Differences in the Occurrence of Cell Wall Components between Distinct Cell Types in Glands of Drosophyllum lusitanicum.

Carnivorous plants are mixotrophs that have developed the ability to lure, trap, and digest small organisms and utilize components of the digested bodies. Leaves of Drosophyllum lusitanicum have two kinds of glands (emergences): stalked mucilage glands and sessile digestive glands. The stalked mucilage glands perform the primary role in prey lure and trapping. Apart from their role in carnivory, they absorb water condensed from oceanic fog; thus, plants can survive in arid conditions. To better understand the function of carnivorous plant emergences, the molecular composition of their cell walls was investigated using immunocytochemical methods. In this research, Drosophyllum lusitanicum was used as a study system to determine whether cell wall immunocytochemistry differs between the mucilage and digestive glands of other carnivorous plant species. Light and electron microscopy were used to observe gland structure. Fluorescence microscopy revealed the localization of carbohydrate epitopes associated with the major cell wall polysaccharides and glycoproteins. The mucilage gland (emergence) consists of a glandular head, a connecting neck zone, and stalk. The gland head is formed by an outer and inner layer of glandular (secretory) cells and supported by a layer of endodermoid (barrier) cells. The endodermoid cells have contact with a core of spongy tracheids with spiral-shaped thickenings. Lateral tracheids are surrounded by epidermal and parenchymal neck cells. Different patterns of cell wall components were found in the various cell types of the glands. Cell walls of glandular cells generally are poor in both low and highly esterified homogalacturonans (HGs) but enriched with hemicelluloses. Cell walls of inner glandular cells are especially rich in arabinogalactan proteins (AGPs). The cell wall ingrowths in glandular cells are significantly enriched with hemicelluloses and AGPs. In the case of cell wall components, the glandular cells of Drosophyllum lusitanicum mucilage glands are similar to the glandular cells of the digestive glands of Aldrovanda vesiculosa and Dionaea muscipula.

Immunocytochemical Analysis of Bifid Trichomes in Aldrovanda vesiculosa L. Traps.

The two-armed bifids (bifid trichomes) occur on the external (abaxial) trap surface, petiole, and stem of the aquatic carnivorous plant Aldrovanda vesiculosa (Droseracee). These trichomes play the role of mucilage trichomes. This study aimed to fill the gap in the literature concerning the immunocytochemistry of the bifid trichomes and compare them with digestive trichomes. Light and electron microscopy was used to show the trichome structure. Fluorescence microscopy revealed the localization of carbohydrate epitopes associated with the major cell wall polysaccharides and glycoproteins. The stalk cells and the basal cells of the trichomes were differentiated as endodermal cells. Cell wall ingrowths occurred in all cell types of the bifid trichomes. Trichome cells differed in the composition of their cell walls. The cell walls of the head cells and stalk cells were enriched with arabinogalactan proteins (AGPs); however, they were generally poor in both low- and highly-esterified homogalacturonans (HGs). The cell walls in the trichome cells were rich in hemicelluloses: xyloglucan and galactoxyloglucan. The cell wall ingrowths in the basal cells were significantly enriched with hemicelluloses. The presence of endodermal cells and transfer cells supports the idea that bifid trichomes actively transport solutes, which are polysaccharide in nature. The presence of AGPs (which are considered plant signaling molecules) in the cell walls in these trichome cells indicates the active and important role of these trichomes in plant function. Future research should focus on the question of how the molecular architecture of trap cell walls changes in cells during trap development and prey capture and digestion in A. vesiculosa and other carnivorous plants.

Immunocytochemical Analysis of the Wall Ingrowths in the Digestive Gland Transfer Cells in Aldrovanda vesiculosa L. (Droseraceae).

