A protective role for the polyamine spermine against drought stress in Arabidopsis

Cellular polyamine content often changes in response to abiotic stresses. However, its physiological relevance is unknown. We found that an Arabidopsis mutant plant (acl5/spms), which cannot produce spermine, is hypersensitive to high salt. Examination of drought sensitivity of the mutant and comparison with wild type plants indicated hypersensitivity to drought. This phenotype was cured by spermine pretreatment but not by the other polyamines putrescine and spermidine, suggesting that drought-hypersensitivity exhibited by the mutant is due to spermine deficiency. The water loss rate of wild type and mutant plants were similar until 20 min after onset of dehydration stress, but after a longer exposure the rate in mutant plants was higher than in wild type plants. Consistent with this result, the stomata of the mutant leaves remained open while in wild type leaves they closed. Based on the collected data, we discuss a role for spermine in response to drought stress.     

Underexpression of the plant <Emphasis Type="Italic">NOTCHLESS</Emphasis> gene,encoding a WD-repeat protein,causes pleitropic phenotype during plant development

WD-repeat proteins are involved in a breadth of cellular processes. While the WD-repeat protein encoding gene NOTCHLESS has been involved in the regulation of the Notch signaling pathway in Drosophila, its yeast homolog Rsa4p was shown to participate in 60S ribosomal subunit biogenesis. The plant homolog ScNLE was previously characterized in Solanum chacoense (ScNLE) as being involved in seed development. However, expression data and reduced size of ScNLE underexpressing plants suggested in addition a role during shoot development. We here report the detailed phenotypic characterization of ScNLE underexpressing plants during shoot development. ScNLE was shown to be expressed in actively dividing cells of the shoot apex. Consistent with this, ScNLE underexpression caused pleiotropic defects such as a reduction in aerial organ size, a reduction in some organ numbers, delayed flowering, and an increase in stomatal index. Analysis of adaxial epidermal cells revealed that both cell number and cell size were reduced in mature leaves of ScNLE underexpressing lines. Two-hybrid screens with the Nle domain and the WD-repeat domain of ScNLE allowed the isolation of homologs of yeast MIDASIN and NSA2 genes, the products of which are involved in 60S ribosomal subunit biogenesis in yeast. A ScNLE-GFP chimeric protein was localized in both the cytoplasm and nucleus. These data altogether suggest that ScNLE likely plays a role in 60S ribosomal subunit biogenesis, which is essential for proper cellular growth and proliferation during plant development.

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A simple method for testing leaf responses of tall tropical forest trees to elevated CO2

The effects of atmospheric CO₂ enrichment on mature trees in their natural environment are largely unknown. Here we present a new, and inexpensive technique which can be used in situ to address some key physiological questions related to the CO₂ problem. Small, light-weight cups mounted on the lower side of rigid leaves at the top of tall tropical forest trees were supplied with CO₂-enriched air derived from a low-technology air mixing device utilizing forest floor CO₂ evolution. We present the scientific rationale for such field experiments, technical details, an assessment of potential cup artifacts and first results illustrating effects of elevated CO₂ on stomata and carbohydrate accumulation in the canopies of mature trees.     

Rose leaf structure in relation to different stages of micropropagation

Summary The Mme Isaac Pereire rose was investigated in an attempt to establish how micropropagated roses might best be weaned into normal growth conditions. Leaves of in vitro grown plants, weaned plants and the stock plant were studied, using light microscopy and different scanning and transmission electron microscopical techniques. Features that varied in the different growing conditions were leaf size and thickness, amount of wax, thickness of cuticle and external epidermal cell wall, number and aperture of the stomata, size of the epidermal cells, number of layers of the palisade cells, and size of the chloroplasts in the mesophyll. The rose in the present study had wax on the in vitro cultured plants; this wax was of similar ultrastructural appearance to that of the stock plant, even though in smaller quantities. Weaned plants had an intermediate amount of wax. The cuticle was thin, ranging from 0.04 m on plants growing in vitro to 0.3-0.6 m on weaned plants and stock plants. Stomata were always wide-open on leaves taken from cultures with a relative humidity of 100%. After four weeks in a humidity lowered to 85% stomata had closed.Abbreviations BAP 6-benzyl-aminopurine - CPD critical point drier - CTEM conventional transmission electron microscopy - NAA a-naphthaleneacetic acid - psi pounds per square inch - SEM scanning electron microscopy - TEM transmission electron microscopy     

