Integrating Spectroscopy with Potato Disease Management.

Spectral phenotyping is an efficient method for the nondestructive characterization of plant biochemical and physiological status. We examined the ability of a full range (350 to 2,500 nm) of foliar spectral data to (i) detect Potato virus Y (PVY) and physiological effects of the disease in visually asymptomatic leaves, (ii) classify different strains of PVY, and (iii) identify specific potato cultivars. Across cultivars, foliar spectral profiles of PVY-infected leaves were statistically different (F = 96.1, P ≤ 0.001) from noninfected leaves. Partial least-squares discriminate analysis (PLS-DA) accurately classified leaves as PVY infected (validation κ = 0.73) and the shortwave infrared spectral regions displayed the strongest correlations with infection status. Although spectral profiles of different PVY strains were statistically different (F = 6.4, P ≤ 0.001), PLS-DA did not classify different strains well (validation κ = 0.12). Spectroscopic retrievals revealed that PVY infection decreased photosynthetic capacity and increased leaf lignin content. Spectral profiles of potato cultivars also differed (F = 9.2, P ≤ 0.001); whereas average spectral classification was high (validation κ = 0.76), the accuracy of classification varied among cultivars. Our study expands the current knowledge base by (i) identifying disease presence before the onset of visual symptoms, (ii) providing specific biochemical and physiological responses to disease infection, and (iii) discriminating between multiple cultivars within a single plant species.

The effects of boiling and leaching on the content of potassium and other minerals in potatoes.

The white potato (Solanum tuberosum L.) is a valuable source of potassium in the human diet. While most consumers benefit from high levels of potassium in potato tubers, individuals with compromised kidney function must minimize their potassium intake. This study was undertaken to determine the effects of leaching and boiling on levels of potassium and other minerals in potato tubers. Leaching alone did not significantly reduce levels of potassium or other minerals in tubers. Boiling tuber cubes and shredded tubers decreased potassium levels by 50% and 75%, respectively. Reductions in mineral amounts following boiling were observed for phosphorus, magnesium, sulfur, zinc, manganese, and iron. There was no difference between the leaching and boiling treatment and the boiling treatment. In addition, mineral levels in tubers of 6 North American potato cultivars are reported. Significant differences in mineral levels were detected among cultivars, but they were too small to be nutritionally important. Individuals wishing to maximize the mineral nutrition benefits of consuming potatoes should boil them whole or bake, roast, or microwave them. Those who must reduce potassium uptake should boil small pieces before consuming them.

Enzymes that scavenge reactive oxygen species are down-regulated prior to gibberellic acid-induced programmed cell death in barley aleurone.

Gibberellins (GAs) initiate a series of events that culminate in programmed cell death, whereas abscisic acid (ABA) prevents this process. Reactive oxygen species (ROS) are key elements in aleurone programmed cell death. Incubation of barley (Hordeum vulgare) aleurone layers in H2O2 causes rapid death of all cells in GA- but not ABA-treated layers. Sensitivity to H2O2 in GA-treated aleurone cells results from a decreased ability to metabolize ROS. The amounts and activities of ROS scavenging enzymes, including catalase (CAT), ascorbate peroxidase, and superoxide dismutase are strongly down-regulated in aleurone layers treated with GA. CAT activity, protein, and Cat2 mRNA decline rapidly following exposure of aleurone layers to GA. In ABA-treated layers, on the other hand, the amount and activity of CAT and Cat2 mRNA increases. Incubation in ABA maintains high amounts of ascorbate peroxidase and superoxide dismutase, whereas GA brings about a rapid reduction in the amounts of these enzymes. These data imply that GA-treated cells loose their ability to scavenge ROS and that this loss ultimately results in oxidative damage and cell death. ABA-treated cells, on the other hand, maintain their ability to scavenge ROS and remain viable.

Cell death of barley aleurone protoplasts is mediated by reactive oxygen species.

