Hydrophobic protein from soybean (HPS) is present in soybean dust and is an allergen (Gly m 1) that causes asthma in allergic individuals. Past studies have shown that HPS occurs on the seed surface. To determine the microscopic localization of HPS during seed development, monoclonal antibodies to HPS were used to visualize the protein by fluorescence and transmission electron microscopy. Seed coat and endocarp sections were also examined for pectin, cellulose, callose, starch, and protein by histochemical staining. HPS is present in the endocarp epidermal cells at 18 to 28 days post anthesis. At later stages of seed development, HPS occurs in extracellular secretions that accumulate unevenly on the endocarp epidermis and seed surface. HPS is synthesized by the endocarp epidermis and deposited on the seed surface as part of a heterogeneous matrix.

Suberin is a complex biopolymer composed of a poly(aliphatic) domain (SPAD) and a poly(phenolic) domain (SPPD). Suberin is typically confined to specialized cell types including root exodermal cells, but its synthesis in a maturing exodermis is still not well understood. For the current work, Allium cepa roots were used as a model to analyze SPAD and SPPD synthesis in a maturing uniseriate exodermis. Roots were divided into four maturation zones based on their growth rate and the deposition of suberin lamellae in maturing exodermal cells. Exodermal and epidermal cell layers were separated from the underlying layers in each maturation zone, then soluble and insoluble suberin monomers were extracted chemically, and quantified and identified by gas chromatography – mass spectrometry. Temporal patterns for the synthesis of the SPAD, but not for the SPPD, were revealed upon resolution of the metabolite profiles. The composition of the soluble fraction was essentially unchanged as the exodermis matured. In contrast, the SPAD composition differed during maturation, mainly owing to significant increases in the deposition of C18:1 α,ω-dioic acid and ω-OH fatty acids. It is proposed that the exodermal maturation zones with their corresponding metabolite profiles be used as targets for the functional enzymatic characterization of suberin biosynthetic pathways.

Background and Aims Casparian bands are characteristic of the endodermis and exodermis of roots, but also occur infrequently in other plant organs, for example stems and leaves. To date, these structures have not been detected in phellem cells of a periderm. The aim of this study was to determine whether Casparian bands occur in phellem cells using tests that are known to detect Casparian bands in cells that also contain suberin lamellae. Both natural periderm and wound-induced structures were examined in shoots and roots.Methods Using Pelargonium hortorum as a candidate species, the following tests were conducted: (1) staining with berberine and counterstaining with aniline blue, (2) mounting sections in concentrated sulphuric acid and (3) investigating the permeability of the walls with berberine as an apoplastic, fluorescent tracer.Key Results (1) Berberine-aniline blue staining revealed a modification in the radial and transverse walls of mature phellem cells in both stems and roots. Three days after wounding through to the cortex of stems, the boundary zone cells (pre-existing, living cells nearest the wound) had developed vividly stained primary walls. By 17 d, staining of mature phellem cells of wound-induced periderm was similar to that of natural periderm. (2) Mature native phellem cells of stems resisted acid digestion. (3) Berberine was excluded from the anticlinal (radial and transverse) walls of mature phellem cells in stems and roots, and from the wound-induced boundary zone.Conclusions Casparian bands are present in mature phellem cells in both stems and roots of P. hortorum. It is proposed that Casparian bands act to retard water loss and pathogen entry through the primary cell walls of the phellem cells, thus contributing to the main functions of the periderm.

The endodermis and exodermis are the inner- and outermost cortical layers, respectively, of a root. Both are characterized by the development of Casparian bands in their anticlinal walls. Endodermal Casparian bands normally appear within 10 mm of the root tip, while exodermal Casparian bands are typically deposited farther from the tip. All Casparian bands contain the biopolymers lignin and suberin, allowing the endodermis and exodermis to serve as filtration sites for the passive movement of ions between the soil solution and the stele. Later in development, suberin lamellae are frequently deposited as secondary walls, which will reduce the transmembrane transport of ions and water. In some species, tertiary walls are also formed; their main function is postulated to be mechanical support of the root. Recent research with fluorescence and electron microscopy has revealed some important details of development and structure of these wall modifications. Further, chemical analyses of enzymatically isolated wall modifications have shown the chemical basis for the endodermis and exodermis as apoplastic barriers. Studies of Arabidopsis at the molecular level are shedding light on the genetic control of endodermal morphogenesis. In contrast, molecular aspects of exodermal development are totally unknown. Future work will benefit from a combined molecular and biochemical approach to the endodermis and exodermis.