Leaf chlorosis in oilseed rape plants (Brassica napus) grown on cadmium-polluted soil: causes and consequences for photosynthesis and growth

Brassica napus L. (oilseed rape) was grown from seeds on a reconstituted soil contaminated with cadmium (100 mg Cd kg⁻¹ dry soil), resulting in a marked chlorosis of the leaves which was investigated using a combination of biochemical, biophysical and physiological methods. Spectroscopic and chromatographic analyses of the photosynthetic pigments indicated that chlorosis was not due to a direct interaction of Cd with the chlorophyll biosynthesis pathway. In addition, mineral deficiency and oxidative stress were apparently not involved in the pigment loss. Leaf chlorosis was attributable to a marked decrease in the chloroplast density caused by a reduction in the number of chloroplasts per cell and a change in cell size, suggesting that Cd interfered with chloroplast replication and cell division. Relatively little Cd was found in the chloroplasts and the properties of the photosynthetic apparatus (electron transport, protein composition, chlorophyll antenna size, chloroplast ultrastructure) were not affected appreciably in plants grown on Cd-polluted soil. Depth profiling of photosynthetic pigments by phase-resolved photoacoustic spectroscopy revealed that the Cd-induced decrease in pigment content was very pronounced at the leaf surface (stomatal guard cells) compared to the leaf interior (mesophyll). This observation was consistent with light transmission and fluorescence microscopy analyses, which revealed that stomata density in the epidermis was noticeably reduced in Cd-exposed leaves. Concomitantly, the stomatal conductance estimated from gas-exchange measurements was strongly reduced with Cd. When plants were grown in a high-CO₂ atmosphere (4,000 μl CO₂ l⁻¹), the inhibitory effect of Cd on growth was not cancelled, suggesting that the reduced availability of CO₂ at the chloroplast level associated with the low stomatal conductance was not the main component of Cd toxicity in oilseed rape. Received: 14 July 2000 / Accepted: 27 August 2000     

Microtubules and morphogenesis in stomata of the water fernAzolla: An unusual mode of guard cell and pore development

Summary A mature stomate of the water fernAzolla consists of a single apparently unspecialized annular guard cell (GC) with two nuclei surrounding an elongated pore aligned longitudinally in the leaf. During development, the guard mother cell develops a preprophase band (PPB) of microtubules (MTs) oriented transverse to the leaf axis. This is followed by a cell plate which fuses with the parental walls at the PPB site. Subsequently only the central part of the cell plate is consolidated, while the parts to either side become perforated and tenuous and may disperse completely, forming a single composite GC.Meanwhile, a dense array of MTs appears along both faces of the central part of the new wall, oriented normal to the leaf surface. Further MT arrays radiate out across the periclinal walls from the region of the consolidated cell plate. Putative MT nucleating sites are seen along the cell edges between these anticlinal and periclinal arrays. Polarized light microscopy reveals cellulose deposition parallel to the periclinal MT arrays. At the same time lamellar material is deposited within the new anticlinal wall. As the GC complex elongates, a split appears in these lamellae creating an initially transverse slit which then opens up to become first circular and ultimately an elongated pore aligned in the long axis of the leaf,i.e., at right angles to the wall in which it originated. The radiating pattern of cellulose microfibrils in the periclinal walls contributes to the shaping of the pore. Elongation at the apical and basal ends of the GC is restricted by longitudinal microfibril orientation, while that at the sides is facilitated by transverse alignment.     

