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PubMed1991

Cadmium uptake in Norway spruce (Picea abies (L.) Karst.) seedlings.

Root elongation of Norway spruce (Picea abies (L.) Karst.) seedlings was inhibited in the presence of 5 microM Cd, but was unaffected by a Cd concentration at 0.05 microM. Nutrient solutions labeled with (109)Cd were used to investigate the influence of pH, cations and the metabolic inhibitor 2,4-dinitrophenol (DNP) on the uptake of Cd by roots of intact spruce seedlings. Extracellular Cd was removed by washing the roots, and the relative amounts of Cd in the root apoplast and symplast estimated. In the presence of DNP, Cd uptake was reduced at 0.05 microM (non-toxic) but not at 5 microM Cd (toxic). At 0.05 microM Cd, the uptake of Cd into both the apoplast and symplast was dependent on the pH of the nutrient solution. Lower pH decreased Cd accumulation. Aluminum supplied at 100 or 500 microM lowered the Cd concentrations of both the apoplast and symplast. An increase in Ca or Mg supply reduced the Cd concentration of the apoplast but not of the symplast. In the presence of 5 microM Mn, the concentration of Cd in the symplast decreased by 44% compared to the control (1 microM Mn). High concentrations of Zn or Hg did not affect the Cd concentration of the roots.

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PubMed1991

Dynamics of swaying of Picea sitchensis.

Six 26-year-old Sitka spruce (Picea sitchensis (Bong.) Carr.) trees growing in a Scottish plantation were swayed manually to determine their mechanical dynamics. The natural frequency of sway of the intact trees (mean height 14.2 m and mean stem diameter 14.5 cm at 1.3 m) was on average 0.35 Hz. The variation of this frequency with tree size was found to be well described by engineering mechanics theory. In particular, shape parameters could be defined for both intact and branchless trees, which, along with stem size, density and elasticity, could predict the natural sway frequency using a simple formula. The damping of sway was found to consist of three components, (1) interference of branches with those of neighbors, (2) aerodynamic drag on foliage, and (3) damping in the stem. For the sample of six trees, which spanned the diameter range at the experimental site, the importance of these three components to overall damping was in the ratio 5/4/1 for the median sized tree. Interference between neighbors depended on the distance to neighbors, as well as on the sizes of the chosen tree and its neighbors. Aerodynamic damping was larger for larger trees and the energy lost to this force was similar in magnitude to that calculated to be lost using drag coefficients from published wind tunnel and other studies. The amount of damping from the stem was linearly related to stem diameter.

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PubMed1992

Purification of Two Superoxide Dismutase Isozymes and Their Subcellular Localization in Needles and Roots of Norway Spruce (Picea abies L.) Trees.

Two isozymes of superoxide dismutase (SOD; EC 1.15.1.1) were purified from Norway spruce (Picea abies L.) needles to apparent electrophoretic homogeneity. Purification factors were 354 for SOD I and 265 for SOD II. The native molecular mass of both purified enzymes was approximately 33 kD, as determined by gel filtration. The subunit molecular weights, as estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis, were 20,000 for SOD I and 16,000 for SOD II in the presence of 2-mercaptoethanol, and 15,800 and 15,000, respectively, in its absence. These results indicate that the native enzymes were homodimers whose subunits contained intrachain disulfide bonds. Isoelectric points determined by nondenaturing isoelectric focusing were 4.5 and 5.5 for SOD I and II, respectively. NH(2)-terminal sequence analysis of the first 22 to 23 amino acids revealed 70 to 75% sequence identity with chloroplastic CuZn SODs from other plant species for SOD I, and 75% sequence identity with the cytosolic CuZn SOD from Scots pine for SOD II. SOD I was the major activity in needles and it was associated with chloroplasts. SOD II activity was dominant in roots.

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PubMed1994

Apoplastic Peroxidases and Lignification in Needles of Norway Spruce (Picea abies L.).