The exodermis of Iris germanica roots is multiseriate. Its outermost layer matures first with typical Casparian bands and suberin lamellae. But as subsequent layers mature, the Casparian band extends into the tangential and anticlinal walls of their cells. Compared with roots in which the endodermis represents the major transport barrier, the multiseriate exodermis (MEX) was expected to reduce markedly radial water and solute transport. To test this idea, precocious maturation of the exodermis was induced with a humid air gap inside a hydroponic chamber. Hydraulic conductivity (Lppc) was measured on completely submerged roots (with an immature exodermis) and on air-gap-exposed root regions (with two mature exodermal layers) using a pressure chamber. Compared with regions of roots with no mature exodermal layers, the mature MEX reduced Lppc from 8.5×10−8 to 3.9×10−8 m s−1 MPa−1. Puncturing the MEX increased Lppc to 19×10−8 m s−1 MPa−1, indicating that this layer constituted a substantial hydraulic resistance within the root (75% of the total). Alternatively, a root pressure probe was used to produce pressure transients from which hydraulic conductivity was determined, but this device measured mainly flow through the endodermis in these wide-diameter roots. The permeability of roots to NaCl and ethanol was also reduced in the presence of two mature MEX layers. The data are discussed in terms of the validity of current root models and in terms of a potential role for I. germanica MEX during conditions of drought and salt stress.

AteIF5A3, one of three genes encoding eukaryotic translation initiation factor 5A (eIF5A) in Arabidopsis thaliana, and corresponding genes PdeIF5A3 from Populus deltoides (eastern cottonwood) and SleIF5A4 from Solanum lycopersicum (tomato) were constitutively over-expressed in A. thaliana. The resultant transgenic plants exhibited enhanced vegetative and reproductive growth. Indeed, the increase in seed yield relative to empty vector controls for the PdeIF5A3 over-expressing plants ranged from 50% to 300% depending on the line. The PdeIF5A3 over-expressing plants also exhibited enhanced fitness when exposed to osmotic and nutrient (N, P and K) stress. The spatial localization of AteIF5A3 was visualized by confocal microscopy using transgenic plants expressing P(AteIF5A3) :GFP-AteIF5A3. GFP fluorescence reflecting expression of AteIF5A3 was detectable in the phloem, particularly companion cells, of roots, stems and leaves, in the epidermal cells of the root tip, in the columella cells of the root cap and in the chalazal tissue of fertilized ovules, which all play a pivotal role in nutrient or hormone translocation. Thus, AteIF5A3 appears to be involved in supporting growth and to play a regulatory role in the response of plants to sub-lethal osmotic and nutrient stress.

Iris germanica roots develop a multiseriate exodermis (MEX) in which all mature cells contain suberin lamellae. The location and lipophilic nature of the lamellae contribute to their function in restricting radial water and solute transport. The objective of the current work was to identify and quantify aliphatic suberin monomers, both soluble and insoluble, at specific stages of MEX development and under differing growth conditions, to better understand aliphatic suberin biosynthesis. Roots were grown submerged in hydroponic culture, wherein the maturation of up to three exodermal layers occurred over 21 days. In contrast, when roots were exposed to a humid air gap, MEX maturation was accelerated, occurring within 14 days. The soluble suberin fraction included fatty acids, alkanes, fatty alcohols, and ferulic acid, while the suberin poly(aliphatic) domain (SPAD) included fatty acids, α,ω-dioic acids, ω-OH fatty acids, and ferulic acid. In submerged roots, SPAD deposition increased with each layer, although the composition remained relatively constant, while the composition of soluble components shifted toward increasing alkanes in the innermost layers. Air gap exposure resulted in two significant shifts in suberin composition: nearly double the amount of SPAD monomers across all layers, and almost three times the alkane accumulation in the first layer. The localized and abundant deposition of C18:1 α,ω-dioic and ω-OH fatty acids, along with high accumulation of intercalated alkanes in the first mature exodermal layer of air gap-exposed roots indicate its importance for water retention under drought compared with underlying layers and with entire layers developing under water.

