The development and ultrastructure of the testa and tracheid bar inErythrina lysistemon Hutch. (Leguminosae: Papilionoideae)

Summary The development of the testa was studied inErythrina lysistemon using both light and electron microscopy. Cells of the outer epidermis of the outer integument divide anticlinally and undergo radial elongation to form a palisade layer. The outer tangential walls are thickened at an early stage, and deposition of fluted thickenings on the radial walls occurs at maturity. Palisade cells in the hilar region differentiate from sub-funicular tissue, and at maturity the outer ends of the cells undergo extensive deposition of secondary walls and associated lignification. The light line occurs at the junction between the outer, thickened portions of the cells and the inner, less thickened portions. An electron-translucent (suberised) cap develops in the outer tangential walls of the palisade cells at a late stage. Microtubules and dictyosomes are closely associated with the developing thickenings in palisade and tracheid bar, and the microtubules run parallel to the wall microfibrils. Differentiation of the tracheid bar coincides with final secondary wall deposition and lignification in the hilar palisade. The cells of the tracheid bar are dead at maturity, but are surrounded by sheaths of elongate parenchyma.     

Formation of macromolecular lignin in ginkgo xylem cell walls as observed by field emission scanning electron microscopy

Formation of macromolecular lignin in ginkgo cell walls. In the lignifying process of xylem cell walls, macromolecular lignin is formed by polymerization of monolignols on the pectic substances, hemicellulose and cellulose microfibrils that have deposited prior to the start of lignification. Observation of lignifying secondary cell walls of ginkgo tracheids by field emission scanning electron microscopy suggested that lignin-hemicellulose complexes are formed as tubular bead-like modules surrounding the cellulose microfibrils (CMFs), and that the complexes finally fill up the space between CMFs. The size of one tubular bead-like module in the middle layer of the secondary wall (S2) was tentatively estimated to be about 16+/-2 nm in length, about 25+/-1 nm in outer diameter, with a wall thickness of 4+/-2 nm; the size of the modules in the outer layer of the secondary wall (S1) was larger and they were thicker-walled than that in the middle layer (S2). Aggregates of large globular modules were observed in the cell corner and compound middle lamella. It was suggested that the structure of non-cellulosic polysaccharides and mode of their association with CMFs may be important factors controlling the module formation and lignin concentration in the different morphological regions of the cell wall.     

Tensile growth stress and lignin distribution in the cell walls of yellow poplar, Liriodendron tulipifera Linn.

Five specimens that contained a continuous gradient of wood, from normal to tension wood regions, were collected from an inclined yellow poplar (Liriodendron tulipifera), and the released strain of tensile growth stress was quantified. Ultraviolet (UV) microspectrophotometry was used to examine the relationship between lignin distribution in the cell wall and the intensity of tensile growth stress. The UV absorption of the secondary wall and the cell corner middle lamella decreased with increasing tensile released strain (i.e., tensile growth stress). The UV absorption in the compound middle lamella region remained virtually constant, irrespective of the tensile released strain. The absorption maximum (5_max) remained virtually constant in the secondary wall, the cell corner middle lamella, and the compound middle lamella region at 273-274, 277-278, and 275-278 nm respectively, irrespective of the tensile released strain. The ratios of the UV absorbance at 280 to 260 nm and 280 to 273 nm of the secondary wall decreased with increasing tensile released strain. The ratios in the cell corner and compound middle lamella region remained constant, irrespective of the tensile released strain. The lignin content of the secondary wall decreased, while the syringyl/guaiacyl ratio increased with increasing tensile released strain. Gelatinous fibers were not observed in the tension wood regions, but the secondary wall became gelatinous-layer-like, i.e., the lignin content and microfibrillar angle decreased and the cellulose content increased. A definite gelatinous layer seems to be important for generating greater tensile growth stress. It is concluded that a decrease in lignin and an increase in cellulose microfibrils parallel to the fiber axis in the secondary wall are necessary to produce large tensile growth stress.     

Free-space marker studies on the leaf ofZea mays L.

