J.A. Heyes

Role of a Single Aquaporin Isoform in Root Water Uptake

Aquaporins are ubiquitous channel proteins that facilitate the transport of water across cell membranes. Aquaporins show a typically high isoform multiplicity in plants, with 35 homologs in Arabidopsis. The integrated function of plant aquaporins and the function of each individual isoform remain poorly understood. Matrix-assisted laser desorption/ionization time-of-flight analyses suggested that Plasma Membrane Intrinsic Protein2;2 (PIP2;2) is one of the abundantly expressed aquaporin isoforms in Arabidopsis root plasma membranes. Two independent Arabidopsis knockout mutants of PIP2;2 were isolated using a PCR-based strategy from a library of plant lines mutagenized by the insertion of Agrobacterium tumefaciens T-DNA. Expression in transgenic Arabidopsis of a PIP2;2 promoter–β-glucuronidase gene fusion indicated that PIP2;2 is expressed predominantly in roots, with a strong expression in the cortex, endodermis, and stele. The hydraulic conductivity of root cortex cells, as measured with a cell pressure probe, was reduced by 25 to 30% in the two allelic PIP2;2 mutants compared with the wild type. In addition, free exudation measurements revealed a 14% decrease, with respect to wild-type values, in the osmotic hydraulic conductivity of roots excised from the two PIP2;2 mutants. Together, our data provide evidence for the contribution of a single aquaporin gene to root water uptake and identify PIP2;2 as an aquaporin specialized in osmotic fluid transport. PIP2;2 has a close homolog, PIP2;3, showing 96.8% amino acid identity. The phenotype of PIP2;2 mutants demonstrates that, despite their high homology and isoform multiplicity, plant aquaporins have evolved with nonredundant functions.

Molecular physiology of aquaporins in plants

In plants, membrane channels of the major intrinsic protein (MIP) super-family exhibit a high diversity with, for instance, 35 homologues in the model species Arabidopsis thaliana. As has been found in other organisms, plant MIPs function as membrane channels permeable to water (aquaporins) and in some cases to small nonelectrolytes. The aim of the present article is to integrate into plant physiology what has been recently learned about the molecular and functional properties of aquaporins in plants. Exhaustive compilation of data in the literature shows that the numerous aquaporin isoforms of plants have specific expression patterns throughout plant development and in response to environmental stimuli. The diversity of aquaporin homologues in plants can also be explained in part by their presence in multiple subcellular compartments. In recent years, there have been numerous reports that describe the activity of water channels in purified membrane vesicles, in isolated organelles or protoplasts, and in intact plant cells or even tissues. Altogether, these data suggest that the transport of water and solutes across plant membranes concerns many facets of plant physiology. Because of the high degree of compartmentation of plant cells, aquaporins may play a critical role in cell osmoregulation. Water uptake in roots represents a typical process in which to investigate the role of aquaporins in transcellular water transport, and the mechanisms and regulations involved are discussed.

Physiological changes associated with Sandersonia aurantiaca flower senescence in response to sugar

Abstract Sandersonia aurantiaca is a liliaceous cut flower in which senescence is not regulated by ethylene. We stood flower stems in solutions of deionized water (control) or sucrose (2%) and monitored the pattern of senescence of individual flowers attached to the stems and quantified the amount of carbohydrate and protein present in the flowers. Treatment with sucrose extended the postharvest life of flower stems by delaying the senescence of individual flowers attached to the flowering stem. Flowers on sucrose-treated stems were larger, firmer and brighter orange than the control flowers. Flowers that were treated with sucrose also contained greater quantities of carotenoids, soluble and storage carbohydrates and soluble protein than the control flowers. The first visible signs of senescence occurred prior to any net loss of soluble carbohydrate or protein.