Carnivorous plants are unique due to their ability to attract small animals or protozoa, retain them in specialized traps, digest them, and absorb nutrients from the dissolved prey material; however, to this end, these plants need a special secretion-digestive system (glands). A common trait of the digestive glands of carnivorous plants is the presence of transfer cells. Using the aquatic carnivorous species Aldrovanda vesiculosa, we showed carnivorous plants as a model for studies of wall ingrowths/transfer cells. We addressed the following questions: Is the cell wall ingrowth composition the same between carnivorous plant glands and other plant system models? Is there a difference in the cell wall ingrowth composition between various types of gland cells (glandular versus endodermoid cells)? Fluorescence microscopy and immunogold electron microscopy were employed to localize carbohydrate epitopes associated with major cell wall polysaccharides and glycoproteins. The cell wall ingrowths were enriched with arabinogalactan proteins (AGPs) localized with the JIM8, JIM13, and JIM14 epitopes. Both methylesterified and de-esterified homogalacturonans (HGs) were absent or weakly present in the wall ingrowths in transfer cells (stalk cells and head cells of the gland). Both the cell walls and the cell wall ingrowths in the transfer cells were rich in hemicelluloses: xyloglucan (LM15) and galactoxyloglucan (LM25). There were differences in the composition between the cell wall ingrowths and the primary cell walls in A. vesiculosa secretory gland cells in the case of the absence or inaccessibility of pectins (JIM5, LM19, JIM7, LM5, LM6 epitopes); thus, the wall ingrowths are specific cell wall microdomains. Even in the same organ (gland), transfer cells may differ in the composition of the cell wall ingrowths (glandular versus endodermoid cells). We found both similarities and differences in the composition of the cell wall ingrowths between the A. vesiculosa transfer cells and transfer cells of other plant species.

Asymmetric wall ingrowth deposition in Arabidopsis phloem parenchyma transfer cells is tightly associated with sieve elements.

In Arabidopsis, polarized deposition of wall ingrowths in phloem parenchyma (PP) transfer cells (TCs) occurs adjacent to cells of the sieve element/companion cell (SE/CC) complex. However, the spatial relationships between these different cell types in minor veins, where phloem loading occurs, are poorly understood. PP TC development and wall ingrowth localization were compared with those of other phloem cells in leaves of Col-0 and the transgenic lines AtSUC2::AtSTP9-GFP (green fluorescent protein) and AtSWEET11::AtSWEET11-GFP that identify CCs and PP cells, respectively. The development of PP TCs in minor veins, indicated by deposition of wall ingrowths, proceeded basipetally in leaves. However, not all PP cells develop wall ingrowths, and higher levels of deposition occur in abaxial- compared with adaxial-positioned PP TCs. Furthermore, the deposition of wall ingrowths was exclusively initiated on and preferentially covered the PP TC/SE interface, rather than the PP TC/CC interface, and only occurred in PP cells that were adjacent to SEs. Collectively, these results demonstrate a tight association between SEs and wall ingrowth deposition in PP TCs and suggest the existence of two subtypes of PP cells in leaf minor veins. Compared with PP cells, PP TCs showed more abundant accumulation of AtSWEET11-GFP, indicating functional differences in phloem loading between PP and PP TCs.

Arabinogalactan Proteins in the Digestive Glands of Dionaea muscipula J.Ellis Traps.

The arabinogalactan proteins (AGP) play important roles in plant growth and developmental processes. However, to the best of our knowledge, there is no information on the spatial distribution of AGP in the plant organs and tissues of carnivorous plants during their carnivorous cycle. The Dionaea muscipula trap forms an "external stomach" and is equipped with an effective digestive-absorbing system. Because its digestive glands are composed of specialized cells, the hypothesis that their cell walls are also very specialized in terms of their composition (AGP) compared to the cell wall of the trap epidermal and parenchyma cells was tested. Another aim of this study was to determine whether there is a spatio-temporal distribution of the AGP in the digestive glands during the secretory cycle of D. muscipula. Antibodies that act against AGPs, including JIM8, JIM13 and JIM14, were used. The localization of the examined compounds was determined using immunohistochemistry techniques and immunogold labeling. In both the un-fed and fed traps, there was an accumulation of AGP in the cell walls of the gland secretory cells. The epitope, which is recognized by JIM14, was a useful marker of the digestive glands. The secretory cells of the D. muscipula digestive glands are transfer cells and an accumulation of specific AGP was at the site where the cell wall labyrinth occurred. Immunogold labeling confirmed an occurrence of AGP in the cell wall ingrowths. There were differences in the AGP occurrence (labeled with JIM8 and JIM13) in the cell walls of the gland secretory cells between the unfed and fed traps.