The barley aleurone layer is a terminally differentiated secretory tissue whose activity is hormonally controlled. The plant hormone gibberellic acid (GA) stimulates the secretion of hydrolytic enzymes and triggers the onset of programmed cell death (PCD). Abscisic acid (ABA) antagonizes the effects of GA and inhibits enzyme secretion and PCD. Reactive oxygen species (ROS) are key players in many types of PCD, and data presented here implicate ROS in hormonally regulated death of barley aleurone cells. Incubation of aleurone layers or protoplasts in H(2)O(2)-containing media results in death of GA-treated but not ABA-treated aleurone cells. Cells that are programmed to die are therefore less able to withstand ROS than cells that are programmed to remain alive. Illumination of barley aleurone protoplasts with blue or UV-A light results in a rapid increase in intracellular H(2)O(2) production. GA-treated protoplasts die rapidly in response to this increase in intracellular H(2)O(2) production, but ABA-treated protoplasts do not die. The rate of light-induced death could be slowed by antioxidants, and incubating protoplasts in the dark with the antioxidant butylated hydroxy toluene reduces the rate of hormonally induced death. Taken together, these data demonstrate that GA-treated aleurone protoplasts are less able than ABA-treated protoplasts to tolerate internally generated or exogenously applied H(2)O(2), and strongly suggest that ROS are components of the hormonally regulated cell death pathway in barley aleurone cells.

Vacuoles and prevacuolar compartments.

Plant vacuoles are complex and dynamic organelles. Important advances have been made in our understanding of the transporters present in the tonoplast and of the molecular interactions that allow targeting to vacuoles. Despite these advances, markers that permit vacuoles to be defined unambiguously have not yet been identified.

Barley aleurone cell death is not apoptotic: characterization of nuclease activities and DNA degradation.

Barley aleurone cells undergo programmed cell death (PCD) when exposed to gibberellic acid (GA), but incubation in abscisic acid (ABA) prevent PCD. We tested the hypothesis that PCD in aleurone cells occurs by apoptosis, and show that the hallmark of apoptosis, namely DNA cleavage into 180 bp fragments, plasma membrane blebbing, and the formation of apoptotic bodies do not occur when aleurone cells die. We show that endogenous barley aleurone nucleases and nucleases present in enzymes used for protoplast preparation degrade aleurone DNA and that DNA degradation by these nucleases is rapid and can result in the formation of 180 bp DNA ladders. Methods are described that prevent DNA degradation during isolation from aleurone layers or protoplasts. Barley aleurone cells contain three nucleases whose activities are regulated by GA and ABA. CA induction and ABA repression of nuclease activities correlate with PCD in aleurone cells. Cells incubated in ABA remain alive and do not degrade their DNA, but living aleurone cells treated with GA accumulate nucleases and hydrolyze their nuclear DNA. We propose that barley nucleases play a role in DNA cleavage during aleurone PCD.

Gibberellin signaling.

Recent findings provide insights into the gibberellin signaling system in plants. Genes for gibberellin biosynthetic enzymes have been cloned, and an emerging theme is that gibberellin biosynthesis is negatively regulated by gibberellin responses. Mutants defective in gibberellin signaling have been analyzed, and an important finding is that gibberellin represses growth inhibition. The list of intracellular gibberellin signal-transduction elements has been expanded to include G-proteins and protein kinases.

Isolation of Intact Protein Storage Vacuoles from Barley Aleurone (Identification of Aspartic and Cysteine Proteases).

Within the cereal aleurone reserve proteins are stored in specialized organelles, the protein storage vacuoles (PSV). We developed an aqueous method for the isolation of intact PSV. Barley (Hordeum vulgare L. cv Himalaya) aleurone protoplasts were gently lysed by passing them through a syringe needle. PSV were separated from cytoplasmic components by microfiltration and low-speed centrifugation. Isolated PSV appeared by light microscopy to be identical with those within barley aleurone protoplasts. Luminal contents were retained throughout the isolation procedure. We used isolated PSV to identify and characterize PSV-associated proteolytic activities. Isolated PSV contained cysteine proteases and aspartic proteases (APs). Gibberellic acid treatment of protoplasts increased cysteine protease activity. Protein blots probed with anti-H. vulgare aspartic proteinase (HvAP) indicated that one PSV-AP was HvAP. Immunocytochemical localization by electron microscopy confirmed the presence of HvAP within the lumen of PSV. We conclude that isolated barley aleurone PSV will be useful in further characterizing this organelle.

