DEVELOPMENTAL ANATOMY OF THE STEM OF WELFIA GEORGII,IRIARTEA GIGANTEA,AND OTHER ARBORESCENT PALMS: IMPLICATIONS FOR MECHANICAL SUPPORT

Changes in stem anatomy with radial position and height were studied for the arborescent palms Welfia georgii, Iriartea gigantea, Socratea durissima, Euterpe macrospadix, Prestoea decurrens, and Cryosophila albida. Vascular bundles are concentrated toward the stem periphery and peripheral bundles contain more fibers than central bundles. Expansion and cell wall thickening of fibers within vascular bundles continues throughout the life of a palm, even in the oldest tissue. Within individual vascular bundles, the fibers nearest the phloem expand first and fiber cell walls become heavily thickened. A front of expanding fibers moves outward from the phloem until all fibers within a vascular bundle are fully expanded and have thick cell walls. Peripheral vascular bundles differentiate first and inner bundles later. In the stem beneath the crown, vascular bundles and ground tissue cells show little or no size increase, but marked cell wall thickening during development for Welfia georgii. Beneath the crown, diameters of peripheral vascular bundles increase more than twofold for Iriartea gigantea, while diameters of central bundles do not increase. In Iriartea stems, ground tissue cells at the periphery elongate to accommodate expanding vascular bundles and cell walls become thickened to a lesser degree than in fibers; central ground tissue cells elongate markedly, but cell walls do not become thickened; and large lacunae form between central parenchyma cells. For Iriartea, Socratea, and Euterpe, sustained cell expansion results in limited, but significant increases in stem diameter. For all species, sustained cell wall thickening results in dramatic increases in stem stiffness and strength.     

Functional xylem in the post-veraison grape berry

A number of studies have shown a transition from a primarily xylem to a primarily phloem flow of water as fleshy fruits develop, and the current hypothesis to explain this transition, particularly in grape (Vitis vinifera L.) berries, is that the vascular tissue (tracheids) become non-functional as a result of post-veraison berry growth. In most studies, pedicels have been dipped in a vial containing an apoplastic dye, which was taken up into the entire peripheral and axial xylem vasculature of pre-veraison, but not post-veraison berries. The pressure plate/pressure membrane apparatus that is commonly used to study soil moisture characteristics was adapted and the pre- to post-veraison change in xylem functionality in grape berries was re-evaluated by establishing a hydrostatic (tension) gradient between the pedicel and a cut surface at the stylar end of the berry. Under the influence of this applied hydrostatic gradient, movement of the apoplastic tracer dye, basic fuchsin, was found in the pedicel and throughout the axial and peripheral xylem of the berry mesocarp. A similar movement of dye could be obtained by simply adjoining the stylar cut surface to a dry, hydrophilic wicking material. Since both pre- and post-veraison berries hydrate when the pedicel is dipped in water, it is hypothesized that the absence of dye movement into the vasculature of post-veraison berries indicates not a loss of xylem function, but rather the loss of an appropriate driving force (hydrostatic gradient) in the berry apoplast. Based on this hypothesis, and the substantial decrease in xylem flows that occur in intact grape berries at veraison, it is suggested that there may be significant changes in the pattern of solute partitioning between the fruit symplast and apoplast at veraison. It is further suggested that diurnal patterns in symplast/apoplast solute partitioning in grapes and other fleshy fruit, may explain the observed minimal xylem contribution to the water budgets of these fruits.     

Histochemical changes in the developing abscission layer in fruits of Prunus cerasus L.

Summary Abscission layer formation in the sour cherry (Prunus cerasus L.) during fruit maturation occurred in the transition zone between the fruit and the pedicel. The abscission layer, consisting of 5–8 rows of cells, was first identified by its low affinity for haematoxylin. The walls of cells in the abscission layer contained less total polysaccharides than adjacent cells. The pectins were degraded and the cellulose was partially broken down resulting in cell separation. The Ca level in the abscission zone decreased and Ca and Mg were lost from the walls of cells in the layer during abscission. After the abscission layer formed, cells associated with the layer had a lower capacity to bind ⁴⁵Ca than cells distal or proximal to the layer.Michigan Agricultural Experiment Station Journal Article No. 4607     