The objective of the present study was to investigate the correlation of soluble apoplastic peroxidase activity with lignification in needles of field-grown Norway spruce (Picea abies L.) trees. Apoplastic peroxidases (EC 1.11.1.7) were obtained by vacuum infiltration of needles. The lignin content of isolated cell walls was determined by the acetyl bromide method. Accumulation of lignin and seasonal variations of apoplastic peroxidase activities were studied in the first year of needle development. The major phase of lignification started after bud break and was terminated about 4 weeks later. This phase correlated with a transient increase in apoplastic guaiacol and coniferyl alcohol peroxidase activity. NADH oxidase activity, which is thought to sustain peroxidase activity by production of H2O2, peaked sharply after bud break and decreased during the lignification period. Histochemical localization of peroxidase with guaiacol indicated that high activities were present in lignifying cell walls. In mature needles, lignin was localized in walls of most needle tissues including mesophyll cells, and corresponded to 80 to 130 [mu]mol lignin monomers/g needle dry weight. Isoelectric focusing of apoplastic washing fluids and activity staining with guaiacol showed the presence of strongly alkaline peroxidases (isoelectric point [greater than or equal to] 9) in all developmental stages investigated. New isozymes with isoelectric points of 7.1 and 8.1 appeared during the major phase of lignification. These isozymes disappeared after lignification was terminated. A strong increase in peroxidase activity in autumn was associated with the appearance of acidic peroxidases (isoelectric point [less than or equal to] 3). These results suggest that soluble alkaline apoplastic peroxidases participate in lignin formation. Soluble acidic apoplastic peroxidases were apparently unrelated to developmentally regulated lignification in spruce needles.

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PubMed1977

Effects of Quality, Intensity, and Duration of Light Breaks during a Long Night on Dormancy in Blue Spruce (Picea pungens Engelm.) Seedlings.

Blue spruce (Picea pungens Engelm.) seedlings grow continuously when exposed to photoperiods exceeding 16 hours and enter dormancy within 4 weeks under photoperiods of 12 hours or less. Dormancy was prevented under 12-hour photoperiods by 2-hour light breaks of red light (1.70 muw/cm(2) at 650 nm) or high intensity white light (2,164.29 muw/cm(2) at 400 to 800 nm) given in the middle of the 12-hour night, and by continuous low intensity white light (204.76 muw/cm(2) at 400 to 800 nm). Two-hour light breaks of far red light (1.80 muw/cm(2) at 730 nm), red light followed by far red light, or low intensity white light were not effective in delaying dormancy. The results imply that the phytochrome system mediates the photoperiodic control of dormancy in blue spruce seedlings. The similarity of results obtained using the low intensity, long duration as against the high intensity, short duration light treatments suggests that the law of reciprocity applies in this response.

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PubMed1978

Physiological Effects of Surface Waxes: I. Light Reflectance for Glaucous and Nonglaucous Picea pungens.

Foliage reflectance was studied on glaucous and nonglaucous foliage of blue spruce (Picea pungens Engel.). Current-year and 1-year-old glaucous and nonglaucous foliage from mature trees and seedling glaucous and nonglaucous foliage had similar reflectance patterns in the 350 to 800 nanometer region. The highest reflectance was in the 750 to 800 nanometer region and the lowest reflectance was in the 670 nanometer region. Glaucous foliage had a higher percentage of light reflectance at all of the wavelengths of light. The largest difference of reflectance between glaucous and nonglaucous foliage was in the 350 nanometer region with a general decline in the difference to the smallest difference at the 800 nanometer region.

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PubMed1983

Metabolism of Tritiated Gibberellins A(4) and A(9) in Norway Spruce, Picea abies (L.) Karst. : Effects of a Cultural Treatment Known to Enhance Flowering.

