOX9 ectopic expression enhances the lam1 mutant phenotype, and antisense expression partially rescues the lam1 mutant, both overexpression and knockout of NsWOX9 in N. sylvestris resulted in a range of severe leaf blade distortions, indicating important role in blade development. Our results indicate that direct repression of WOX9 by WUS clade repressor STF/LAM1 is required for correct blade architecture and patterning in M. truncatula and N. sylvestris. These findings suggest that controlling transcriptional activation and repression mechanisms by direct interaction of activator and repressor WOX genes may be required for cell proliferation and differentiation homeostasis, and could be an evolutionarily conserved mechanism for the development of complex and diverse morphology in flowering plants.

Leaflet initiation and blade expansion are separable in compound leaf development

Compound leaves are composed of multiple separate blade units termed leaflets. In tomato (Solanum lycopersicum) compound leaves, auxin promotes both leaflet initiation and blade expansion. However, it is unclear how these two developmental processes interact. With highly variable complexity, tomato compound leaves provide an ideal system to address this question. In this study, we obtained and analyzed mutants of the WUSCHEL-RELATED HOMEOBOX (WOX) family gene SlLAM1 from tomato, whose orthologs in tobacco (Nicotiana sylvestris) and other species are indispensable for blade expansion. We show that SlLAM1 is expressed in the middle and marginal domains of leaves, and is required for blade expansion in leaflets. We demonstrate that sllam1 mutants cause a delay of leaflet initiation and slightly alter the arrangement of first-order leaflets, whereas the overall leaflet number is comparable to that of wild-type leaves. Analysis of the genetic interactions between SlLAM1 and key auxin signaling components revealed an epistatic effect of SlLAM1 in determining the final leaf form. Finally, we show that SlLAM1 is also required for floral organ growth and affects the fertility of gametophytes. Our data suggest that SlLAM1 promotes blade expansion in multiple leaf types, and leaflet initiation can be largely uncoupled from blade expansion during compound leaf morphogenesis.

Research on Effect of Icing Degree on Performance of NACA4412 Airfoil Wind Turbine

In order to study the effect of icing on the wind turbine blade tip speed ratio and wind energy utilization coefficient under working conditions, it is important to better understand the growth characteristics of wind turbine blade icing under natural conditions. In this paper, the icing test of the NACA4412 airfoil wind turbine was carried out using the natural low temperature wind turbine icing test system. An evaluation model of icing degree was established, and the influence of wind speed and icing degree on the performance parameters of wind turbines was compared and analyzed. It is shown that icing is mainly concentrated on the leading edge of the blade and the windward side. The icing on the leeward surface is concentrated near the leading edge of the blade. The amount of icing varies significantly along the direction of blade expansion, with the least amount of ice accumulation at 0.2R, followed by 0.6R, and most at 0.95R. At the beginning of the experiment, the icy thickness at the 0.2R, 0.6R and 0.95R locations increased at a rate of 12.03%, 8.87% and 6.00% respectively, and then the growth rate gradually decreased and stabilized. At the same wind speed, the blade tip speed ratio and wind energy utilization coefficient are inversely related to the change trend of the icing degree. When the blade icing degree is 0.82%, 1.49% and 2.73%, the maximum tip speed ratio is reduced by 7.1%, 14.1% and 21.2% and the maximum wind energy utilization coefficient is reduced by 36.3%, 51.2% and 61.6% respectively.

Identification of Candidate Adaxial-Abaxial-Related Genes Regulating Petal Expansion During Flower Opening in Rosa chinensis "Old Blush".

Petal expansion is the main process by which flower opening occurs in roses (Rosa chinensis). Although the regulation of leaf expansion has been extensively studied, little is known about the mechanisms controlling petal expansion. The regulation of leaf dorsoventral (adaxial–abaxial) polarity is important for blade expansion and morphogenesis, but the mechanisms involved adaxial–abaxial regulation in petals are unknown. We found that auxin, a key hormonal regulator of leaf adaxial–abaxial patterning, is unevenly distributed in rose petals. The transcriptomes of the adaxial and abaxial petal tissues were sequenced at three developmental stages during flower opening. Genes that were differentially expressed between the two tissues were filtered for those known to be involved in petal expansion and phytohormone biosynthesis, transport, and signaling, revealing potential roles in petal expansion, especially auxin pathway genes. Using a weighted gene coexpression network analysis (WGCNA), we identified two gene modules that may involve in adaxial–abaxial regulation, 21 and five hub genes have been found respectively. The qRT-PCR validation results were consistent with the RNA-seq data. Based on these findings, we propose a simple network of adaxial–abaxial-related genes that regulates petal expansion in R. chinensis “Old Blush.” For the first time, we report the adaxial–abaxial transcriptional changes that occur during petal expansion, providing a reference for the study of the regulation of polarity in plant development.