The Anatomy of Fruit Abscission in Loganberries

Loganberry fruits abscise at the base of the receptacle, justdistal to the sepals. As the fruit ripens, all cells of theabscission zone expand. The central parenchyma cells, due totheir position, appear to be the driving force behind abscission.Their expansion causes early cell-separation within a superficialzone of small cells and rupture of the epidermis at the sepal/receptaclejunction without prior dissolution of cell walls. However, othercells within the abscission zone have their walls degraded,mostly in the region of the middle lamella, as ripening progresses. Xylem transfer cells are found in abundance in the vascularbundles supplying the sepals. The outward curve of these bundlesinto the sepals brings the transfer cells into close proximitywith the abscission zone. A comparison of their distributionin loganberries with that in raspberries (MacKenzie, 1979),which are closely related but abscise at a different site, suggeststhat transfer cells may be implicated in the abscission process. The inevitable structural weakness brought about by the paucityof vascular tissue in the abscission zone relative to the morerobust pedicel may also predispose this area to separation. Anatomy, abscission, loganberry, Rubus idaeus x R, ursinus, Mailing Sunberry, transfer cells structure, fruit     

X-ray microanalysis of ion distribution within root cortical cells of the halophyte Suaeda maritima (L.) Dum.

The ion content of compartments within cortical cells of mature roots of the halophyte Suaeda maritima grown at 200 mol·m^-3 NaCl has been studied by X-ray microanalysis of freeze-substituted thin sections. Sodium and Cl were found in the vacuoles at about four-times the concentration in the cytoplasm or cell walls, whereas K was more concentrated in the cell walls and cytoplasm than in vacuoles. The vacuolar Na concentration was 12- to 13-times higher than that of K. The Na concentration of cell walls of cortical cells was about 95 mol·m^-3 of analysed volume. The cytoplasmic K concentration within the mature cortical cells was estimated to be 55 mol·m^-3 of analysed volume.     

Die induktion eines trenngewebes bei früchten von Prunus avium L. durch 2-Chloräthylphosphonsäure

Summary 2-Chloroethylphosphonic acid (CEPA) facilitates the separation of the fruit from the pedicel significantly. The application of 2,000 and 4,000 ppm CEPA in four sweet cherry varieties during maturation resulted in the formation of a complete abscission layer in the transition zone between pedicel and fruit. In contrast, in the untreated fruit no abscission layer was evident at maturity. The walls of the cells in the abscission layer contained less total polysaccharides than adjacent cells. Cellulose was partially broken down, and the pectins were degraded. The Ca and Mg content in the cell walls decreased. Thus the same histochemical changes are involved in natural and CEPA induced abscission.     

Separation of vascular cell walls from cortical cell walls of plant roots

Summary The cortex was physically separated from the stele of corn roots. The isolated walls from cortical cells were less dense than the walls isolated from stelar cells. The cell walls from each tissue were centrifuged on a step gradient composed of 50 and 60% sucrose for 5 min at 900 g. After the short centrifugation time, the cortical cell walls banded at the 50/60% interface while the vascular tissue walls pelleted through 60% sucrose. An aliquot of vascular cell walls was then marked with cytochromec. The marked cell walls were mixed with cortical cell walls and centrifuged on a 50/60% sucrose gradient and after 5 min, the vascular tissue walls were cleanly separated from the cortical cell walls. The experiment was repeated without prior separation of tissue types with another corn variety, carrot roots grown in culture, and pea roots. A clean separation of cell wall types was achieved after homogenization of intact roots in liquid nitrogen, extrusion from a nitrogen bomb, and centrifugation in sucrose gradients.     

Patterns of incorporation of d-galactose into cell wall polysaccharide of growing maize roots

Summary ¹⁴C-1-d-galactose was rapidly taken up by excised corn root-tips and efficiently converted to hexose units in cell wall polysaccharides. The label recovered in both hydrolysed pectin and hemicellulose was predominantly in galactose and only the -cellulose contained appreciable amounts of labelled glucose. There was no evidence for breakdown of labelled units after incorporation into the cell wall. It is suggested that the utilisation of this free galactose has not appreciably affected the normal metabolic pathway by which galactose is incorporated into plant cell walls.Advantage was taken of the specificity of this labelling to follow patterns of galactosyl incorporation in roots. Autoradiographs were prepared from adjacent longitudinal sections that had been extracted with ammonium oxalate solution and 24% (w/v) KOH respectively. The distribution of silver grains over these sections was compared with that over an unextracted section. Galactosyl units of pectin were incorporated in young cell walls in all tissues investigated. The pattern closely resembled that noted in earlier work for uronosyl and pentosyl incorporation. In pith and cortical cells, galactosyl units of hemicellulose were deposited at a maximum rate in walls approaching the end of their growth when pentose incorporation was low. Because branched alkali-soluble polysaccharides containing galactose and pentose have been isolated from several tissues of corn, similar compounds are likely to exist in the root. It is proposed that the continued elaboration of such a polysaccharide might continue after deposition, and the addition of galactosyl units may be a factor which limits further plastic extension of the wall.     

