Lise Bolt Jørgensen

Autophagic Components Contribute to Hypersensitive Cell Death in Arabidopsis

Autophagy has been implicated as a prosurvival mechanism to restrict programmed cell death (PCD) associated with the pathogen-triggered hypersensitive response (HR) during plant innate immunity. This model is based on the observation that HR lesions spread in plants with reduced autophagy gene expression. Here, we examined receptor-mediated HR PCD responses in autophagy-deficient Arabidopsis knockout mutants (atg), and show that infection-induced lesions are contained in atg mutants. We also provide evidence that HR cell death initiated via Toll/Interleukin-1 (TIR)-type immune receptors through the defense regulator EDS1 is suppressed in atg mutants. Furthermore, we demonstrate that PCD triggered by coiled-coil (CC)-type immune receptors via NDR1 is either autophagy-independent or engages autophagic components with cathepsins and other unidentified cell death mediators. Thus, autophagic cell death contributes to HR PCD and can function in parallel with other prodeath pathways.

Protein-accumulating cells and dilated cisternae of the endoplasmic reticulum in three glucosinolate-containing genera: Armoracia, Capparis, Drypetes

Three glucosinolate-containing species, Armoracia rusticana Gaertner, Meyer et Scherbius (Brassicaceae), Capparis cynophallophora L. (Capparaceae) and Drypetes roxburghii (Wall.) Hurusawa (Euphorbiaceae), are shown by both light and electron microscopy to contain protein-accumulating cells (PAC). The PAC of Armoracia and Copparis (former “myrosin cells”) occur as idioblasts. The PAC of Drypetes are usual members among axial phloem parenchyma cells rather than idioblasts. In Drypetes the vacuoles of the PAC are shown ultrastructurally to contain finely fibrillar material and to originate from local dilatations of the endoplasmic reticulum. The vacuoles in PAC of Armoracia and Capparis seem to originate in the same way; but ultrastructurally, their content is finely granular. In addition, Armoracia and Capparis are shown by both light and electron microscopy to contain dilated cisternae (DC) of the endoplasmic reticulum in normal parenchyma cells, in accord with previous findings for several species within Brassicaceae. The relationship of PAC and DC to glucosinolates and the enzyme myrosinase is discussed.

Localization of Plant Myrosinases and Glucosinolates

Summary Recent studies of Arabidopsis flower stalks show that the constituents of the mustard-oil bomb, namely myrosinase and glucosinolates, are stored, in adjacent cells. In vivo degradation of glucosinolates, possibly involving myrosin cells, and the transport of glucosinolates affect the localization of the myrosinase-glucosinolate system, showing a dynamic nature that is complementary to the “static” toxic mustard oil-bomb. Large variability in expression, of myrosinase among different species and organs have been detected, and this may indicate different primary roles of the myrosinase-glucosinolate system. Further research areas should include characterization of transporters and proteins responsible for the in vivo degradation, and also localization studies of the biosynthetic genes. This knowledge will create a basis for functional and evolutionary studies of the myrosinase-glucosinolate system.

Ozone Flux to Plasmalemma in Barley and Wheat is controlled by Stomata rather than by direct Reaction of Ozone with Cell Wall Ascorbate

Summary Morning, midday and afternoon values of stomatal conductance and apoplastic and whole-leaf ascorbate concentrations in leaves of barley and wheat were measured on the 7 th day of exposure of young seedlings to ozone enriched air (75–100 nL/L, 8 h d -1 ) in open-top chambers. The aim of the study was to explore whether the contributions of these factors to the limitation of ozone flux to plasmalemma are different in these species. Until midday, ozone flux density to the ozone-exposed mesophyll cell surface was found to be negligibly (4%) lower in barley compared with wheat. After midday, this difference increased to 12%, mostly due to a more rapid afternoon decrease of stomatal conductance in barley. The diurnal decline in apoplastic ascorbate concentration in barley was less pronounced than in wheat. The differences in diurnal courses of apoplastic ascorbate were apparently not related to whole-leaf ascorbate levels, found to be stable during the day in both species. Due to the thin cell wall (0.12 μm in barley and 0.10 μm in wheat), only a minor part of the ozone flux entering the mesophyll cell surface was calculated to be detoxified in the direct reaction with apoplastic ascorbate (7–14% in barley and 4–13% in wheat), causing a 5–6 times lower contribution of this reaction to total ozone decay than the contribution of the stomata. The contribution of other direct apoplastic ozone scavengers is estimated to be even less than that of ascorbate. The overall resultant difference in calculated ozone fluxes to mesophyll plasmalemma in barley and wheat (5% in the morning and 14% in the afternoon) was statistically insignificant. An increase of this difference is suggested if singlet oxygen and/or hydroxyl radical are generated in the apoplast under ozone.