Summary Two free-space marker procedures (Prussian blue and lanthanum nitrate) were employed to determine the pathway(s) followed by water and solutes in the transpiration stream after their introduction into the xylem of small and intermediate bundles, and the effectiveness of the suberin lamellae of the bundle-sheath cells as a barrier to the movement of tracer ions (Fe³⁺ and La³⁺). Judged from the distribution of Prussian-blue crystals (insoluble, crystalline deposits resulting from the precipitation of ferric ions by ferrocyanide anions) and lanthanum deposits, water and the tracer ions moved readily from the lumina of the vessels into the apoplast (cell wall continuum) of the phloem and bundle-sheath cells via portions of vessel primary walls not bearing lignified secondary wall thickenings. Prussian blue and lanthanum deposits were abundant on the bundlesheath cell side of the bundle sheath/mesophyll interface but few occurred on that of the mesophyll, indicating that the suberin lamella is an effective barrier to apoplastic movement of both ferric and lanthanum ions. The presence of Prussian-blue crystals and lanthanum deposits in the compound middle lamella of the radial wall of the bundle-sheath cells indicates that the compound middle lamella provides an apoplastic pathway for transpirational water from the xylem to the evaporating surfaces of the mesophyll and epidermal cells.     

Cytochemical studies of pectin digestion in epidermis with specific cell separation

Summary This investigation concerns a unique type of epidermal cells in the anther ofStrelitzia reginae. At dehiscence these cells are released and form multicellular threads. The radial and tangential middle lamella regions of their cell walls disintegrate by the formation of numerous growing and fusing cavities. The possibility that this process could be due to digestion by pectinase was elucidated by use of cytochemical methods. In immature ordinary and thread-forming cells staining for pectin with hydroxylamine-ferric chloride yielded reaction products mainly in the middle lamella region and the subcuticular layer. After the appearance of cavities reaction took place around but not inside these formations. Treatment with fungal pectinase caused degradation of cell walls in ordinary epidermal tissue. Mature cell walls appeared more resistant to the lytic action than immature ones. In thread-forming tissue, independent of the stage of maturation, digestion of the pectin rich regions was induced. However, the fungal enzyme was not able to produce cavities. No pectin reaction with hydroxylamine-ferric chloride was obtained after pectinase treatment.     

Microfibrillar structure of growing cell wall in a coenocytic green alga,Boergesenia forbesii

A fine structure of cell wall lamellae in a coenocytic green algaBoergesenia forbesii was examined by electron microscopy. The wall has a polylamellate structure containing cellulose microfibrils 25 to 30 nm in diameter. The outer surface of the cell was covered by a thin structureless lamella, underneath which existed a lamella containing randomly-oriented microfibrils. The major part of the wall consisted of two types of lamellae, multifibrillar lamella and a transitional, matrix-rich one. In the former, microfibrils were densely arranged more or less parallel with each other. In the transitional lamella, existing between the multifibrillar ones, the microfibril orientation shifted about 30° within the layer. The fibril orientation also shifted 30° between adjacent transitional and multifibrillar layers, and consequently the microfibril orientation in the neighboring multifibrillar layers shifted 90°. It was concluded that the orientation rotated counterclockwise when observed from inside the cell. Each lamella in the thallus wall become thinner with cell expansion, but no reorientation of microfibrils in the outer old layers was observed. In the rhizoid, the outer lamellae sloughed off with the tip growth.     

Dynamic Changes in Distribution of Lignin and Hemicelluloses in Cell Walls During Differentiation of Secondary Xylem in Eucommia ulmoides (in English)

The dynamic changes in the distribution of lignin and hemicelluloses (xylans and xyloglucans) in cell walls during the differentiation of secondary xylem in Eucommia ulmoides Oliv. were studied by means of ultraviolet light microscopy and transmission electron microscopy combined with immunogold labelling. In the cambial zone and cell expansion zone, xyloglucans were localized both in the tangential and radial walls, but no xylans or lignin were found in these regions. With the formation of secondary wall S1 layer, lignin occurred in the cell corners and middle lamella, while xylans appeared in S1 layer, and xyloglucans were localized in the primary walls and middle lamella. In pace with the formation of secondary wall S2 and S3 layer, lignification extended to S1, S2 and S3 layer in sequence, showing a patchy style of lignin deposition. Concurrently, xylans distributed in the whole secondary walls and xyloglucans, on the other hand, still localized in the primary walls and middle lamella. The results indicated that along with the formation and lignification of the secondary wall, great changes had taken place in the cell walls. Different parts of cell walls, such as cell corners, middle lamella, primary walls and various layers of secondary walls, had different kinds of hemicelluloses, which formed various cell wall architecture combined with lignin and other cell wall components.     