The path of photosynthate translocation into citrus fruit

Abstract The path of [¹⁴C]photosynthate translocation into citrus fruit was examined to determine which anatomical and physiological features were involved in this process. Experiments were conducted during the final pre-harvest months of 2 years grapefruit crops (Citrus paradisi Macf. cv. ‘Marsh’). A source leaf nearest the fruit was exposed to ¹⁴CO₂ for 1 h + 5 h ambient air, followed by dissection of vascular and phloem-free tissues in the fruit quarter directly aligned with the source. Radioactivity in each tissue was quantified after separation and extraction in boiling 80% ethanol. Peel (flavedo+albedo) contained an average 35% of the label in the quarter fruit, but an additional 20% was localized entirely in dorsal vascular bundles along exterior walls of juice segments. Less [¹⁴C]photosynthate was recovered from other vascular tissues and was nearly absent from adjacent mature seeds. Radioactivity in the single layer of segment epidermis, however, averaged 17% of that in the quarter fruit. Juice tissues interior to this accumulated only 17% of the total. No phloem tissue was evident in either the segment epidermis or juice tissues, but over 70% of the [¹⁴C]assimilates in the latter were localized in thread-like stalks which attach juice vesicles to dorsal vascular bundles. In addition, labelled hexose/sucrose ratios in these structures increased with distance from the vascular bundle. The majority of photosynthates, therefore, entered citrus fruit via dorsal vascular bundles and were partially hydrolysed during slow transfer through non-vascular segment epidermis and juice stalks.     

Mechanisms for shaping,orienting, positioning and patterning plant secondary cell walls

Xylem vessels are cells that develop a specifically ornamented secondary cell wall to ensure their vascular function, conferring both structural strength and impermeability. Further plasticity is given to these vascular cells by a range of different patterns described by their secondary cell walls that—as for the growth of all plant organs—are developmentally regulated. Microtubules and their associated proteins, named MAPs, are essential to define the shape, the orientation, the position and the overall pattern of these secondary cell walls. Key actors in this process are the land-plant specific MAP70 proteins which not only allow the secondary cell wall to be positioned at the cell cortex but also determine the overall pattern described by xylem vessel secondary cell walls.Key words: xylem/wood vessels, tracheary elements, secondary cell wall, cell wall patterning, microtubules, microtubule-associated proteins, MAP70Xylem formation has been one of the key steps of plant evolution. These physically strong tube cells allowed plants to colonize land by reinforcing their upright position against gravity and resisting desiccation by permitting water conduction throughout the plant body. This double role is fulfilled by specific conducting wood cells—the tracheary elements (TEs). These cells represent the cellular units of the adjustable plant vasculature, which relies on the three structural characteristics of TEs: (1) these cells develop a secondary cell wall to resist pressure exerted by the sap they will conducted, (2) these cells undergo programmed cell death (PCD) to hollow out their entire cytoplasmic content to form a conduit for the sap and (3) these cells will undergo a terminal perforation at their basal end (with respect to the corresponding meristem) to form a complete functional vascular cylinder which will connect with the underlying vascular vessels once terminally differentiated.^1,2 TEs are further characterized by a diversity of organizational pattern described by their secondary cell wall, which can be annular or spiral (referred to as protoxylem-type ornamentations) reticulate or pitted (referred to as metaxylem-type ornamentations).^3,4 These differently ornamented TEs are developmentally regulated and for protoxylemtype TEs appear during the development of early primary tissues (annular TEs are mostly observed in developing embryos) while metaxylem-type TEs appear in the later development of primary and secondary tissues (they represent the TEs present in wood). Annular and spiral TEs are first formed in organs undergoing primary growth and are considered to be “extendable” (their pattern in rings and spirals does not oppose further extension of the TE cell) during the growth of this organ. Once the growing organ has attained a certain size these TEs will be crushed by the surrounding tissue whilst the more heavily reinforced reticulate and pitted TEs will form to insure the vascular flow and strengthen the entire organ. In short, the modularity and plasticity of this plant vascular system is directly dependant on the differentiation and the type of cell wall ornamentation of its constituent TEs. The establishment of such regular patterning of secondary cell walls has been attributed to the underlying cortical microtubule array that predefines the cell wall depositions (reviewed in ref. ²). Pharmacological modulation of microtubule properties in both whole plants and in vitro TE differentiating systems leads to severe defects in the patterning, orientation, smoothness and deposition of TE secondary cell walls (reviewed in ref. ²).     

