Xylem Cell Development Research Articles

Transcriptomic Remodeling Occurs During Cambium Activation and Xylem Cell Development in Taxodium ascendens

Taxodium ascendens has been extensively cultivated in the wetlands of the Yangtze River in south China and has significantly contributed to ecology and timber production. Until now, research on T. ascendens genomics has yet to be conducted due to its large and complex genome, which hinders the development of T. ascendens genomic resources. Combined with the microstructural changes during cambium cell differentiation across various growth periods, we investigate the transcriptome expression and regulatory mechanisms governing cambium activity in T. ascendens. Using RNA sequencing (RNA-Seq) technology, we identified the genes involved in the cambium development of cells at three stages (dormancy, reactivation, and activity). These genes encode the regulatory and control factors associated with the cambial activity, cell division, cell expansion, and biosynthesis of cell wall components. Blast comparison revealed that three genes (TR_DN69961_c0_g1, TRINITY_DN17100_c1_g1, TRINITY_DN111727_c0_g1) from the MYB and NAC families might regulate transcription during lignin formation in wood thickening. These results illustrate the dynamic changes in the transcriptional network during vascular cambium development. Additionally, they shed light on the genetic regulation mechanism of secondary growth in T. ascendens and guide further elucidation of the candidate genes involved in regulating cambium differentiation and wood formation.

Single-cell transcriptomics unveils xylem cell development and evolution

BackgroundXylem, the most abundant tissue on Earth, is responsible for lateral growth in plants. Typical xylem has a radial system composed of ray parenchyma cells and an axial system of fusiform cells. In most angiosperms, fusiform cells comprise vessel elements for water transportation and libriform fibers for mechanical support, while both functions are performed by tracheids in other vascular plants such as gymnosperms. Little is known about the developmental programs and evolutionary relationships of these xylem cell types.ResultsThrough both single-cell and laser capture microdissection transcriptomic profiling, we determine the developmental lineages of ray and fusiform cells in stem-differentiating xylem across four divergent woody angiosperms. Based on cross-species analyses of single-cell clusters and overlapping trajectories, we reveal highly conserved ray, yet variable fusiform, lineages across angiosperms. Core eudicots Populus trichocarpa and Eucalyptus grandis share nearly identical fusiform lineages, whereas the more basal angiosperm Liriodendron chinense has a fusiform lineage distinct from that in core eudicots. The tracheids in the basal eudicot Trochodendron aralioides, an evolutionarily reversed trait, exhibit strong transcriptomic similarity to vessel elements rather than libriform fibers.ConclusionsThis evo-devo framework provides a comprehensive understanding of the formation of xylem cell lineages across multiple plant species spanning over a hundred million years of evolutionary history.

Comparison of Stomatal Structure and Distribution between Ovules and Leaves in Ginkgo biloba

Stomata are plant epidermal structures that play essential roles in photosynthesis, respiration, and transpiration. Although stomata on plant leaves have been extensively studied, their structure and distribution on other organs remain poorly understood. The “living fossil “, Ginkgo biloba, has naked ovules that are thought to be primitive reproductive structures in ancient seed plants. Therefore, we hypothesized that there are some distinct stoma features in G. biloba ovules that have not been reported. In this study, we investigated the morphological development of stomata on ovules and leaves of Ginkgo biloba using scanning electron microscopy, then examined the anatomical characteristics of the general stalk and petiole using semi-thin sectioning. We found that stomata were distributed on the epidermis of the whole ovule, except near the micropyle; these stomata persisted until harvest, indicating that ovules perform gross photosynthesis to an extent similar to the photosynthesis observed in leaves, which is beneficial to ovule development. Ovule and leaf stomata share similar orientation, composition, and development; however, their distribution and subsidiary cell morphology significantly differ. The morphology of the general stalk was similar to the morphology of the petiole, but xylem cell development was minimal, and no sclerenchyma cells were present beneath the epidermis; these findings suggested that the general stalk is biomechanically weaker than the petiole. Overall, these results suggest that despite their differences, G. biloba ovules and leaves share many morphological and anatomical similarities in terms of stomatal architecture and stalk anatomy. These findings will help to elucidate the leaf origins of “flowers” in ancient plants.