Levels of short-chain fatty acids and of abscisic acid in water-stressed and non-stressed leaves and their effects on stomata in epidermal strips and excised leaves

Straight-chain saturated fatty acids (C6-C11) and abscisic acid (ABA) accumulate in the leaves of Phaseolus vulgaris L. and Hordeum vulgare L. under water stress. ABA and certain of the fatty acids, particularly decanoic and undecanoic acid, can inhibit stomatal opening and cause stomatal closure in epidermal strips of Commelina communis L. depending on the incubating medium used. 10^-4 M (±)-ABA inhibits opening in media containing either high or relatively low concentrations of KCl but causes closure only in the latter medium. The fatty acids (at 10^-4 M) prevent opening in both media while significant closure of open stomata was caused only by undecanoic acid in both media and, additionally, by decanoic acid in the low-KCl medium. 10^-4 M formic acid also caused stomatal closure and prevented opening to significant extents in the low-KCl medium (it was not tested in the high-KCl medium). The efficacy of undecanoic acid in causing 50% inhibition of opening is about three orders of magnitude lower than that of ABA. At a concentration of 10^-3 M, nonanoic, decanoic and particularly undecanoic acid and all-trans-farnesol cause increased cell leakage in Beta vulgaris L. root tissue. Undecanoic acid (10^-4 M) also causes some loss of guard cell integrity in C. communis within 1.5 h of treatment. ABA (10^-4 M) reduces transpiration rates in barley and C. communis leaves when applied via the transpiration stream but decanoic and undecanoic acids did not have this effect. Transpiration was not affected when ABA or the fatty acids were applied to the leaf surfaces.Abbreviations ABA abscisic acid - RWC relative water content - SCFA short-chain fatty acids Deceased May 1977     

Secondary phenolic products in isolated guard cell,epidermal cell and mesophyll cell protoplasts from pea (Pisum sativum L.) leaves: Distribution and determination

Summary Guard cells and epidermal cells of the abaxial (lower) and adaxial (upper) epidermis ofPisum sativum L., mutant Argenteum, are the predominant sites of flavonoid accumulation within the leaf. This was demonstrated by the use of a new method of simultaneous isolation and separation of intact, highly-purified guard cell and epidermal cell protoplasts from both epidermal layers and of protoplasts from the mesophyll. Isolated guard and epidermal protoplasts retained flavonoid patterns of the parent epidermal tissue; quercetin 3-triglucoside and its p-coumaric acid ester as major constituents, kaempferol 3-triglucoside and its p-coumaric acid ester as minor compounds. Total flavonoid content in the lower epidermis was estimated to be ca. 80 fmol per guard cell protoplast and 500 fmol per epidermal cell protoplast. Protoplasts isolated from the upper epidermis had about 20–30% as much of these flavonoids. Mesophyll protoplasts retained only about 25 fmol total flavonoid per protoplast.By fluorescence microscopy, using the alkaline-induced yellow-green fluorescence characteristics of flavonols, we suggest that these flavonol glycosides are present in cell vacuoles. There was no indication for the presence of flavine-like compounds.Abbreviations uE adaxial (upper) epidermis - IE abaxial (lower) epidermis - GCP guard cell protoplasts - ECP epidermal cell protoplasts - MCP mesophyll cell protoplasts - PP protoplasts - HPLC high performance liquid chromatography - TLC thin layer chromatography - CC column chromatography - HOAc acetic acid     

Direct and indirect effects of light on stomata. I. In Scots pine and Sitka spruce

Abstract. The response of stomatal conductance to broadband blue and red light was measured in whole shoots of Scots pine and Sitka spruce, two species which have low stomatal sensitivity to CO². In Scots pine, blue light was more than three times more effective than red light (on an incident quantum basis) in opening stomata, particularly at low quantum flux densities (<100μmiol m⁻² s⁻¹). However, the apparent quantum yield of net CO² assimilation rate in blue light was only half that in red light. The contrasting effects of red and blue light on conductance and assimilation led to higher intercellular CO² concentrations (C_i) in blue light (up to 100 μmol mol⁻¹ higher) than in red light. Similar results were obtained with Sitka spruce shoots, though differences in the effectiveness of red and blue light were less marked. In both species, both red and blue light increased conductance in normal and CO²-free air, indicating that neither red nor blue light exert effects through changes in C_i or mesophyll assimilation. However, decreases in C_i caused increases in conductance in both red and blue light, suggesting that these direct effects of light are not wholly independent of CO².