Cellulose and pectin localization in roots of mycorrhizalAllium porrum: labelling continuity between host cell wall and interfacial material

Two different types of contacts (or interfaces) exist between the plant host and the fungus during the vesicular-arbuscular mycorrhizal symbiosis, depending on whether the fungus is intercellular or intracellular. In the first case, the walls of the partners are in contact, while in the second case the fungal wall is separated from the host cytoplasm by the invaginated host plasmamembrane and by an interfacial material. In order to verify the origin of the interfacial material, affinity techniques which allow identification in situ of cell-wall components, were used. Cellobiohydrolase (CBH I) that binds to cellulose and a monoclonal antibody (JIM 5) that reacts with pectic components were tested on roots ofAllium porrum L. (leek) colonized byGlomus versiforme (Karst.) Berch. Both probes gave a labelling specific for the host cell wall, but each probe labelled over specific and distinct areas. The CBH I-colloidal gold complex heavily labelled the thick epidermal cell walls, whereas JIM 5 only labelled this area weakly. Labelling of the hypodermis was mostly on intercellular material after treatment with JIM 5 and only on the wall when CBH I was used. Suberin bands found on the radial walls were never labelled. Cortical cells were mostly labelled on the middle lamella with JIM 5 and on the wall with CBH I. Gold granules from the two probes were found in interfacial material both near the point where the fungus enters the cell and around the thin hyphae penetrating deep into the cell. The ultrastructural observations demonstrate that cellulose and pectic components have different but complementary distributions in the walls of root cells involved in the mycorrhizal symbiosis. These components show a similar distribution in the interfacial material laid down around the vesicular-arbuscular mycorrhizal fungus indicating that the interfacial material is of host origin.     

Anatomic Characteristics Associated with Head Splitting in Cabbage (Brassica oleracea var. capitata L.)

Cabbage belonging to Brassicaceae family is one of the most important vegetables cultivated worldwide. The economically important part of cabbage crop is head, formed by leaves which may be of splitting and non-splitting types. Cabbage varieties showing head splitting causes huge loss to the farmers and therefore finding the molecular and structural basis of splitting types would be helpful to breeders. To determine which anatomical characteristics were related to head-splitting in cabbage, we analyzed two contrasting cabbage lines and their offspring using a field emission scanning electron microscope. The inbred line “747” is an early head-splitting type, while the inbred line “748” is a head-splitting-resistant type. The petiole cells of “747” seems to be larger than those of “748” at maturity; however, there was no significant difference in petiole cell size at both pre-heading and maturity stages. The lower epidermis cells of “747” were larger than those of “748” at the pre-heading and maturity stages. “747” had thinner epidermis cell wall than “748” at maturity stage, however, there was no difference of the epidermis cell wall thickness in the two lines at the pre-heading stage. The head-splitting plants in the F₁ and F₂ population inherited the larger cell size and thinner cell walls of epidermis cells in the petiole. In the petiole cell walls of “747” and the F₁ and F₂ plants that formed splitting heads, the cellulose microfibrils were loose and had separated from each other. These findings verified that anomalous cellulose microfibrils, larger cell size and thinner-walled epidermis cells are important genetic factors that make cabbage heads prone to splitting.     

Ultrastructure of the haustorial interface and apoplastic continuum between host and the root hemiparasiteOlax phyllanthi (Labill.) R. Br. (Olacaceae)

Summary Structural features of haustorial interface parenchyma of the root hemiparasiteOlax phyllanthi are described. Walls contacting host xylem are thickened non-uniformly with polysaccharides, not lignin, and show only a thin protective wall layer when abutting pits in walls of host xylem vessels or tracheids. Lateral walls of interface parenchyma exhibit an expanded middle layer of open fibrillar appearance, sometimes with, but mostly lacking adjoining layers of dense wall material. Free ribosomes and rough endoplasmic reticulum are prominent and occasional wall ingrowths present. Experiments involving transpirational feeding of the apoplast tracers lanthanum nitrate or uranyl acetate to host roots cut below haustorial connections, indicate effective apoplastic transfer from host to parasite root via the haustorium. Deposits of the tracers suggest a major pathway for water flow through host xylem pits, across the thin protective wall layer, and thence into the haustorium via the electronopaque regions of the terminal and lateral walls of the contact parenchyma. Graniferous tracheary elements and walls of parenchyma cells of the body of the haustorium appear to participate in tracer flow as do walls of cortical cells, stele parenchyma and xylem conducting elements of the parasite root, suggesting that both vascular and non-vascular routes are involved in extracytoplasmic transfer of xylem sap from host to parasite. The Casparian strip of the endodermis and the suberin lamella of the exodermis of theOlax root act as barriers to flow within the system.     