Long-term single-cell imaging and simulations of microtubules reveal principles behind wall patterning during proto-xylem development

Plants are the tallest organisms on Earth; a feature sustained by solute-transporting xylem vessels in the plant vasculature. The xylem vessels are supported by strong cell walls that are assembled in intricate patterns. Cortical microtubules direct wall deposition and need to rapidly re-organize during xylem cell development. Here, we establish long-term live-cell imaging of single Arabidopsis cells undergoing proto-xylem trans-differentiation, resulting in spiral wall patterns, to understand microtubule re-organization. We find that the re-organization requires local microtubule de-stabilization in band-interspersing gaps. Using microtubule simulations, we recapitulate the process in silico and predict that spatio-temporal control of microtubule nucleation is critical for pattern formation, which we confirm in vivo. By combining simulations and live-cell imaging we further explain how the xylem wall-deficient and microtubule-severing KATANIN contributes to microtubule and wall patterning. Hence, by combining quantitative microscopy and modelling we devise a framework to understand how microtubule re-organization supports wall patterning.

The Cytoskeleton and Its Role in Determining Cellulose Microfibril Angle in Secondary Cell Walls of Woody Tree Species.

Recent advances in our understanding of the molecular control of secondary cell wall (SCW) formation have shed light on molecular mechanisms that underpin domestication traits related to wood formation. One such trait is the cellulose microfibril angle (MFA), an important wood quality determinant that varies along tree developmental phases and in response to gravitational stimulus. The cytoskeleton, mainly composed of microtubules and actin filaments, collectively contribute to plant growth and development by participating in several cellular processes, including cellulose deposition. Studies in Arabidopsis have significantly aided our understanding of the roles of microtubules in xylem cell development during which correct SCW deposition and patterning are essential to provide structural support and allow for water transport. In contrast, studies relating to SCW formation in xylary elements performed in woody trees remain elusive. In combination, the data reviewed here suggest that the cytoskeleton plays important roles in determining the exact sites of cellulose deposition, overall SCW patterning and more specifically, the alignment and orientation of cellulose microfibrils. By relating the reviewed evidence to the process of wood formation, we present a model of microtubule participation in determining MFA in woody trees forming reaction wood (RW).

PCR-based detection of single sequence variants from a natural collection of the non-model tree species European Aspen Populus tremula (L.)

Abstract In the present study we present and discuss the identification of species-specific SNPs to rule out any experimental influence of species-specific primer design (Populus tremula vs. the closely related model-species Populus trichocarpa) on the detectability of SNPs. Applying a species-optimized method, partial sequences of 14 genes involved in xylem cell development, xylogenesis, pectin formation, and drought stress reaction were analyzed at the genomic level. About 3 Mb of sequence information were generated by Sanger sequencing technology and 258 sequence variants were identified. 15 out of these represent insertions /deletions located exclusively in non-coding regions and the remaining 243 are SNPs found in coding and non-coding regions of candidate genes. The introduction of a species-specific SNP detection pipeline will help to detect nucleotide variants in P. tremula and to conduct association mapping in natural P. tremula populations.

Cambial activity, annual rhythm of xylem production in relation to phenology and climatic factors and lignification pattern during xylogenesis in drum-stick tree (Moringa oleifera)

The interrelationship among seasonality of cambium, wood formation, cell size variation, lignification, tree phenology and climatic factors has been examined in Moringa oleifera, a tropical evergreen tree. The vascular cambium in Moringa is a storied with a distinct seasonal variation in its structure due to dimensional changes in rays. Though cambium remains active throughout the year it is sensitive to water availability. Peak cambial cell division and rate of xylem differentiation are influenced by average rainfall during the monsoon period. Cambial cell division reaches higher up in the tree trunk when it is supporting a high number of branches and leaves. Statistical analysis of cell size variation and climate factors revealed that xylem cell development is greatly influenced by rainfall and rarely by temperature. Lengths of fusiform initials and vessel elements are positively correlated. The pattern of lignification during xylogenesis shows that the vessels are the first element to develop lignified walls and ray cells are the last elements to become lignified. Fiber cell walls show more syringyl lignin, while the cell walls of other xylem elements are characterized by relatively more guaiacyl lignin units.

Radial stem growth in response to microclimate and soil moisture in a drought-prone mixed coniferous forest at an inner Alpine site.