Microtubule rearrangements accompanying dedifferentiation in mesophyll cultures ofZinnia elegans L.

Summary To determine the orientation of cortical microtubule arrays in mesophyll cells ofZinnia, a new technique designed to increase the rate of fixation of excised leaf tissue and subsequent permeabilization of mesophyll cell walls was developed. This procedure resulted in immunolabeling of high percentages of mesophyll cells, making it possible to quantify cells with different types of cortical microtubule arrays. When developing palisade mesophyll cells were fixed in situ, most of the cells had cortical microtubules organized in parallel arrays oriented transverse to the long axis. Delay in the transfer of leaf tissue to fixative resulted in increased numbers of cells with random cortical microtubule orientations, indicating that arrays may become reoriented rapidly during leaf excision and cell isolation procedures. The role of wound-induced microtubule reorientation in mesophyll dedifferentiation and tracheary element development is discussed.Abbreviations BSA bovine serum albumin - CMT cortical microtubule - TE tracheary element - TBS tris-buffered saline     

Interactions between Developing Citrus Fruits and their Supportive Vascular System

The developing fruit is a strong sink, which demands large amountsof assimilates. A correlation between grapefruit (Citrus ParadisiMacf., var. Marsh seedless) fruit size and its pedicel crosssectional area (CSA) can be demonstrated, suggesting a closeinteraction between them. The presence of fruits seems to determinethe developmental pattern of the vascular tissues within thebranches on which the fruits are borne. The pedicel normally terminates its diametric growth prior tothe linear phase of fruit growth. Fruit thinning (90%) and trunkgirdling, performed in order to minimize carbohydrate limitations,result in dramatic increases in fruit growth rate and pedicelCSA. Partial girdling of the pedicel causes a transient decreasein fruit growth. An increase in specific mass transport (SMT)through the existing vascular routes is the immediate response,due to the instantaneous upsurge of carbohydrate supply to individualfruit. Nevertheless, the rapid development of new vascular tissueshas been the major factor responsible for the long term enhancement,or recovery, of fruit growth, suggesting that limitation intransport capacity does occur. The cause and effect relationships between fruit and vasculardevelopment are discussed.Copyright 1995, 1999 Academic Press Source, sink, fruit growth, vascular development, transport limitation, specific mass transport (SMT), carbohydrate availability, competition, Citrus     

Soluble metabolites in intercellular regions of squash hypocotyl tissues: Implications for exudation

Summary Amino acids, organic acids, and sugars make up the bulk of ¹⁴C-assimilates that leach from freshly excised sections of subterranean hypocotyls of squash plants allowed to fix ¹⁴CO₂ photosynthetically. Based on kinetics of leakage, it appears that metabolites are present in significant levels in intercellular regions of squash hypocotyls and that they may be the major source of hypocotyl exudates. Although the anatomical relations of epidermal cells hinder excessive metabolite losses by subterranean portions of hypocotyls, avenues of diffusion are continuous along cell walls and middle lamellae between cortical tissue and the hypocotyl surface. re]19751115     

Extracellular complexation of Cd in the Hartig net and cytosolic Zn sequestration in the fungal mantle of Picea abies – Hebeloma crustuliniforme ectomycorrhizas