Stellate Trichomes in Dionaea muscipula Ellis (Venus Flytrap) Traps, Structure and Functions.

The digestive organs of carnivorous plants have external (abaxial) glands and trichomes, which perform various functions. Dionaea muscipula Ellis (the Venus flytrap) is a model carnivorous plant species whose traps are covered by external trichomes. The aim of the study was to fill in the gap regarding the structure of the stellate outer trichomes and their immunocytochemistry and to determine whether these data support the suggestions of other authors about the roles of these trichomes. Light and electron microscopy was used to show the trichomes' structure. Fluorescence microscopy was used to locate the carbohydrate epitopes that are associated with the major cell wall polysaccharides and glycoproteins. The endodermal cells and internal head cells of the trichomes were differentiated as transfer cells, and this supports the idea that stellate trichomes transport solutes and are not only tomentose-like trichomes. Trichome cells differ in the composition of their cell walls, e.g., the cell walls of the internal head cells are enriched with arabinogalactan proteins (AGPs). The cell walls of the outer head cells are poor in both low and highly homogalacturonans (HGs), but the immature trichomes are rich in the pectic polysaccharide (1-4)-ß-D-galactan. In the immature traps, young stellate trichomes produce mucilage which may protect the trap surface, and in particular, the trap entrance. However, the role of these trichomes is different when the outer head cells collapse. In the internal head cells, a thick secondary wall cell was deposited, which together with the thick cell walls of the outer head cells played the role of a large apoplastic space. This may suggest that mature stellate trichomes might function as hydathodes, but this should be experimentally proven.

Molecular mechanisms of maize endosperm transfer cell development.

Endosperm transfer cells function as the nutrient transporter, antimicrobic barrier, and signal mediator between filial and maternal tissues. Sugar supply of maternal tissues, sugar demand of filial tissues, and requirement for defence against pathogens are three elemental factors inducing differentiation of endosperm transfer cells. Epigenetic factors, especially MEG1, moderate the key genetic factor ZmMRP-1 to activate endosperm transfer cell-specific genes that control the flange wall ingrowth formation and defensin-like protein secretion in maize. Auxin and cytokinin are primary hormones involved in development of maize endosperm transfer cells. Crosstalk between glucose and hormone signaling regulates endosperm transfer cell development via modifying ZmMRP-1 expression. This review summarizes the current knowledge on maize endosperm transfer cell development, and discusses its potential molecular mechanisms. It is expected to strengthen the theoretical basis for structural and functional optimization of endosperm transfer cells, and yield improvement of kernels in maize.

Cell wall polymers in the Phaeoceros placenta reflect developmental and functional differences across generations.

The placenta of hornworts is unique among bryophytes in the restriction of transfer cells that are characterized by elaborate wall labyrinths to the gametophyte generation. During development, cells around the periphery of the sporophyte foot elongate, forming smooth-walled haustorial cells that interdigitate with gametophyte cells. Using immunogold labeling with 22 antibodies to diverse cell wall polymers, we examined compositional differences in the developmentally and morphologically distinct cell walls of gametophyte transfer cells and sporophyte haustorial cells in the placenta of Phaeoceros. As detected by Calcofluor White fluorescence, cellulose forms the cell wall scaffolding in cells on both sides of the placenta. Homogalacturonan (HG) and rhamnogalacturonan I (RG-I) pectins are abundant in both cell types, and haustrorial cells are further enriched in methyl-esterified HGs. The abundance of pectins in placental cell walls is consistent with the postulated roles of these polymers in cell wall porosity and in maintaining an acidic apoplastic pH favorable to solute transport. Xyloglucan hemicellulose, but not mannans or glucuronoxylans, are present in cell walls at the interface between the two generations with a lower density in gametophytic wall ingrowths. Arabinogalactan proteins (AGPs) are diverse along the plasmalemma of placental cells and are absent in surrounding cells in both generations. AGPs in placental cell walls may play a role in calcium binding and release associated with signal transduction as has been speculated for these glycoproteins in other plants. Callose is restricted to thin areas in cell walls of gametophyte transfer cells. In contrast to studies of transfer cells in other systems, no reaction to the JIM12 antibody against extensin was observed in Phaeoceros.