Lactic Acid Bacteria Convert Glucosinolates to Nitriles Efficiently Yet Differently from Enterobacteriaceae

Glucosinolates from the genus Brassica can be converted into bioactive compounds known to induce phase II enzymes, which may decrease the risk of cancers. Conversion via hydrolysis is usually by the brassica enzyme myrosinase, which can be inactivated by cooking or storage. We examined the potential of three beneficial bacteria, Lactobacillus plantarum KW30, Lactococcus lactis subsp. lactis KF147, and Escherichia coli Nissle 1917, and known myrosinase-producer Enterobacter cloacae to catalyze the conversion of glucosinolates in broccoli extract. Enterobacteriaceae consumed on average 65% glucoiberin and 78% glucoraphanin, transforming them into glucoiberverin and glucoerucin, respectively, and small amounts of iberverin nitrile and erucin nitrile. The lactic acid bacteria did not accumulate reduced glucosinolates, consuming all at 30-33% and transforming these into iberverin nitrile, erucin nitrile, sulforaphane nitrile, and further unidentified metabolites. Adding beneficial bacteria to a glucosinolate-rich diet may increase glucosinolate transformation, thereby increasing host exposure to bioactives.

Textural and physiological changes during pepino (Solanum muricatum Ait.) ripening

Abstract Pepino fruit (cultivar ‘El Camino’) softened throughout ripening while attached to the bush. An Instron Universal Testing Machine was used to define a range of compressive and tensile textural parameters. A novel tensile test for fruit tissues is described. The pattern of textural change implied that softening was primarily a result of a progressive decline in cell wall strength and loss of cell-to-cell adhesion. In vitro treatments to lower tissue turgor, apoplastic pH, or free calcium were equally effective in softening tissue rings. Pectinmethylesterase and polygalacturonase (PG) activities increased during ripening. PG activity appeared just before the first externally visible sign of ripening (purple striping), and increased during the later stages of fruit softening. Pepino fruit are non-climacteric, since propylene accelerated colour change and fruit softening, but fruit produced little ethylene at harvest and did not produce ethylene autocatalytically in response to exogenous propylene. Fruit respiration increased transiently in response to propylene, but in control fruit the respiration rate increased only slowly during post-harvest ripening. During prolonged storage at 20°C the only fruit to produce ethylene at rates over 0.2 μl kg −1 h −1 were those which subsequently developed rots.

Vase solutions containing sucrose result in changes to cell walls of sandersonia (Sandersonia aurantiaca) flowers

It is well established that vase solutions containing sugar can improve the vase-life of many cut flower crops. Since cut sandersonia flowers supplied with 2% sucrose are firmer during wilting compared to water-fed controls, we have examined whether the effects of sucrose treatment extend to alterations in cell wall structure in the floral tissues, which may influence the wilting-related flower softening. Mature but not fully opened individual flowers were removed from the stems of sandersonia plants and were fed continuously with either 2% sucrose solution or water for up to 10 days. Sucrose supplementation resulted in decreased amounts of chelator-soluble pectin and increased amounts of Na2CO3-soluble pectin per individual flower, and also changed the molecular size profiles of both these pectin fractions compared to the water-fed controls. The molecular size differences were obvious after 3 days in vase solutions, and diminished with subsequent vase time. Senescence-related galactose loss was delayed in sucrose-fed flowers but there was no difference in the levels of β-galactosidase activity present in these flowers compared to controls. The observed differences in cell wall pectins due to sucrose feeding were not reflected in differences to the overall firmness of pre-senescent flowers (up to day 3). High levels of galactose persisted into the wilting phase when sucrose-fed flowers were firmer than water-fed controls. We conclude that while sucrose induced significant quantitative and qualitative differences in pectin fractions and galactose content, firmness of floral tissue, particularly during senescence, was not governed by these events alone.