Compartmentation of heavy metals on or within mycorrhizal fungi may serve as a protective function for the roots of forest trees growing in soils containing elevated concentrations of metals such as Cd and Zn. In this paper we present the first quantitative measurements by X‐ray microanalysis of heavy metals in high‐pressure frozen and cryosectioned ectomycorrhizal fungal hyphae. We used this technique to analyse the main sites of Cd and Zn in fungal cells of mantle and Hartig net hyphae and in cortical root cells of symbiotic Picea abies – Hebeloma crustuliniforme associations to gain new insights into the mechanisms of detoxification of these two metals in Norway spruce seedlings. The mycorrhizal seedlings were exposed in growth pouches to either 1 mM Cd or 2 mM Zn for 5 weeks. The microanalytical data revealed that two distinct Cd‐ and Zn‐binding mechanisms are involved in cellular compartmentation of Cd and Zn in the mycobiont. Whereas extracellular complexation of Cd occurred predominantly in the Hartig net hyphae, both extracellular complexation and cytosolic sequestration of Zn occurred in the fungal tissue. The vacuoles were presumed not to be a significant pool for Cd and Zn storage. Cadmium was almost exclusively localized in the cell walls of the Hartig net (up to 161 mmol kg ^{? 1} DW) compared with significantly lower concentrations in the cell walls of mantle hyphae (22 mmol kg ^{? 1} DW) and in the cell walls of cortical cells (15 mmol kg ^{? 1} DW). This suggests that the apoplast of the Hartig net is a primary accumulation site for Cd. Zinc accumulated mainly in the cell walls of the mantle hyphae (111 mmol kg ^{? 1} DW), the Hartig net hyphae (130 mmol kg ^{? 1} DW) and the cortical cells (152 mmol kg ^{? 1} DW). In addition, Zn occurred in high concentrations in the cytoplasm of the fungal mantle hyphae (up to 164 mmol kg ^{? 1} DW) suggesting that both the cell walls and the cytoplasm of fungal tissue are the main accumulation sites for Zn in P. abies resulting in decreased Zn transfer from the fungus to the root.     

ULTRASTRUCTURE OF THE CORALLINACEAE (RHODOPHYTA) II. VEGETATIVE CELLS OF LITHOTHRIX ASPERGILLUM1

The ultrastructure of the calcareous red coralline alga Lithothrix aspergillum Gray and the development of the various tissue types has been studied. The sub-apical meristematic tissue alternately produces genicular or intergenicular cells. The genicular cells rapidly elongate and their cell walls thicken and become denser as more fibrillar wall material is laid down within the cell wall. These cells contain little cytoplasm and few organelles. The inter genicular cells which elongate only slightly during development have a small vacuole and many free starch grains in the cytoplasm. The peripheral cells in each inter genicular layer remain meristematic and form a cortical cell layer over the genicular cells. These cortical cells and the apical meristematic cells are covered by small epidermal cells which have extensive cell wall ingrowths between the chloroplasts. The inter genicular cells are calcified. Although the CaCO₃ is laid down within the cell walls, there is always a thin layer of CaCO₃-free organic cell wall material between the plasmalemma and the CaCO₃ impregnated wall. Only the distal tips of the genicular cells are calcified. In old genicular tissues of Lithothrix, secondary deposits of CaCO₃ of unknown crystallography are also found in the spaces between the cell walls. Thus there appear to be at least two mechanisms of calcification in this alga.     

Metabolism of sugars and organic acids in immature grape berries

Individual intact excised immature Sultana berries were supplied through the cut pedicel with ¹⁴C-sugars and organic acids. When ¹⁴C-hexoses were supplied malic and tartaric acids accounted for 25% and 10% of the total activity extracted after 24 hours, and sucrose was synthesized. It is proposed that the changes in the levels of organic acids during ripening are related to changes in the ability of the berry to synthesize them. Although administration of uniformly labeled sucrose resulted in the unequal labeling of glucose and fructose, the results indicate breakdown of sucrose by invertase. It is suggested that the route of entry of the pedicel-fed sugars into the berry may be different from the route taken by sugar translocated from the leaf.     

Extending the Microtubule/Microfibril Paradigm : Cellulose Synthesis Is Required for Normal Cortical Microtubule Alignment in Elongating Cells

The cortical microtubule array provides spatial information to the cellulose-synthesizing machinery within the plasma membrane of elongating cells. Until now data indicated that information is transferred from organized cortical microtubules to the cellulose-synthesizing complex, which results in the deposition of ordered cellulosic walls. How cortical microtubules become aligned is unclear. The literature indicates that biophysical forces, transmitted by the organized cellulose component of the cell wall, provide a spatial cue to orient cortical microtubules. This hypothesis was tested on tobacco (Nicotiana tabacum L.) protoplasts and suspension-cultured cells treated with the cellulose synthesis inhibitor isoxaben. Isoxaben (0.25–2.5 μm) inhibited the synthesis of cellulose microfibrils (detected by staining with 1 μg mL⁻¹ fluorescent dye and polarized birefringence), the cells failed to elongate, and the cortical microtubules failed to become organized. The affects of isoxaben were reversible, and after its removal microtubules reorganized and cells elongated. Isoxaben did not depolymerize microtubules in vivo or inhibit the polymerization of tubulin in vitro. These data are consistent with the hypothesis that cellulose microfibrils, and hence cell elongation, are involved in providing spatial cues for cortical microtubule organization. These results compel us to extend the microtubule/microfibril paradigm to include the bidirectional flow of information.     