Super-Resolution Fluorescence Imaging of Arabidopsis thaliana Transfer Cell Wall Ingrowths using Pseudo-Schiff Labelling Adapted for the Use of Different Dyes.

To understand plant growth and development, it is often necessary to investigate the organization of plant cells and plant cell walls. Plant cell walls are often fluorescently labeled for confocal imaging with the dye propidium iodide using a pseudo-Schiff reaction. This reaction binds free amine groups on dye molecules to aldehyde groups on cellulose that result from oxidation with periodic acid. We tested a range of fluorescent dyes carrying free amine groups for their ability to act as pseudo-Schiff reagents. Using the low-pH solution historically used for the Schiff reaction, these alternative dyes failed to label cell walls of Arabidopsis cotyledon vascular tissue as strongly as propidium iodide but replacing the acidic solution with water greatly improved fluorescence labeling. Under these conditions, rhodamine-123 provided improved staining of plant cell walls compared to propidium iodide. We also developed protocols for pseudo-Schiff labeling with ATTO 647N-amine, a dye compatible for super-resolution Stimulated Emission Depletion (STED) imaging. ATTO 647N-amine was used for super-resolution imaging of cell wall ingrowths that occur in phloem parenchyma transfer cells of Arabidopsis, structures whose small size is only slightly larger than the resolution limit of conventional confocal microscopy. Application of surface-rendering software demonstrated the increase in plasma membrane surface area as a consequence of wall ingrowth deposition and suggests that STED-based approaches will be useful for more detailed morphological analysis of wall ingrowth formation. These improvements in pseudo-Schiff labeling for conventional confocal microscopy and STED imaging will be broadly applicable for high-resolution imaging of plant cell walls.

Sucrose regulates wall ingrowth deposition in phloem parenchyma transfer cells in Arabidopsis via affecting phloem loading activity.

In Arabidopsis thaliana, phloem parenchyma transfer cells (PPTCs) occur in leaf minor veins and play a pivotal role in phloem loading. Wall ingrowth formation in PPTCs is induced by the phloem loading activity of these cells, which is regulated by sucrose (Suc). The effects of endogenous versus exogenous Suc on wall ingrowth deposition, however, differ. Elevating endogenous Suc levels by increased light enhanced wall ingrowth formation, whereas lowering endogenous Suc levels by dark treatment or genetically in ch-1 resulted in lower levels of deposition. In contrast, exogenously applied Suc, or Suc derived from other organs, repressed wall ingrowth deposition. Analysis of pAtSUC2::GFP plants, used as a marker for phloem loading status, suggested that wall ingrowth formation is correlated with phloem loading activity. Gene expression analysis revealed that exogenous Suc down-regulated expression of AtSWEET11 and 12, whereas endogenous Suc up-regulated AtSWEET11 expression. Analysis of a TREHALOSE 6-PHOSPHATE (T6P) SYNTHASE overexpression line and the hexokinase (HXK)-null mutant, gin2-1, suggested that Suc signalling of wall ingrowth formation is independent of T6P and HXK. Collectively, these results are consistent with the conclusion that Suc regulates wall ingrowth formation via affecting Suc exporting activity in PPTCs.

Transfer cells: what regulates the development of their intricate wall labyrinths?