Relationship between Mg, B and Mn status and tomato tolerance against Cd toxicity

Distinct tomato genotypes possess different tolerance degree to cadmium (Cd), but the mechanisms behind this phenomenon are scarcely understood. To this end, the physiological, biochemical, anatomical, nutritional and molecular mechanisms associated to the plant tolerance against Cd toxicity were investigated in five tomato accessions with contrasting sensitivity to Cd exposure. Firstly, the data revealed that larger biomass loss was not always coupled to higher Cd concentration, indicating that other events, in addition to the internal Cd accumulation, impact tomato performance at early stages of Cd exposure. Secondly, the results indicated that the fine regulation of nutrient status, particularly magnesium (Mg), boron (B) and manganese (Mn), is associated to the mitigation of Cd toxicity. Magnesium status was coupled to the modulation of root development, resulting in changes in root hair formation and biomass allocation. Boron accumulation in leaves was linked to Cd toxicity, suggesting that tolerance mechanisms involved strategies to decrease or even avoid B excess in photosynthetic tissues. Disturbances in Mn status, i.e. Mn excess in leaves and Mn deficiency in roots, were also related to tomato sensitivity to Cd exposure. Thirdly, plant capacity to maintain leaf blade expansion is a relevant strategy for a better tomato development after short-term Cd exposure. Fourthly, tomato tolerance to Cd-induced stress does not depend on CAT activity enhancements in such conditions. In conclusion, tomato ability to quickly manage its nutritional status is necessary for alleviation of the Cd effects at early stages of exposure to this metal. The better understanding about tolerance mechanisms and mode of action of Cd toxicity in plants can help in the establishment of strategies to mitigate its impacts on crops.

Seed germination at different stratification temperatures and development of Phytelephas macrocarpa Ruiz & Pavón seedlings

Abstract: Phytelephas macrocarpa (ivory palm) is an Amazonian palm vulnerable to exploitation pressure, as its seeds are widely used in regional handicrafts. The aims of this study were to evaluate the effectiveness of different stratification temperatures in overcoming seed dormancy and to analyze the seedling development stages of this species. In germination under stratification, the seeds were placed in plastic bags containing moistened vermiculite, and maintained at the constant temperatures of 25, 30, 35 and 40 °C, and an alternating temperature from 26 to 40 °C. In the study about the development of seedlings, seeds were sown in vermiculite under plastic cover (growing house), and the evolution of the seedling was evaluated, from the formation of germinative button to the complete blade expansion of the first eophyll. Stratification at alternating temperatures (26 to 40 °C) helped overcoming seed dormancy. Stratification at 25 °C kept the seed viability for nine months. Germination, characterized by the formation of the germinative button took an average of 114 ± 24 days, and the seedling development until the first extended eophyll lasted 244 ± 57 days.

The effect of cold priming on the fitness of Arabidopsis thaliana accessions under natural and controlled conditions

Priming improves an organism's performance upon a future stress. To test whether cold priming supports protection in spring and how it is affected by cold acclimation, we compared seven Arabidopsis accessions with different cold acclimation potentials in the field and in the greenhouse for growth, photosynthetic performance and reproductive fitness in March and May after a 14 day long cold-pretreatment at 4 °C. In the plants transferred to the field in May, the effect of the cold pretreatment on the seed yield correlated with the cold acclimation potential of the accessions. In the March transferred plants, the reproductive fitness was most supported by the cold pretreatment in the accessions with the weakest cold acclimation potential. The fitness effect was linked to long-term effects of the cold pretreatment on photosystem II activity stabilization and leaf blade expansion. The study demonstrated that cold priming stronger impacts on plant fitness than cold acclimation in spring in accessions with intermediate and low cold acclimation potential.

Anatomy, Histochemistry, and Antifungal Activity of Anacardium humile (Anacardiaceae) Leaf.

Leaves of Anacardium humile are used in Brazilian traditional medicine for the treatment of intestinal disturbances and skin lesions. This study aimed to define leaf diagnostic structural characters, to propose a new method of phytochemical analysis of secretions, prospect flavonoids and alkaloids, and to evaluate their inhibitory activity on Candida albicans. Common anatomical, phytochemical, and microbiological methods were used. Leaves of Anacardium occidentale were used as a reference for the structural analyses. The main structural characters were closed vascular system, absence of ducts at the medulla, bilateral mesophyll, absence of bundle sheath extension, and secretory idioblasts at the xylem. The ducts present schizogenous origin, and secreting activity is restricted to the initial phases of leaf blade expansion. The proposed new phytochemical method is practical and inexpensive and has potential for wide application. The abundance of tannins and flavonoids is related to medicinal use. A single peak in high-performance liquid chromatography indicated the presence of a pure substance not previously reported. The extract had a strong inhibitory effect on C. albicans. The obtained results confirm the potential of A. humile for the prospection of new bioactive compounds.