Whether a seed coat of a soybean (Glycine max L. Mer.) seed is permeable or non-permeable is governed by a number of quantitative trait loci further influenced by environmental factors. In soybean seeds, water loss is controlled by a thin, inconspicuous outer cuticle. When intact, the outer cuticle constitutes a barrier to water passage; however, the presence of minute cracks in the cuticle results in the ready passage of water. We explored the timing of cuticular development in soybean seeds by measuring the deposition of the cutin in relation to seed growth and cell viability. Cutin deposition occurred early in the development and ceased just prior to the final stage of rapid seed expansion. Cracks in the cuticle appeared after cutin synthesis ceased while the seed continued to grow. In permeable seeds (regardless of genotype) the resistance of the cuticle to water passage increased steadily during development until seed expansion was maximal and cracks appeared in the cuticle. Once cracks formed, they became the primary site of water passage and the cuticle lost its ability to control the process. In non-permeable seeds, no cracks appeared at this critical point and the cuticle continued to restrict water passage. Microarray analysis of gene expression during seed coat development revealed a complex transcriptome with many genes uniquely expressed in the seed coat. However, the expression patterns were remarkably similar between permeable and non-permeable types, in keeping with the complexity of the underlying genetics of seed coat permeability.

BACKGROUND AND AIMS:Most studies of exodermal structure and function have involved species with a uniseriate exodermis. To extend this work, the development and apoplastic permeability of Iris germanica roots with a multiseriate exodermis (MEX) were investigated. The effects of different growth conditions on MEX maturation were also tested. In addition, the exodermises of eight Iris species were observed to determine if their mature anatomy correlated with habitat.METHODS:Plants were grown in soil, hydroponics (with and without a humid air gap) or aeroponics. Roots were sectioned and stained with various dyes to detect MEX development from the root apical meristem, Casparian bands, suberin lamellae and tertiary wall thickenings. Apoplastic permeability was tested using dye (berberine) and ionic (ferric) tracers.KEY RESULTS:The root apical meristem was open and MEX development non-uniform. In soil-grown roots, the exodermis started maturing (i.e. Casparian bands and suberin lamellae were deposited) 10 mm from the tip, and two layers had matured by 70 mm. In both hydro- and aeroponically grown roots, exodermal maturation was delayed. However, in areas of roots exposed to an air gap in the hydroponic system, MEX maturation was accelerated. In contrast, maturation of the endodermis was not influenced by the growth conditions. The mature MEX had an atypical Casparian band that was continuous around the root circumference. The MEX prevented the influx and efflux of berberine, but had variable resistance to ferric ions due to their toxic effects. Iris species living in well-drained soils developed a MEX, but species in water-saturated substrates had a uniseriate exodermis and aerenchyma.CONCLUSIONS:MEX maturation was influenced by the roots' growth medium. The MEX matures very close to the root tip in soil, but much further from the tip in hydro- and aeroponic culture. The air gap accelerated maturation of the second exodermal layer. In Iris, the type of exodermis was correlated with natural habitat suggesting that a MEX may be advantageous for drought tolerance.

Phytophthora sojae is the causal agent of root and stem rot of soybean (Glycine max). Various cultivars with partial resistance to the pathogen have been developed to mitigate this damage. Herein, two contrasting genotypes, the cultivar Conrad (with strong partial resistance) and the line OX760-6 (with weak partial resistance), were compared regarding their amounts of preformed and induced suberin components, and to early events during the P. sojae infection process. To colonize the root, hyphae grew through the suberized middle lamellae between epidermal cells. This took 2 to 3 h longer in Conrad than in OX760-6, giving Conrad plants more time to establish their chemical defenses. Subsequent growth of hyphae through the endodermis was also delayed in Conrad. This cultivar had more preformed aliphatic suberin than the line OX760-6 and was induced to form more aliphatic suberin several days prior to that of OX760-6. However, the induced suberin was formed subsequent to the initial infection process. Eventually, the amount of induced suberin (measured 8 days postinoculation) was the same in both genotypes. Preformed root epidermal suberin provides a target for selection and development of new soybean cultivars with higher levels of expression of partial resistance to P. sojae.