Transfer cells (TCs) support high nutrient rates into, or at symplasmic discontinuities within, the plant body. Their transport capacity is conferred by an amplified plasma membrane surface area, enriched in nutrient transporters, supported on an intricately invaginated wall labyrinth (WL). Thus, development of the WL is at the heart of TC function. Enquiry has shifted from describing WL architecture and formation to discovering mechanisms regulating WL assembly. Experimental systems used to examine these phenomena are critiqued. Considerable progress has been made in identifying master regulators that commit stem cells to a TC fate (e.g. the maize Myeloblastosis (MYB)-related R1-type transcription factor) and signals that induce differentiated cells to undergo trans-differentiation to a TC phenotype (e.g. sugar, auxin and ethylene). In addition, signals that provide positional information for assembly of the WL include apoplasmic hydrogen peroxide and cytosolic Ca2+ plumes. The former switches on, and specifies the intracellular site for WL construction, while the latter creates subdomains to direct assembly of WL invaginations. Less is known about macromolecule species and their spatial organization essential for WL assembly. Emerging evidence points to a dependency on methyl-esterified homogalacturonan accumulation, unique patterns of cellulose and callose deposition and spatial positioning of arabinogalactan proteins.

Female gametophyte development in Sedum sediforme (Jacq.) Pau (Crassulaceae): an anatomical, cytochemical and ultrastructural analysis.

Available documentation about the development of the female gametophyte of Crassulaceae is very limited. The aim of this study was to extend the embryological knowledge of Crassulaceae by analysing the development of the embryo sac in Sedum sediforme. Transmission electron microscopy and light microscopy including Nomarski optics (DIC) were used to observe individual stages of female gametophyte development. Cytochemical staining enabled detection of lipids, insoluble polysaccharides and proteins in gametophyte cells during their formation. Their increased accumulation was observed during nucellar cell and unfunctional cell degeneration in the embryo sac at the coenocytic and cellular stages (megagametogenesis). The female gametophyte develops in anatropous, bitegmic and crassinucellate ovules. The mature embryo sac is built of seven cells but after antipodes degeneration it is formed by the egg apparatus and a central cell. The monosporic Polygonum type was observed. One megaspore mother cell (MMC) formed three cells after meiosis. A triad was formed from a functional megaspore (placed chalazally), one uninucleate megaspore and a binucleate cell located at the micropylar end. Plasmodesmata with adhering electron-dense dome were noticed in walls of the coenocytic embryo sac and in the outer walls of ephemeral antipodes. Moreover, similar to synergids, antipodes form wall ingrowths. Here, we report new structural features of the antipodal cells (the presence of plasmodesmata with an electron-dense dome) which have not been described before. This new structural observation indicates that these cells participate in substance transport and that this process can probably be additionally regulated.

Transcript Profiling Identifies NAC-Domain Genes Involved in Regulating Wall Ingrowth Deposition in Phloem Parenchyma Transfer Cells of Arabidopsis thaliana.

Transfer cells (TCs) play important roles in facilitating enhanced rates of nutrient transport at key apoplasmic/symplasmic junctions along the nutrient acquisition and transport pathways in plants. TCs achieve this capacity by developing elaborate wall ingrowth networks which serve to increase plasma membrane surface area thus increasing the cell's surface area-to-volume ratio to achieve increased flux of nutrients across the plasma membrane. Phloem parenchyma (PP) cells of Arabidopsis leaf veins trans-differentiate to become PP TCs which likely function in a two-step phloem loading mechanism by facilitating unloading of photoassimilates into the apoplasm for subsequent energy-dependent uptake into the sieve element/companion cell (SE/CC) complex. We are using PP TCs in Arabidopsis as a genetic model to identify transcription factors involved in coordinating deposition of the wall ingrowth network. Confocal imaging of pseudo-Schiff propidium iodide-stained tissue revealed different profiles of temporal development of wall ingrowth deposition across maturing cotyledons and juvenile leaves, and a basipetal gradient of deposition across mature adult leaves. RNA-Seq analysis was undertaken to identify differentially expressed genes common to these three different profiles of wall ingrowth deposition. This analysis identified 68 transcription factors up-regulated two-fold or more in at least two of the three experimental comparisons, with six of these transcription factors belonging to Clade III of the NAC-domain family. Phenotypic analysis of these NAC genes using insertional mutants revealed significant reductions in levels of wall ingrowth deposition, particularly in a double mutant of NAC056 and NAC018, as well as compromised sucrose-dependent root growth, indicating impaired capacity for phloem loading. Collectively, these results support the proposition that Clade III members of the NAC-domain family in Arabidopsis play important roles in regulating wall ingrowth deposition in PP TCs.