Dendroclimatological studies in a dry inner Alpine environment (750 m a.s.l.) revealed different growth response of co-occurring coniferous species to climate, which is assumed to be caused by a temporal shift in wood formation among species. The main focus of this study therefore was to monitor intra-annual dynamics of radial increment growth of mature deciduous and evergreen coniferous species (Pinus sylvestris, Larix decidua and Picea abies) during two consecutive years with contrasting climatic conditions. Radial stem growth was continuously followed by band dendrometers and modelled using Gompertz functions to determine time of maximum growth. Histological analyses of tree ring formation allowed determination of temporal dynamics of cambial activity and xylem cell development. Daily fluctuations in stem radius and radial stem increments were extracted from dendrometer traces, and correlations with environmental variables were performed. While a shift in temporal dynamics of radial growth onset and cessation was detected among co-occurring species, intra-annual radial growth peaked synchronously in late May 2011 and early June 2012. Moist atmospheric conditions, i.e. high relative air humidity, low vapour pressure deficit and low air temperature during the main growing period, favoured radial stem increment of all species. Soil water content and soil temperature were not significantly related to radial growth. Although a temporal shift in onset and cessation of wood formation was detected among species, synchronous culmination of radial growth indicates homogenous exogenous and/or endogenous control. The close coupling of radial growth to atmospheric conditions points to the importance of stem water status for intra-annual growth of drought-prone conifers.

Concomitant analysis of cambial abscisic acid and cambial growth activity in poplar

Endogenous levels of cambial region abscisic acid (ABA) were quantified by immunoassay and assessed together with cambial growth activity in poplar (Populus nigra L. × P. maximowiczii Henry, clone Kamabuchi) over the course of a growing season. The level of cambial region ABA increased from spring to late-summer but decreased sharply in autumn. Cambial growth activity, measured as the radial number of undifferentiated cambial cells and enlarging xylem cells, also increased from spring to summer and decreased sharply in autumn, indicating the onset of cambial dormancy. Exogenous ABA, applied laterally to poplar stems at two times within the growing season, enhanced cambial growth activity, as the radial number of undifferentiated cambial cells increased in ABA-treated trees subsequent to the two application times. Xylem cell development was also affected by exogenous ABA as fibre length increased significantly in ABA-treated trees at both application times. The positive correlation of cambial region ABA and cambial growth activity as well as the positive effects of exogenous ABA application thereon sheds new light on the role of this hormonal growth regulator.

Allelopathic effects of aqueous extracts of Artistolochia esperanzae O.Kuntze on development of Sesamum indicum L. seedlings

Aristolochia esperanzae is a climbing plant that occurs in the savanna regions of Brazil. The aim of this work was to identify the effects of aqueous extracts of A. esperanzae on germination, root growth and xylem cell development of sesame seedlings. Leaf and shoot extracts were prepared at concentrations of 1.5 and 3%. Extracts caused marked changes in germination and seedling growth with greatest inhibition produced by root extracts. Morphological changes and decreased growth and development of seedlings were also observed. The extracts of A. esperanzae caused a reduction of 50% in the size of root xylem cells and marked changes in the primary root and in the number of secondary roots.

Effects of climate variables on intra-annual stem radial increment in Pinus cembra (L.) along the alpine treeline ecotone

Within the alpine treeline ecotone tree growth is increasingly restricted by extreme climate conditions. Although intra-annual stem growth recorded by dendrometers can be linked to climate, stem diameter increments in slow-growing subalpine trees are masked by changes in tree water status.We tested the hypothesis that intra-annual radial stem growth in Pinus cembra is influenced by different climate variables along the treeline ecotone in the Austrian Alps. Dendrometer traces were compared with dynamics of xylem cell development to date onset of cambial activity and radial stem growth in spring.Daily fluctuations in stem radius reflected changes in tree water status throughout the treeline ecotone. Extracted daily radial increments were significantly correlated with air temperature at the timberline and treeline only, where budburst, cambial activity and enlargement of first tracheids also occurred quite similarly. A close relationship was detected between radial increment and number of enlarging tracheids throughout the treeline ecotone.We conclude that (i) the relationship between climate and radial stem growth within the treeline ecotone is dependent on a close coupling to atmospheric climate conditions and (ii) initiation of cambial activity and radial growth in spring can be distinguished from stem re-hydration by histological analysis.

Influence of overexpression of a gibberellin 20-oxidase gene on the kinetics of xylem cell development in hybrid poplar (Populus tremula L. and P. tremuloides Michx.)