Shoots of mature grafted propagules of Picea abies (L.) Karst. metabolized [(3)H]gibberellin A(4) (GA(4)) to at least 14 acidic substances, two of which were tentatively identified by gas-liquid radiochromatography as GA(2) (possibly an artifact) and GA(34). [(3)H]GA(9) was converted into a number of metabolites, one of which was chromatographically similar to, but not identical with, GA(4). Metabolism was maximally 61 and 57% over 48 hours for GA(4) and GA(9), respectively, and was correlated with the rate of change (i.e. increase followed by decrease) in endogenous GA-like substances as shoot elongation progressed. Propagules covered with a clear plastic film, a treatment which promotes flowering, metabolized [(3)H]GA(4) more slowly than did control plants in the open. Inasmuch as a GA(4/7) mixture can also promote flowering in P. abies, the retarded metabolism of [(3)H]GA(4) may reflect the manner in which trees under plastic metabolize endogenous GA-like substances. If so, then the stimulating effect of this cultural treatment on flowering may come about through an increased level of endogenous, less polar GA-like substances.

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PubMed1984

Stilbene Synthase and Chalcone Synthase : Two Different Constitutive Enzymes in Cultured Cells of Picea excelsa.

Cultured cells of Picea excelsa capable of forming stilbenes and flavanoids have been established. Unlike needles of intact plants containing piceatannol (3,3',4',5-tetrahydroxystilbene) and stilbene glycosides the cultured cells converted phenylalanine and p-coumaric acid primarily into resveratrol monomethyl ether (3,4'-dihydroxy-5-methoxystilbene) and naringenin. Partially purified enzyme preparations were assayed for chalcone synthase as well as for stilbene synthase activity converting malonyl-CoA plus p-coumaroyl-CoA into 3,4',5-trihydroxystilbene (resveratrol).Although stilbene synthase and chalcone synthase use the same substrates and exhibit similar molecular properties, i.e. molecular weight and subunit molecular weight, they are two different proteins. This difference was demonstrated by gel electrophoresis and by means of monospecific antibodies.

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PubMed1985

Peroxidase Activity in Relation to Suberization and Respiration in White Spruce (Picea glauca [Moench] Voss) Seedling Roots.

Peroxidase (EC 1.11.1.7) activity is associated with suberization during endodermal development and metacutization in roots of white spruce (Picea glauca [Moench] Voss) seedlings. Histochemical analysis indicates a relationship between suberization and peroxidase activity, but peroxidase is ubiquitous. Increased peroxidase activity results from the induction of four anodic peroxidase isozymes in addition to quantitative increases in two anodic peroxidase isozymes. Four of these polymerized eugenol. Cold temperatures induce formation of two anodic isozymes and result in suberization. The increased peroxidase activity associated with suberization is correlated to residual respiration. In an attempt to elucidate this relationship, the effect of respiratory inhibitors on respiration and peroxidase activity are compared.

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PubMed1986

Root Respiration in White Spruce (Picea glauca [Moench] Voss) Seedlings in Relation to Morphology and Environment.

Roots of white spruce seedlings (Picea glauca [Moench] Voss) undergo respiratory changes during the year that are related to changing metabolic requirements. An alternative pathway is always present, functions during most of the year, and operates maximally during periods of root and shoot growth. Although some differences in respiration and the apportioning of respiration can be correlated to root morphology, the environment and the stage of shoot development are also important controls. Differences in respiration related to root morphology are not manifest in mitochondrial structure, but overall rates were found to correlate with the number of mitochondria present. Root respiration in seedlings grown under root growth capacity conditions reflects root and shoot growth at that time rather than root growth capacity.

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PubMed1987

Identification of Gibberellins in Norway Spruce (Picea abies [L.] Karst.) by Combined Gas Chromatography-Mass Spectrometry.