Cyanohydrin glycosides of Passiflora: distribution pattern, a saturated cyclopentane derivative from P. guatemalensis, and formation of pseudocyanogenic α-hydroxyamides as isolation artefacts

Nineteen species of Passiflora (Passifloraceae) were examined for the presence of cyanogenic glycosides. Passibiflorin, a bisglycoside containing the 6-deoxy-beta-D-gulopyranosyl residue, was isolated from P. apetala, P. biflora, P. cuneata, P. indecora, P. murucuja and P. perfoliata. In some cases this glycoside co-occurs with simple beta-D-glucopyranosides: tetraphyllin A, deidaclin, tetraphyllin B, volkenin, epivolkenin and taraktophyllin. P. citrina contains passicapsin, a rare glycoside with the 2,6-dideoxy-beta-D-xylo-hexopyranosyl moiety, while P. herbertiana contains tetraphyllin A, deidaclin, epivolkenin and taraktophyllin, P. discophora tetraphyllin B and volkenin, and P. x violacea tetraphyllin B sulfate. The remaining species were noncyanogenic. The glycosides were identified by 1H and 13C NMR spectroscopy following isolation by reversed-phase preparative HPLC. From P. guatemalensis, a new glucoside named passiguatemalin was isolated and identified as a 1-(beta-D-glucopyranosyloxy)-2,3-dihydroxycyclopentane-1-carbonitrile. An isomeric glycoside was prepared by catalytic hydrogenation of gynocardin. alpha-Hydroxyamides corresponding to the cyanogenic glycosides were isolated from several Passiflora species. These alpha-hydroxyamides, presumably formed during processing of the plant material, behave as cyanogenic compounds when treated with commercial Helix pomatia crude enzyme preparation. Thus, the enzyme preparation appears to contain an amide dehydratase, which converts alpha-hydroxyamides to cyanohydrins that liberate cyanide; this finding is of interest in connection with analysis of plant tissues and extracts using Helix pomatia enzymes.

Substrate specificity in the biosynthesis of cyclopentanoid cyanohydrin glucosides

Abstract The biosynthesis of deidaclin in Turnera angustifolia and of linamarin in Passiflora morifolia were investigated using intact plant tissues. Radiolabelled precursors, 2-(2′-cyclopentenyl)[2- 14 C]glycine and l -[U- 14 C]-valine were fed to freshly harvested shoots either alone or together with the presumed nitrile intermediates, 2-cyclopentenecarbonitrile and 2-methylpropanenitrile. The cyanohydrin glucosides were isolated and purified, and the incorporation of the radioactive labels was determined after enzymatic degradation of the glucosides to cyanide. The labels from the amino acid precursors were incorporated into the nitrile group of their corresponding cyanohydrin glucosides, and the incorporation was in each case strongly inhibited by simultaneous feeding with either of the two nitriles. Turnera angustifolia was able to synthesize linamarin when fed with 2-methylpropanenitrile, even though linamarin could not be demonstrated to be present in this plant naturally.

Biosynthesis of cyanohydrin glucosides from unnatural nitriles in intact tissue of Passiflora morifolia and Turnera angustifolia

Passiflora morifolia, which under natural conditions contains cyanohydrin glucosides linamarin, lotaustralin and epilotaustralin, converted cyclopentanecarbonitrile, 2-cyclopentenecarbonitrile and 3-methylbutanenitrile into the corresponding cyanohydrin glucosides. Turnera angustifolia, which normally produces glucosides of cyclopentenone cyanohydrin, converted cyclopentanecarbonitrile, 2-methylpropanenitrile and 2-methylbutanenitrile, but not 3-methylbutanenitrile, into the corresponding cyanohydrin glucosides. Mixtures of epimers were produced when these glucosides contained chiral cyanohydrin carbon atoms. Feeding with cyclopentanecarbonitrile resulted in formation of 1-(beta-D-glucopyranosyloxy)cyclopentanecarbonitrile, a saturated analogue of deidaclin and tetraphyllin A. Neither plant utilized cyclopropanecarbonitrile as substrate. The experiments demonstrate broad substrate specificity of nitrile hydroxylases present in these plants. A novel glycoside, 2-[6-O-(beta-D-xylopyranosyl)-beta-D-glucopyranosyloxy]propane (isopropyl primeveroside), was isolated from P. morifolia. The compound represents a rare example of natural isopropyl glycoside; its characterization included assignment of all 1H and 13C NMR signals of the primeverosyl group using two-dimensional NMR methods. Biosynthesis of the isopropyl moiety of the primeveroside is unclear, but the formation of alcohols corresponding to natural cyanohydrins may be a previously unrecognized extension of the cyanohydrin biosynthesis pathway in higher plants.

Through-flow of water in leaves of a submerged plant is influenced by the apical opening

Abstract. Submerged aquatic higher plants maintain acropetal water transport to the young leaves in active growth to satisfy their demand for nutrients and hormones derived from the roots. We here present the first measurements of hydraulic properties for a submerged plant, the monocotyledon Sparganium emersum Rehman. The hydraulic conductance per unit length, Kh, was measured in leaf segments without the leaf tip and shown to be greater in old, fully developed leaves (1.5 · 10−10 · m4 · MPa−1 · s−1) than in young leaves (1.0 · 10−10 · m4 · MPa−1 · s−1). In leaves with intact leaf tips, however, Kh was significantly greater in the youngest leaves, which suggests that the leaf tip with the hydathode influences resistance and thus flow. Microscopy confirmed that the hydathodal area, which is an apical opening, undergoes structural changes with leaf age; a matrix of microorganisms develops in the older leaves and probably restricts water flow by clogging the hydathodes. The leaf specific conductivity expressing transport capacity relative to the leaf area supplied, of S. emersum (0.1 · 10−8 to 9 · 10−8 · m2 MPa−1· s−1) was within the same range as for various species of terrestrial ferns, vines and trees. This finding does not support the traditional concept of functionally reduced vascular transport in