AbstractGibberellins (GAs) are important regulators of shoot growth in trees. We studied the kinetics of xylem formation in hybrid poplar (Populus tremulaL.×P. tremuloidesMichx.) in which the key regulatory gene gibberellin acid 20 oxidase (GA20-oxidase) isolated fromArabidopsisis overexpressed. Increments in the height and radius of shoots were registered by high-resolution laser measurements. The anatomical and chemical structure of mature xylem cells was studied by light electron microscopy and UV spectrophotometry. Transgenic plants showed an increase in height growth, but a lower speed of cell elongation during primary growth compared to wild-type plants. During the first year of growth, transgenic plants showed a higher radius increment, an increase in the period of cell expansion of vessels and fibres and their final size, and a higher lignin content of the compound middle lamella between fibres compared to wild-type plants. In contrast, during the third year of growth, only a slight increase in the period of cell expansion of fibre cells was observed in transgenic compared to wild-type plants. Analyses of GA20-oxidase expression in leaves and shoots of 6-month- and 3-year-old plants of three different independent transgenic lines revealed a decrease in its expression only in shoots but not in leaves of the 3-year-old plants. The results indicate that overexpression of the GA20-oxidase gene in young shoots of transgenic poplar predominately affects cell expansion, while no GA20-oxidase expression was observed in shoots of 3-year-old transgenic plants, resulting in wild-type xylem cell development.

Dodeca-CLE Peptides as Suppressors of Plant Stem Cell Differentiation

In plants and animals, small peptide ligands that signal in cell-cell communication have been suggested to be a crucial component of development. A bioassay of single-cell transdifferentation demonstrates that a dodecapeptide with two hydroxyproline residues is the functional product of genes from the CLE family, which includes CLAVATA3 in Arabidopsis. The dodecapeptide suppresses xylem cell development at a concentration of 10(-11) M and promotes cell division. An application, corresponding to all 26 Arabidopsis CLE protein family members, of synthetic dodecapeptides reveals two counteracting signaling pathways involved in stem cell fate.

Trephor: A New Tool for Sampling Microcores from tree stems

Detailed analyses of cambium activity and wood formation during growth need repeated sampling of newly formed xylem. In order to be minimally invasive, wood samples are extracted as microcores. Despite the research done on xylem cell development and the increasing interest in intra-annual studies of xylogenesis, few tools are available for microcore sampling. Methods originally designed for other purposes have often been used, but no details are available on their efficiency and usefulness. Information is also lacking on laboratory preparation techniques for cell analysis of tree-ring formation, leading to difficulties in carrying out these experiments. The advantages and limits of the tools used up to now are described. A new tool, named Trephor (patent pending n° PD2004A000324), specifically designed for long-lasting use is presented. Trephor is chisel-shaped for a fast recovery of 2 mm diameter microcores. The cutting tube is inserted into the wood using a hammer and no other accessory is required. Simple technical characteristics allow high quality samples to be collected from both softwood and hardwood species with minimum damage to the sampled trees. Trephor was tested during the 2004 growing season, demonstrating good resistance to wear and tear and mechanical stress. Embedding the microcores in paraffin for fast section preparation is described.

On the Origin of Intercellular Canals in the Secondary Xylem of Selected Meliaceae Species

The anatomy, frequency, and origin of intercellular canals in the xylem of ten Meliaceae species (Carapa guianensis Aubl., Carapa procera DC., Cedrela odorata L., Cedrela fissilis Vell., Entandrophragma cilindricum Sprague, Entandrophragma utile Sprague, Khaya ivorensis A. Chev., Khaya senegalensis (Desr.) A. Juss., Swietenia macrophylla King, Swietenia mahagoni (L.) Jacq.) were investigated using 327 samples from institutional wood collections, 398 plantation grown trees, and 43 pot cultivated plants. Tangential bands of intercellular canals and single canals were found in the xylem of all ten species. Staining of microtome sections indicated that the chemical composition of the secretion is similar to that of “wound-gums”. Studying the origin of the intercellular canals along the stem axis, it became obvious that the formation of the canals can be induced by wounding of the primary meristems (in particular by insect attacks of Hypsipyla spp., wounding of root tips) and by wounding of the cambium (formation of 43–100% of the intercellular canals). In fast growing trees of Carapa spp., Entandrophragma utile, and Khaya ivorensis, planted at an experimental site near Manaus, Brazil, numerous canals were found which were not induced by wounding of the meristems. In these trees an out of phase sequence of xylem cell development and high growth stresses were observed, which are hypothesised to be a further trigger for the traumatic formation of intercellular canals.