Nectar and oleiferous trichomes as floral attractants in Bulbophyllum saltatorium Lindl. (Orchidaceae).

Although many Orchidaceae have deceit flowers that produce no reward, the most common reward, when present, is nectar. Bulbophyllum, however, is unusual in that the labellar secretions of most species investigated to date lack sugars, and, therefore, cannot be considered true nectar. The African species Bulbophyllum saltatorium is an exception in that it produces not only nectar but also possesses specialized, capitate oleiferous trichomes. The nectary of B. saltatorium is borne on the labellum and is represented by a deep, narrow, median longitudinal groove, having a small aperture, and flanked by trichomes. Isodiametric epidermal cells lining this groove secrete nectar which collects both in the groove and on the surface of the labellum. As well as a nectary, the labellum of B. saltatorium also bears three types of unicellular trichomes: the longest trichomes are borne distally and abaxially; the marginal ones form a rim around the entire labellum, and finally, massive, capitate trichomes occur proximally and adaxially. These are oleiferous, containing large quantities of oil which might function as precursors of volatile components of fragrance or provide a food-reward. To the best of our knowledge, this is the first time for such oleiferous trichomes to be described for Bulbophyllum. Therefore, apart from their color and markings, flowers of this species are able to attract pollinators in at least two, possibly three ways: food-reward in the form of nectar; fragrance; and possibly food-rewards in the form of food-hairs.

A Structurally Specialized Uniform Wall Layer is Essential for Constructing Wall Ingrowth Papillae in Transfer Cells.

Transfer cells are characterized by wall labyrinths with either a flange or reticulate architecture. A literature survey established that reticulate wall ingrowth papillae ubiquitously arise from a modified component of their wall labyrinth, termed the uniform wall layer; a structure absent from flange transfer cells. This finding sparked an investigation of the deposition characteristics and role of the uniform wall layer using a Vicia faba cotyledon culture system. On transfer of cotyledons to culture, their adaxial epidermal cells spontaneously trans-differentiate to a reticulate architecture comparable to their abaxial epidermal transfer cell counterparts formed in planta. Uniform wall layer construction commenced once adaxial epidermal cell expansion had ceased to overlay the original outer periclinal wall on its inner surface. In contrast to the dense ring-like lattice of cellulose microfibrils in the original primary wall, the uniform wall layer was characterized by a sparsely dispersed array of linear cellulose microfibrils. A re-modeled cortical microtubule array exerted no influence on uniform wall layer formation or on its cellulose microfibril organization. Surprisingly, formation of the uniform wall layer was not dependent upon depositing a cellulose scaffold. In contrast, uniform wall cellulose microfibrils were essential precursors for constructing wall ingrowth papillae. On converging to form wall ingrowth papillae, the cellulose microfibril diameters increased 3-fold. This event correlated with up-regulated differential, and transfer-cell specific, expression of VfCesA3B while transcript levels of other cellulose biosynthetic-related genes linked with primary wall construction were substantially down-regulated.

Wall ingrowth deposition in phloem parenchyma transfer cells in Arabidopsis: Heteroblastic variations and a potential role in pathogen defence.