The seed coat is a composite structure that serves as a conduit for nutrient acquisition for the developing embryo: Upon seed desiccation and maturation, the seed coat provides cover and protection for the embryo and may impose dormancy or trigger germination by controlling water uptake. For crop plants such as soybean, traits determined by the seed coat impact the overall quality of the seed and the utility of its derived products. There are remarkable examples of unusual genetic mechanisms that control seed coat pigmentation, lustre, and cracking that have been recently described in soybean. Studies on seed permeability to water have resulted in the formulation of new hypotheses to account for variation of this trait. This review summarizes the current advances in our understanding of the structural properties of the soybean seed coat with an emphasis on the control of permeability to water in mature seeds.

In many plant species, suberin lamellae are deposited in the cells of the endodermis. Some cells near the xylem poles tend to remain without these lamellae and are known as “passage cells”. To develop a three-dimensional picture of the pattern of suberin lamella deposition, procedures were developed to isolate large pieces of onion endodermis and to stain their suberin lamellae for various types of microscopy. Sudan red 7B proved satisfactory with white-light, Fluorol yellow and berberine with epifluorescence, and Nile red with confocal laser scanning microscopy (CLSM). Lamella deposition commenced in a few cells 10 mm from the root tip and then proceeded in a scattered manner. However, lamella deposition in more cells (seen 125–155 mm from the root tip) resulted in the formation of longitudinal files of cells with lamellae alternating with files of passage cells. In older regions (255–285 mm from the tip), almost all cells had deposited suberin lamellae, leaving only a few passage cells. At higher magnification, three-dimensional reconstructions of CLSM images of endodermal suberin lamellae revealed that they are perforated by pores all along the roots. Such pores could serve as points for water and ions to enter the cytoplasm of the endodermis.

Cadmium is readily taken up from soils by plants, depending on soil chemistry, and variably among species and cultivars; altered transpiration and xylem transport and/or translocation in the phloem could cause this variation in Cd accumulation, some degree of which is heritable. Using Triticum turgidum var. durum cvs Kyle and Arcola (high and low grain Cd accumulating, respectively), the objectives of this study were to determine if low-concentration Cd exposure alters transpiration, to relate transpiration to accumulation of Cd in roots and shoots at several life stages, and to evaluate the role of apoplastic bypass in the accumulation of Cd in shoots. The low abundance isotope 106Cd was used to probe Cd translocation in plants which had been exposed to elemental Cd or were Cd-naïve; apoplastic bypass was monitored using the fluorescent dye PTS (8-hydroxy-1,3,6-pyrenetrisulphonate). Differential accumulation of Cd by ‘Kyle’ and ‘Arcola’ could be partially attributed to the effect of Cd on transpiration, as exposure to low concentrations of Cd increased mass flow and concomitant Cd accumulation in ‘Kyle’. Distinct from this, exposure of ‘Arcola’ to low concentrations of Cd reduced translocation of Cd from roots to shoots relative to root accumulation of Cd. It is possible, but not tested here, that sequestration mechanisms (such as phytochelatin production, as demonstrated by others) are the genetically controlled difference between these two cultivars that results in differential Cd accumulation. These results also suggest that apoplastic bypass was not a major pathway of Cd transport from the root to the shoot in these plants, and that most of the shoot Cd resulted from uptake into the stele of the root via the symplastic pathway.