Application of the Gompertz equation for the study of xylem cell development

Summary To assess daily relationships between xylem development and the environment, precise knowledge of tree-ring development is required. This includes accurate estimations of the rate and duration of the cell enlargement and wall thickening phases. This paper presents an application where the Gompertz equation is used to calculate cell number increase and to estimate both the rates and periods of the differentiation phases on a daily scale during the growing season. Tests were performed on two coniferous species, Abies balsamea (L.) Mill. and Pinus cembra L., growing in the Canadian boreal forest and on the Italian Alpine treeline respectively. Wood micro-cores, including the most recent tree-rings, were collected during the growing season of 2001. Cross-sections were cut with a microtome and stained with cresyl fast violet to differentiate xylem cells of the growing tree-ring. The number of cells within the zones of radial cell enlargement, secondary wall thickening and mature xylem were counted. The normalised cell number increases were fitted into a Gompertz function [y=f(t)]. The results showed that the function provided suitable descriptions of the cell increase for both Abies balsamea and Pinus cembra with R2 ranging between 0.60 and 0.90. Subsequently, to assess the development phases in time, the Gompertz equation was expressed in function of the independent variable [t=f(y)]. The use of only one equation for the estimation of both cell division and differentiation throughout the vegetative season has demonstrated to be an important improvement.

Subcellular Quantitative Determination of K and Ca in Phloem, Cambium, and Xylem Cells of Spruce(Picea abies[L.] Karst.) During Earlywood and Latewood Formation

The present investigation was directed to calibrated subcellular quantitative determination of K and Ca in phloem, cambium, and xylem cells of spruce (Picea abies [L.] Karst.) by energy dispersive X-ray spectroscopy (EDXS) in combination with transmission electron microscopy (TEM). The element content was studied during earlywood and latewood formation throughout pnmary and secondary wall formation and lignification. In the experimental stage, increment cores were extracted from three 100-year-old forest trees and either shock-frozen, embedded, and used for semithin cuttings, or prepared for optical emission spectroscopy (ICP-OES) for bulk analysis from tissue sections. The structural dynamics in cell formation of phloem, cambium, and xylem was expressed in terms of the radial cell diameter and the cell wall area. The lignification of the same cells was studied by subcellular UV-spectroscopy. During earlywood formation, differentiating tracheids are a strong physiological sink for K. Particularly during cell enlargement, a strong increase of the symplasmatic K content (> I ng per cell) was detected. An increase of the Ca content of differentiating cells (0.4-1 ng per cell) was found during secondary wall formation, whereas the Ca content of the cell wall decreased remarkably during its lignification (0.1-0.3 ng per cell). From these experiments it can be concluded that K is a driving force in cell enlargement of differentiating tracheids (primary wall phase), whereas Ca is of importance especially for wall synthesis in the secondary phase of xylem cell development and lignification.

Extracellular laccases and peroxidases from sycamore maple (Acer pseudoplatanus) cell-suspension cultures

We have investigated the abilities of extracellular enzymes from dark-grown cell-suspension cultures of sycamore maple (Acer pseudoplatanus L.) to oxidize monolignols, the precursors for lignin biosynthesis in plants, as well as a variety of other lignin-related compounds. Laccase and peroxidase both exist as a multiplicity of isoenzymes in filtrates of spent culture medium, but their abilities to produce water-insoluble, dehydrogenation polymers (DHPs) from the monolignols (in the presence of hydrogen peroxide for the peroxidase reaction) appear identical whether or not the enzymes are purified from the concentrated filtrates or left in a crude mixture. The patterns of bonds formed in these DHPs are identical to those found in DHPs synthesized using horseradish peroxidase or fungal laccase, and many of these bonds are found in the natural lignins extracted from different plant sources. On the other hand, sycamore maple laccase is very much less active on phenolic substrates containing multiple aromatic rings than is sycamore maple peroxidase. We suggst that whereas laccase may function during the early stages of lignification to polymerize monolignols into oligo-lignols, cell-wall peroxidases may function when H2O2 is produced during the later stages of xylem cell development or in response to environmental stresses.

Cytodifferentiation of xylem cells in vitro: an overview.

It is hoped that this paper has summarized the background of xylem cell differentiation studies and has set the scene for the papers to follow in this issue. We have also indicated that many areas of the subject are still contentious and in need of further study. New techniques developed in the area of cell biology should prove useful tools in further study of this most interesting area of cytodifferentiation. It is hoped that further advancement of our knowledge in the study of xylem cell development will help understanding in the field of cell differentiation in general.