Correlation between the distribution of lignin and pectin and distribution of sorbed metal ions (lead and zinc) on coir (Cocos nucifera L.)

Plant fibres are capacious for sorption of metal ions, and can be used in water cleaning. Knowledge about the sorption will help in selection of the fibre and optimisation of its chemical modification, if any. The aim of this paper is to investigate the connection, if any, between the distribution of lignin and pectin and the loading of Pb and Zn on coir (mesocarp fibres from Cocos nucifera L.). The coir consisted mainly of xylem and a fibre sheath. The lignin was evenly distributed in the cell walls of the fibre sheath, but in the xylem, there was no detectable content in the compound middle lamella, and a smaller content of lignin in the secondary walls than in the walls of the fibre sheath. The only detectable content of pectin in the fibre sheath walls was in the middle lamella, cell corners and extracellular matrix, while in the xylem, the pectin was almost evenly distributed in the wall, with a higher concentration in the middle lamella and cell corners. All cell walls facing the lacuna had a high content of pectin. The metal ions were mainly loaded on the xylem and cell walls facing the lacuna, maybe with an additional trend to be loaded on the large fibres. Lead was distributed on and across the whole secondary wall. Zinc was loaded on the secondary walls, but there was no information about the distribution across the wall. If there is a simple correlation between the loading of metal ions and the distribution of lignin or pectin, these investigations point at no correlation with lignin and a positive correlation with pectin. It has to be stressed that these conclusions are made on limited material and are therefore preliminary in nature.     

Development of mestome sheath cells in leaves ofAegilops comosa var.thessalica

Summary The development of mestome sheath cells ofAegilops comosa var.thessalica was studied by electron microscopy. Anatomical and cytological observations show that this grass belongs to the C₃ or non-Kranz plants. In the asymmetrically thickened walls of mestome sheath cells a suberized lamella is present. This lamella is deposited asynchronously. In the midrib and the large lateral bundles it appears first in the outer and inner walls and usually later in the radial walls. In the small lateral bundles its appearance is delayed in the inner walls of those cells situated on the xylem side. At maturity the suberized lamella is observed in all cell walls; however, in the small lateral bundles it is partly or totally absent from the walls of some cells situated on the xylem side. Tertiary wall formation is asynchronous as well, for it generally follows the deposition pattern of the suberized lamella.During the development of the mestome sheath cells microtubules show marked changes in their number and orientation, being fewer and longitudinal during suberin deposition. Dictyosomes are very active and may be involved in primary and tertiary wall formation. Endoplasmic reticulum cisternae are abundant and partly smooth, while plasmalemmasomes may function to reduce the plasmalemma extension. However, cytoplasmic structures that are clearly involved in suberin synthesis could not be identified.Suberized lamellae react strongly with silver hexamine. This is probably due to post-fixation with osmium tetroxide.On the basis of structural characteristics the mestome sheath may be regarded as an endodermis (cf., alsoFahn 1974). The significance of this view for water and assimilate exchange between the mesophyll and the bundle is discussed.This report represents a portion of a doctoral dissertation.     

Changes in microfibril arrangement on the inner surface of the epidermal cell walls in the epicotyl of Vigna angularis ohwi et ohashi during cell growth

Throughout the entire period of cell growth, the microfibrils on the inner surface of the outer tangential walls of the epidermal cells of Vigna angularis epicotyls are running parallel to one another and their orientation differs from cell to cell. Although transverse, oblique and longitudinal microfibrils can be observed irrespective of cell age, the frequency distribution of microfibril orientation changes with age. In young cells, transversely oriented microfibrils predominate. In cells of medium age, which are still undergoing elongation, transverse, oblique and longitudinal microfibrils are present in quite similar frequencies. In old, non-growing cells, longitudinally oriented microfibrils are predominent. A decrease in the relative frequency of transversely oriented microfibrils with cell age was also observed in the radial epidermal walls.     