The environmental and seasonal effects on anatomical traits of Pinus taeda L. seedling roots were studied in the laboratory in three contrasting root growth media and also in typical outdoor nursery culture. Growth media with lower water regimen and high penetration resistance caused a reduction in lengths of the white and condensed tannin (CT) zones and acceleration of development of suberin lamellae in the endodermis. As a possible counter to this reduction in zone lengths, second-order laterals were produced closer to the tips of first-order laterals. This suggested there may be an advantage to producing more shorter roots under stressful conditions. Under outdoor nursery conditions (June to mid-December) the white zone was always a rather small part of the root system surface area (4.5% in December), but it dominated as a provider of cortical plasmalemma surface area (CPSA) in contact with modified soil solution (65% in December) because of its live cortex and capacity to increase nearly three fold the amount of CPSA per unit root length. The CT zone always provided most of the total root surface area (80% in December). Although it had no live cortex, a few cells of the CT zone endodermis remained non-suberized passage cells, perhaps giving this major part of the root system some capacity for ion and water absorption. A late summer increase in CPSA was due largely to the rapid production of mycorrhizae. Root systems were capable of very rapid replacement of roots lost due to undercutting and lateral root pruning. The great variation in CPSA per unit root length contained in the white, mycorrhizal and CT zones suggested a capacity to adapt rapidly to changing conditions.

Seeds of different cultivars of Glycine max (L.) Merr. (soybean) have strikingly different rates of water imbibition. Seeds that readily imbibe water are termed 'soft', while those that remain non-permeable, even after several days in water, are referred to as 'hard', 'stone', or 'impermeable' seeds. What prevents soybean hard seeds from taking up water? Previous work established that the initial imbibition of soft soybean seeds correlates with the presence of small cracks in the outermost cuticle that covers the seed coat, prompting a detailed analysis of soybean seed coat cutin. In this paper, it is shown that the outermost cuticle of the seed coat has an unusual chemical composition, lacking typical mid-chain-hydroxylated fatty acids but being relatively rich in other types of hydroxylated fatty acids. The cuticle of the impermeable cultivar studied contained a disproportionately high amount of hydroxylated fatty acids relative to that of the permeable ones. Moreover, a brief treatment with hot alkali released the omega-hydroxy fatty acid component of the outermost cuticle and created holes in it that caused the seeds to become permeable. This demonstrates that the outermost cuticle of the seed is the critical structure that prevents water uptake by hard seeds.

Soybean (Glycine max L. Merr.) is a versatile and important agronomic crop grown worldwide. Each year millions of dollars of potential yield revenues are lost due to a root rot disease caused by the oomycete Phytophthora sojae (Kaufmann & Gerdemann). Since the root is the primary site of infection by this organism, we undertook an examination of the physicochemical barriers in soybean root, namely, the suberized walls of the epidermis and endodermis, to establish whether or not preformed suberin (i.e. naturally present in noninfected plants) could have a role in partial resistance to P. sojae. Herein we describe the anatomical distribution and chemical composition of soybean root suberin as well as its relationship to partial resistance to P. sojae. Soybean roots contain a state I endodermis (Casparian bands only) within the first 80 mm of the root tip, and a state II endodermis (Casparian bands and some cells with suberin lamellae) in more proximal regions. A state III endodermis (with thick, cellulosic, tertiary walls) was not present within the 200-mm-long roots examined. An exodermis was also absent, but some walls of the epidermal and neighboring cortical cells were suberized. Chemically, soybean root suberin resembles a typical suberin, and consists of waxes, fatty acids, ω-hydroxy acids, α,ω-diacids, primary alcohols, and guaiacyl- and syringyl-substituted phenolics. Total suberin analysis of isolated soybean epidermis/outer cortex and endodermis tissues demonstrated (1) significantly higher amounts in the endodermis compared to the epidermis/outer cortex, (2) increased amounts in the endodermis as the root matured from state I to state II, (3) increased amounts in the epidermis/outer cortex along the axis of the root, and (4) significantly higher amounts in tissues isolated from a cultivar (‘Conrad’) with a high degree of partial resistance to P. sojae compared with a susceptible line (OX760-6). This latter correlation was extended by an analysis of nine independent and 32 recombinant inbred lines (derived from a ‘Conrad’ × OX760-6 cross) ranging in partial resistance to P. sojae: Strong negative correlations (−0.89 and −0.72, respectively) were observed between the amount of the aliphatic component of root suberin and plant mortality in P. sojae-infested fields.