1‐methylcyclopropene extends Cymbidium orchid vaselife and prevents damaged pollinia from accelerating senescence

Abstract A single postharvest application of 1‐methylcyclopropene (MCP) was effective in prolonging vaselife of Cymbidium orchids by 6 or 7 days, to c. 19 days. It also prevented repeated applications of ethylene shortening the vaselife of Cymbidium orchids. The anti‐ethylene effect of one MCP treatment persisted throughout three successive challenges with ethylene at 5‐day intervals. A supra‐optimal concentration of MCP (500 ppb) showed no adverse effects. MCP was also effective in protecting Cymbidium orchids against accelerated senescence caused by damage to the pollinia. Individual flowers with damaged pollinia senesced even more rapidly than ethylene‐treated intact flowers. In the presence of MCP, flowers with damaged pollinia had a vaselife almost equal to that of undamaged flowers. This is a significant finding for the orchid export industry.

Developmental and Ripening-Related Effects on the Cell Wall of Pepino (Solanum muricatum) Fruit

Several cell wall components in ripening pepino fruit have been quantitatively and qualitatively characterised, with the aim of identifying their contributions to the loss of tissue firmness. Pepinos were graded into nine groups based on progressive, characteristic skin colour changes, previously shown to correspond with decreasing fruit firmness. While fruit softening began when the pepinos were still green but with newly acquired purple stripes, the first significant quantitative signs of cell wall modification (total pectin and hemicellulose content declining and CDTA-soluble pectin content increasing, on a fresh weight basis) were detectable later in ripening, when the fruit began to acquire yellow skin pigmentation. Gel fractionation studies demonstrated that there were increased levels of low-molecular-weight pectin and xyloglucan during pepino ripening. The change in molecular weight distribution of CDTA-soluble pectin occurred as fruit started to acquire yellow pigmentation, while xyloglucan polymers were modified at an earlier stage that coincided with the initial loss of firmness.

Textural changes during nectarine (Prunus persica) development and ripening

Abstract Changes in nectarine (Prunus persica (L.) Batsch var nectarina (Ait.) Maxim. cv. ‘Fantasia’) texture were measured during the last 10 weeks of fruit development before commercial maturity, and during ripening. Both tensile and compressive textural characteristics of mesocarp tissue were measured using an Instron Universal Testing Machine. Developmental fruit softening was primarily a consequence of cell expansion. On three occasions during the developmental series we compared changes in texture during 3–5 days of postharvest storage at 20 °C with textural change over the same period on the tree. In the first two periods the fruit were not physiologically mature and harvested fruit did not undergo normal ripening. The distinctive patterns of change in tensile and compressive properties at these times were consistent with changes due to low turgor, as predicted by cellular theories. Harvested fruit from the third period (physiologically mature fruit producing ethylene at around 13 nmol kg−1 h−1) showed an accelerated rate of ripening and softening compared with fruit left on the tree. Reductions in cell wall strength and cell-to-cell adhesion would explain the observed patterns of change in tensile and compressive properties of mature fruit. Scanning electron microscopy confirmed there was a change in the predominant cause of tensile failure from cell rupture in immature fruit to cell separation in mature fruit. A tensile parameter which correlated with this change is described.

ATPase activity of mesocarp membranes during postharvest ripening of nectarines

Abstract Softening during fruit ripening may be initiated by an increase in proton pumping across the plasma membrane. This would lower the cell wall pH and loosen acid-labile bonds, as well as displacing Ca2+ from load-bearing bonds. The enzyme responsible for proton pumping is the plasma membrane H+-ATPase. The activity of this enzyme in nectarine (Prunus persica (L.) Batsch var. nectarina (Ait.) Maxim., cv. Redgold) mesocarp was studied during 4 days of ripening at 20°C after harvest at commercial maturity. Fruit began to produce significant amounts of ethylene (> 1 µlitre/kg per h) from the second day. Flesh firmness began to decline from the third day, but background skin colour did not change until the fourth day. Plasma membranes were extracted from the fruit by phase partitioning. Initial characterisation of the H+-ATPase activity showed that it was Mg2+-dependent, K+-stimulated, and VO4 3−-sensitive. The Km for MgA TP was 0.19 ± 0.02 mM. The Ki value for Ca2+ was estimated at 100 µM free Ca2+. Th...