Microfibril orientation in differentiating and maturing fibre and parenchyma cell walls in culms of bamboo (Phyllostachys viridi-glaucescens (Garr.) Riv. & Riv.)

Results of trials using chemical and enzymatic wall extractants for the removal of matrix materials for in situ observations of newly deposited microfibrils are described. Observations were then made of the orientation of microfibrils on the inner walls of differentiating and maturing fibres and parenchyma cells under the FESEM. Orientation changes were similar in both cell types. During very early primary wall development, deposition of microfibrils was in more or less axial alignment, which was later superseded by microfibrils in transverse orientation (90^o to the long axis). A transverse orientation of microfibrils remained throughout much of primary wall synthesis, until an abrupt shift occurred to a sloped orientation during late primary wall synthesis. Microfibrils of the first secondary wall layer were in axial alignment or steeply sloped. In subsequent secondary wall deposition there was an alternation between a transverse and a sloped or axial alignment in maturing fibres and parenchyma cells.     

Mechanisms of infection of plants by nitrogen fixing organisms

Heterotrophic nitrogen-fixing microorganisms can enter plants via wounds, root hairs or intact epidermises. All at some stage need the ability to digest primary cell walls and/or middle lamellas. None appears to digest secondary walls. The ability of any organism to infect a particular plant reflects (a) the enzymes produced by the microorganism (and possibly, as part of its reaction, the plant); (b) the exact nature of the primary wall; (c) the distribution of secondary walls. Plants may respond to infection by hypersensitive and other reactions which could be triggered by production of cell wall fragments. Infection threads of secondary wall material may be essential for root hair infection and where cell boundaries are crossed. Entry into host cells other than by infection threads involves a delicate balance between endophyte and host. This may only be achieved in one or a few cells, which may then divide repeatedly to produce a symbiotic structure.     

杜仲次生木质部分化过程中木质素与半纤维素组分在细胞壁中分布的动态变化

利用紫外光显微镜、透射电子显微镜结合免疫胶体金标记,研究了杜仲（Eucommia ulmoides Oliv.)次生木质部分化过程中木质素与半纤维素组分（木葡聚糖和木聚糖）在细胞壁分布的动态变化。在形成层及细胞伸展区域,细胞壁具有木葡聚糖的分布,而没有木聚糖和木质素沉积,随着次生壁S1层的形成,木质素出现在细胞角隅和胞间层,木聚糖开始出现在S1层中,此时木葡聚糖则分布在初生壁和胞间层;随着次生,壁S2层及S3层的形成和加厚,木质逐逐步由细胞角隅和胞间层扩展到S1、S2和S3层,其沉积呈现出不均匀的块状或片状沉积模式,在次生壁各层形成与其木质化的同时,木聚糖逐渐分布于整个次生壁中,而木糖聚糖仍局限分布于初生壁和胞间层。结果表明,随着细胞次生壁的形成与木质化,细胞壁结构发生较大变化。细胞壁的不同区域,如细胞角隅、胞间层、初生壁和次生壁各层,具有不同的半纤维素组成,其与木质等细胞壁组分结构构成不同的细胞壁分子结构。     