Transfer cell (TCs) develop unique wall ingrowth networks which amplify plasma membrane surface area and thus maximize nutrient transporter density at key anatomic sites for nutrient exchange within plants and their external environment. These sites fall into 4 main groups corresponding to 4 categories of trans-membrane flux: absorption/secretion of solutes from or to the external environment, and absorption/secretion of solutes from or to internal, extra-cytoplasmic compartments. Research on TC biology over recent decades has demonstrated correlations between wall ingrowth deposition in TCs and enhanced transport capacity in many major agricultural species such as pea, fava bean, cotton and maize. Consequently, there is general consensus that the existence of wall ingrowth morphology implies an augmentation in membrane transport capacity. However, this may not be entirely applicable for phloem parenchyma (PP) TCs in Arabidopsis. Our recent survey of PP TC abundance and distribution in Arabidopsis veins indicated that PP TC development reflects heteroblastic status. A consequence of this observation is the suggestion that PP TCs, or at least wall ingrowth deposition in these cells, potentially act as a physical barrier to defend access of invading pathogens to sugar-rich sieve elements rather than solely in facilitating the export of photoassimilate from collection phloem in leaves.

The influence of salinity on growth, morphology, leaf ultrastructure, and cell viability of the seagrass Halodule wrightii Ascherson.

Halodule wrightii is an ecologically important seagrass; however, little is known about the adaptation of this species in the context of environmental change, particularly changes arising from alterations in salinity of coastal ecosystems. This study aimed to determine the effects of different salinities on growth, morphology, leaf ultrastructure, and cell viability of H. wrightii. To accomplish this, plants were cultivated for 21 days in salinities of 25, 35, and 45. More hydropotens were observed in samples exposed to salinity of 45 with increased invagination of the plasma membrane and cell wall. These invaginations were also observed in other epidermal cells of the leaf blade. In particular, a significant retraction of plasma membrane was seen in samples exposed to salinity of 45, with possible deposition of compounds between the membrane and cell wall. Osmotic stress in samples exposed to salinity of 45 affected the chloroplasts through an increase in plastoglobules and thylakoids by granum in the epidermal chloroplasts of the leaf and decrease in the number of chloroplasts. Overall, this study showed that H. wrightii can survive within salinities that range between 25 and 45 without changing growth rate. However, the plant did have higher cell viability at salinity of 35. Salt stress in mesocosms, at both salinity of 25 and 45, decreased cell viability in this species. H . wrightii had greater changes in salinity of 45; this showed that the species is more tolerant of salinities below this value.

De novo assembly of a genome-wide transcriptome map of Vicia faba (L.) for transfer cell research.

Vicia faba (L.) is an important cool-season grain legume species used widely in agriculture but also in plant physiology research, particularly as an experimental model to study transfer cell (TC) development. TCs are specialized nutrient transport cells in plants, characterized by invaginated wall ingrowths with amplified plasma membrane surface area enriched with transporter proteins that facilitate nutrient transfer. Many TCs are formed by trans-differentiation from differentiated cells at apoplasmic/symplasmic boundaries in nutrient transport. Adaxial epidermal cells of isolated cotyledons can be induced to form functional TCs, thus providing a valuable experimental system to investigate genetic regulation of TC trans-differentiation. The genome of V. faba is exceedingly large (ca. 13 Gb), however, and limited genomic information is available for this species. To provide a resource for future transcript profiling of epidermal TC differentiation, we have undertaken de novo assembly of a genome-wide transcriptome map for V. faba. Illumina paired-end sequencing of total RNA pooled from different tissues and different stages, including isolated cotyledons induced to form epidermal TCs, generated 69.5 M reads, of which 65.8 M were used for assembly following trimming and quality control. Assembly using a De-Bruijn graph-based approach generated 21,297 contigs, of which 80.6% were successfully annotated against GO terms. The assembly was validated against known V. faba cDNAs held in GenBank, including transcripts previously identified as being specifically expressed in epidermal cells across TC trans-differentiation. This genome-wide transcriptome map therefore provides a valuable tool for future transcript profiling of epidermal TC trans-differentiation, and also enriches the genetic resources available for this important legume crop species.