Involvement of nitrate reductase in the ameliorating effect of 5-aminolevulinic acid on NaCl-stressed barley seedlings

Exogenous application of 5-aminolevulinic acid (ALA) is widely used to decrease the effect of different abiotic stresses in various crops while the nitrogen metabolism is a possible target of ALA in plants. We investigated the effect of exogenous ALA on growth, activity of nitrate reductase (NR; EC 1.6.6.1), synthesis of proline and ROS generation in seedlings of the salt-sensitive barley cultivar (Hordeum vulgare L. cv. Gonar). Obtained data indicated that the ALA (20, 40 and 80 mg L−1) increased elongation and leaf blade expansion of barley seedlings exposed to 150 mM NaCl as compared to control plants, which were not treated by ALA. NR gene (Nar1) expression, NR protein content and enzymatic activities increased in ALA-treated plants. ALA also induced accumulation of free proline and decreased generation of superoxide anion radicals. Overall, this study demonstrates that ALA effects on salt-stressed barley involves changes of NR activities both at transcriptional and translational levels and leads to complex physiological modifications, such as improvement of growth and antiradical defense.

Mesophyll-Driven Blade Expansion in <i>Pisum sativum</i> var. <i>argenteum</i> Leaves

The growth rate of any multilayered plant organ is limited by the cell layer with the least extensibility. The dicot leaf blade has two epidermal layers covering the mesophyll layers, in which the vascular network is embedded. There has been a lingering uncertainty about which layer limits the rate of blade expansion in dicot leaves. The current study made use of leaf strips cut from the argenteum variety of Pisum sativum L., in which the epidermal layers can easily be removed with minimal damage. After this procedure, the mesophyll showed accelerated growth in short-term and long-term experiments and light and darkness. Extension of both layers is strongly promoted by acidic solutions. Isolated mesophyll layers expand in response to light. This effect depends on turgor pressure, photosynthesis, and the plasma membrane H+-ATPase. The data allow concluding that expanding leaf blades share with axial stem organs a similar arrangement of tissue tension: an expanding core tissue pushing against a restrictive epidermal envelope. In complete leaves, partial removal of the epidermis from only one side of the blade causes a strong epinastic or hyponastic response. Removal of matching epidermis strips from both sides of complete blades causes the exposed mesophyll strip to elongate in excess of the neighboring tissue: it buckles.

Failure of the Tomato Trans-Acting Short Interfering RNA Program to Regulate AUXIN RESPONSE FACTOR3 and ARF4 Underlies the Wiry Leaf Syndrome

Interfering with small RNA production is a common strategy of plant viruses. A unique class of small RNAs that require microRNA and short interfering (siRNA) biogenesis for their production is termed trans-acting short interfering RNAs (ta-siRNAs). Tomato (Solanum lycopersicum) wiry mutants represent a class of phenotype that mimics viral infection symptoms, including shoestring leaves that lack leaf blade expansion. Here, we show that four WIRY genes are involved in siRNA biogenesis, and in their corresponding mutants, levels of ta-siRNAs that regulate AUXIN RESPONSE FACTOR3 (ARF3) and ARF4 are reduced, while levels of their target ARFs are elevated. Reducing activity of both ARF3 and ARF4 can rescue the wiry leaf lamina, and increased activity of either can phenocopy wiry leaves. Thus, a failure to negatively regulate these ARFs underlies tomato shoestring leaves. Overexpression of these ARFs in Arabidopsis thaliana, tobacco (Nicotiana tabacum), and potato (Solanum tuberosum) failed to produce wiry leaves, suggesting that the dramatic response in tomato is exceptional. As negative regulation of orthologs of these ARFs by ta-siRNA is common to land plants, we propose that ta-siRNA levels serve as universal sensors for interference with small RNA biogenesis, and changes in their levels direct species-specific responses.

Why Wiry? Tomato Mutants Reveal Connections among Small RNAs, Auxin Response Factors, Virus Infection, and Leaf Morphology

Tomato ( Solanum lycopersicum ) wiry mutants posed a mystery because their phenotype, wiry or shoestring leaves that lack leaf blade expansion, resembles virus-infected tomato plants (see [figure][1]). However, the wiry mutant phenotype is not transmissible from plant to plant, as an infection would

Auxin-induced leaf blade expansion in Arabidopsis requires both wounding and detachment

Elevation of leaf auxin (indole-3-acetic acid; IAA) levels in intact plants has been consistently found to inhibit leaf expansion whereas excised leaf strips grow faster when treated with IAA. Here we test two hypothetical explanations for this difference in growth sensitivity to IAA by expanding leaf tissues in vivo versus in vitro. We asked if, in Arabidopsis, IAA-induced growth of excised leaf strips results from the wounding required to excise tissue and/or results from detachment from the plant and thus loss of some shoot or root derived growth controlling factors. We tested the effect of a range of exogenous IAA concentrations on the growth of intact attached, wounded attached, detached intact, detached wounded as well as excised leaf strips. After 24 h, the growth of intact attached, wounded attached, and detached intact leaves was inhibited by IAA concentrations as little as 1 µM in some experiments. Growth of detached wounded leaves and leaf strips was induced by IAA concentrations as low as 10 µM. Stress, in the form of high light, increased the growth response to IAA by leaf strips and reduced growth inhibition response by intact detached leaves. Endogenous free IAA content of intact attached leaves and excised leaf strips was found not to change over the course of 24 h. Together these results indicate growth induction of Arabidopsis leaf blade tissue by IAA requires both substantial wounding as well as detachment from the plant and suggests in vivo that IAA induces parallel pathways leading to growth inhibition.