Evidence for the priming role of the central retinula cell in ommatidium differentiation of Ephestia kuehniella

Summary In the developing compound eye of Ephestia kuehniella, within the advancing front of differentiation, regular cell clusters arise which consist of a central cell and two flanking cells. The central cell is destined to become the basal retinula cell later in development. Its crucial role in ommatidium formation is confirmed by ³H-thymidine labelling. Eye anlagen labelled early in the pupal stage incorporate thymidine within two distinct zones along the front of differentiation. After the ommatidia are completely differentiated, both zones contain labelled nuclei of all cell types which participate in ommatidia formation. Within the posterior zone, however, the basal retinula cells are always unlabelled, whereas in the anterior they show labelled nuclei. From this observation it must be concluded that the basal retinula cell first terminates proliferation (either alone or together with a few other cells) to become differentiated as the central retinula cell. These results agree with those found in Drosophila and indicate that the ordered stepwise addition of cells to a central founder cell is a widespread principle of ommatidia formation in insects.     

PRPs localized to the middle lamellae are required for cortical tissue integrity in Medicago truncatula roots

Key message

A family of repetitive proline-rich proteins interact with acidic pectins and play distinct roles in legume root cell walls affecting cortical and vascular structure.

Abstract

A proline-rich protein (PRP) family, composed of tandemly repeated Pro-Hyp-Val-X-Lys pentapeptide motifs, is found primarily in the Leguminosae. Four distinct size classes within this family are encoded by seven tightly linked genes: MtPRP1, MtPRP2 and MtPRP3, and four nearly identical MtPRP4 genes. Promoter fusions to β-glucuronidase showed strong expression in the stele of hairy roots for all 4 PRP genes tested, with additional expression in the cortex for PRP1, PRP2 and PRP4. All except MtPRP4 are strongly expressed in non-tumorous roots, and secreted and ionically bound to root cell walls. These PRPs are absent from root epidermal cell walls, and PRP accumulation is highly localized within the walls of root cortical and vascular tissues. Within xylem tissue, PRPs are deposited in secondary thickenings where it is spatially exclusive to lignin. In newly differentiating xylem, PRPs are deposited in the regularly spaced paired-pits and pit membranes that hydraulically connect neighboring xylem elements. Hairpin-RNA knock-down constructs reducing PRP expression in Medicago truncatula hairy root tumors disrupted cortical and vascular patterning. Immunoblots showed that the knockdown tumors had potentially compensating increases in the non-targeted PRPs, all of which cross-react with the anti-PRP antibodies. However, PRP3 knockdown differed from knockdown of PRP1 and PRP2 in that it greatly reduced viability of hairy root tumors. We hypothesize that repetitive PRPs interact with acidic pectins to form block-copolymer gels that can play distinct roles in legume root cell walls.

     

Rapid changes in cell wall pectic polysaccharides are closely associated with early stages of aerenchyma formation, a spatially localized form of programmed cell death in roots of maize (Zea mays L.) promoted by ethylene

Aerenchyma formation in roots of maize (Zea mays L.) involves programmed death of cortical cells that is promoted by exogenous ethylene (1 µL L⁻¹) or by endogenous ethylene produced in response to external oxygen shortage (3%, v/v). In this study, evidence that degeneration of the cell wall accompanies apoptotic-like changes previously observed in the cytoplasm and nucleus (Gunawardena et al. Planta 212, 205–214, 2001), has been sought by examining de-esterified pectins (revealed by monoclonal antibody JIM 5), and esterified pectins (revealed by monoclonal antibody JIM 7). In controls, de-esterified wall pectins were found at the vertices of triangular junctions between cortical cells (untreated roots). Esterified pectins in control roots were present in the three walls bounding triangular cell-to-cell junctions. After treatment with 3% oxygen or 1 µL L⁻¹ ethylene, this pattern was lost but walls surrounding aerenchyma gas spaces became strongly stained. The results showed that cell wall changes commenced within 0·5 d and evidently were initiated by ethylene in parallel with cytoplasmic and nucleoplasmic events associated with classic intracellular processes of programmed cell death.