Ca2+-Calmodulin Modulates Ion Channel Activity in Storage Protein Vacuoles of Barley Aleurone Cells.

Many plant ion channels have been identified, but little is known about how these transporters are regulated. We have investigated the regulation of a slow vacuolar (SV) ion channel in the tonoplast of barley aleurone storage protein vacuoles (SPV) using the patch-clamp technique. SPV were isolated from barley aleurone protoplasts incubated with CaCl2 in the presence or absence of gibberellic acid (GA) or abscisic acid (ABA). A slowly activating, voltage-dependent ion channel was identified in the SPV membrane. Mean channel conductance was 26 pS when 100 mM KCl was on both sides of the membrane, and reversal potential measurements indicated that most of the current was carried by K+. Treatment of protoplasts with GA3 increased whole-vacuole current density compared to SPV isolated from ABA- or CaCl2-treated cells. The opening of the SV channel was sensitive to cytosolic free Ca2+ concentration ([Ca2+]i) between 600 nM and 100 [mu]M, with higher [Ca2+]i resulting in a greater probability of channel opening. SV channel activity was reduced greater than 90% by the calmodulin (CaM) inhibitors W7 and trifluoperazine, suggesting that Ca2+ activates endogenous CaM tightly associated with the membrane. Exogenous CaM partially reversed the inhibitory effects of W7 on SV channel opening. CaM also sensitized the SV channel to Ca2+. In the presence of ~3.5 [mu]M CaM, specific current increased by approximately threefold at 2.5 [mu]M Ca2+ and by more than 13-fold at 10 [mu]M Ca2+. Since [Ca2+]i and the level of CaM increase in barley aleurone cells following exposure to GA, we suggest that Ca2+ and CaM act as signal transduction elements mediating hormone-induced changes in ion channel activity.

Stomatal and Nonstomatal Components to Inhibition of Photosynthesis in Leaves of Capsicum annuum during Progressive Exposure to NaCl Salinity.

Young bell pepper (Capsicum annuum L.) plants grown in nutrient solution were gradually acclimated to 50, 100, or 150 moles per cubic meter NaCl, and photosynthetic rates of individual attached leaves were measured on several occasions during the salinization period at external CO(2) concentrations ranging from approximately 70 to 1900 micromoles per mole air. Net CO(2) assimilation (A) was plotted against computed leaf internal CO(2) concentration (C(i)), and the initial slope of this A-C(i) curve was used as a measure of photosynthetic ability. During the 10 to 14 days after salinization began, leaves from plants exposed to 50 moles per cubic meter NaCl showed little change in photosynthetic ability, whereas those treated to 100 or 150 moles per cubic meter NaCl had up to 85% inhibition, with increase in CO(2) compensation point. Leaves appeared healthy, and leaf chlorophyll content showed only a 14% reduction at the highest salinity levels. Partial stomatal closure occurred with salinization, but reductions in photosynthesis were primarily nonstomatal in origin. Photosynthetic ability was inversely related to the concentration of either Na(+) or Cl(-) in the leaf laminas sampled at the end of the experimental period. However, the concentration of Cl(-) expressed on a tissue water basis was greater, exceeding 300 moles per cubic meter, and Cl(-) was more closely associated (R(2) = 0.926) with the inhibition of photosynthetic ability. Leaf turgor was not reduced by salinization and leaf osmotic potential decreased to a slightly greater extent than the osmotic potential decreases of the nutrient solutions. Concentration of accumulated Na(+) and Cl(-) (on a tissue water basis) accounted quantitatively for maintenance of leaf osmotic balance, assuming that these ions were sequestered in the vacuoles.