Antibody localization of extensin in cell walls of carrot storage roots

The accumulation and cross-linking of hydroxyproline-rich glycoproteins (HRGPs) in cell walls of dicotyledonous plants has been correlated with a number of wall-strengthening phenomena. Polyclonal antibodies raised against glycosylated extensin-1, the most abundant HRGP in carrot (Daucus carota L.) cell walls, recognize this antigen on gel and dot blots and on thin sections of epoxy-embedded carrot-root cell walls. Since wall labeling can be largely reduced by preincubating the antibodies with purified extensin-1, most labeling can be attributed to recognition of this antigen. The remaining label may be the result of recognition of extensin-2, a second carrot HRGP, or other wall components (cellulose, hemicellulose and pectin are not recognized). Extensin-1 label was distributed quite uniformly across the cell wall but was absent from the expanded middle lamella at the intersection of three or more cells and was reduced in the narrow middle lamella between two cells. This distribution is essentially the same as that of cellulose. Because of limitations of this labeling technique, it is not possible to construct a complete model of the structure of the cross-linked extensin matrix. Nonetheless, short, linear arrays of gold particles may represent small portions of the extensin matrix or of individual extensin molecules as they are exposed on the surface of sections. These and other results presented here indicate that: a) newly synthesized extensin is added to the wall by intussusception; b) extensin cannot cross the middle lamella separating the walls of adjacent cells; and c) incorporation of extensin is a late event in the development of phloem-parenchyma cell walls in carrot.Abbreviations dE-1 antibodies antibodies raised against deglycosylated extensin 1 - ELISA enzyme-linked immunosorbant assay - gE-1 antibodies antibodies raised against glycosylated extensin 1 - HRGP hydroxyproline-rich glycoprotein - PAGE polyacrylamide gel electrophoresis - RG-1 rhamnogalacturonan I - SDS sodium dodecyl sulfate     

ON THE ULTRASTRUCTURE OF CAMBIUM AND ITS VASCULAR DERIVATIVES. III. THE SECONDARY WALLS OF THE SIEVE ELEMENTS OF PINUS STROBUS

The sieve elements of Pinus strobus have thick, lamellate secondary walls, which are composed predominantly of cellulose and lesser amounts of polyuronides and pectins. Eight to 10 lamellae may be present in walls 2-3 μ in thickness. Each lamella represents a plane of high cellulose density which results from intersection of two parallel sets of fibrils. Polyuronides and pectins are more or less evenly distributed in the wall, possibly with a greater concentration near the middle lamella and the inner surface. Resemblance of these walls to the so-called nacré walls is indicated, and it is possible that the two represent the same structure.     

Antibodies raised against tobacco aquaporins of the PIP2 class label viscin tissue of the explosive dwarf mistletoe fruit

Dwarf mistletoes, genus Arceuthobium , are parasitic flowering plants and forest pests. In western North America, Arceuthobium americanum (lodgepole pine dwarf mistletoe) is principally found on Pinus contorta var. latifolia (lodgepole pine). Dwarf mistletoes disperse their seeds by an explosive process that involves the buildup of hydrostatic pressure within a mucilaginous fruit tissue called the 'viscin'. Living viscin tissue envelops the discharged seeds. This study examined the possibility that aquaporins, critical in plant water relations, might be found in the dwarf mistletoe fruit, specifically the viscin cells. An antibody raised against a tobacco plasma membrane intrinsic 2 (PIP2) aquaporin was used with a gold-labeled secondary antibody to probe dwarf mistletoe fruit at various developmental stages. Viscin cell plasma membranes were successfully labeled with the anti-tobacco probe, and the validity of the immunolabeling was supported by Western blot analysis, showing a strong signal at about 30 kDa, which is at the expected size of a PIP2. A definitive immunolabeling pattern, supported by quantification of gold signal per membrane length, was observed: viscin cells sampled early in development had abundant gold label at their plasma membranes (1.93 ± 0.13 to 2.13 ± 0.33 gold particles per μm membrane), while other areas of the cells had no discernible label. Viscin cells sampled near the time of explosive discharge had significantly less label at the plasma membrane (0.21 gold particles ± 0.11 per μm membrane, P   <   0.05), and label was seen at vesicular membranes. Aquaporins likely have a role in directing water to the viscin mucilage early in development, but are retrieved via endocytosis to prevent excess water loss from viscin cells when discharge is imminent.     

Localization of glucuronoxylans in Japanese beech visualized by immunogold labelling

Summary An antiserum against glucuronoxylans (GXs) has been raised from a mouse. The dot-blot immunoassay and competitive inhibition test indicated that the antibodies could bind specifically to GXs. Therefore, the antiserum was used for immunogold labelling to investigate the localization of GXs in Japanese beech. Labelling of GXs was seen only in the secondary walls of xylem cells, but not in the primary walls or the middle lamella. GXs were evenly distributed in the secondary walls except for the outer part of the outer secondary-wall layer in which they were less abundant. The labelling density in each secondary-wall layer (S₁, S₂, and S₃) increased during cell wall formation. This result strongly suggests that the deposition of GXs occurs in a penetrative way.