Gibberellins A(1) (GA(1)), A(3) and A(9) were identified from extracts of shoots of 6-month old Norway spruce (Picea abies) seedlings by the use of sequential reverse and normal phase high performance liquid chromatography (HPLC), bioassay, radioimmunoassay (RIA) and combined gas chromatography-mass spectrometry (GC-MS). The bioassay and RIA were used after fractionation by HPLC to detect the GA-containing fractions, which were then examined by GC-MS. The GAs identified are considered to be endogenous.

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PubMed1989

Assay and Electrophoresis of Superoxide Dismutase from Red Spruce (Picea rubens Sarg.), Loblolly Pine (Pinus taeda L.), and Scotch Pine (Pinus sylvestris L.) : A Method for Biomonitoring.

This paper reports a method for extracting the antioxidant enzyme superoxide dismutase (SOD) from the needles of red spruce (Picea rubens Sarg.), loblolly pine (Pinus taeda L.), and scotch pine (Pinus sylvestris L.) with high efficiency and free from interfering compounds. The extraction employs phosphate buffer with polyvinylpolypyrrolidone and Triton X-100 followed by dialysis overnight. The isozymes of SOD in each species were separated electrophoretically and tested for their sensitivity to KCN and H(2)O(2). An isozyme resistant to these inhibitors was found in the spruce but not the pine needles. The isozymes from the spruce needles were examined for individual responses to aging and H(2)O(2) inhibition. Four of the five CuZn isozymes in spruce were found to have increased significantly but equally by October of their first year and two of those four isozymes were found to be more sensitive to H(2)O(2). The response of the SOD isozymes in loblolly pine seedlings to O(3) was also examined and the isozymes were found to be induced equally. Because the SOD activity in the young pine needles was too low to electrophorese, the SOD activity from the pines in the O(3) experiment had to be partially purified using CHCl(3) and ethanol, then concentrated.

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PubMed1989

A N and N Nuclear Magnetic Resonance Study of Nitrogen Metabolism in Shoot-Forming Cultures of White Spruce (Picea glauca) Buds.

Nitrogen-14 and nitrogen-15 nuclear magnetic resonance (NMR) spectra were recorded for freshly dissected buds of Picea glauca and for buds grown for 3, 6 and 9 weeks on shoot-forming medium. Resonances for Glu (and other alphaNH(2) groups), Pro, Ala, and the side chain groups in Gln, Arg, Orn, and gamma-aminobutyric acid could be detected in in vivo(15)N NMR spectra. Peaks for alpha-amino groups, Pro, NO(3) (-) and NH(4) (+) could also be identified in (14)N NMR spectra. Perfusion experiments performed for up to 20 hours in the NMR spectrometer showed that (15)N-labeled NH(4) (+) and NO(3) (-) are first incorporated into the amide group of Gln and then in the alphaNH(2) pool. Subsequently, it also emerges in Ala and Arg. These data suggest that the glutamine synthetase/ glutamate synthase pathway functions under these conditions. The assimilation of NH(4) (+) is much faster than that of NO(3) (-). Consequently after 10 days of growth more than 70% of the newly synthesized internal free amino acid pool derives its nitrogen from NH(4) (+) rather than NO(3) (-). If NH(4) (+) is omitted from the medium, no NO(3) (-) is taken up during 9 weeks and the buds support limited growth by utilizing their endogenous amino acid pools. It is concluded that NH(4) (+) and NO(3) (-) are both required for the induction of nitrate- and nitrite reductase.

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PubMed1990

Changes in Protein Synthesis upon Cytokinin-Mediated Adventitious Bud Induction and during Seedling Development in Norway Spruce, Picea abies.

A pulse-treatment of embryos of Picea abies (L.) Karst with cytokinin efficiently and reproducibly induces the coordinate de novo formation of bud primordia from subepidermal cells. The cytokinin treatment also affects the germinative development of the embryo; chloroplast maturation is delayed, and cell elongation is completely suppressed. We have analyzed the protein patterns in developing spruce embryos with the aim of identifying proteins which are differentially synthesized during early bud-differentiation and germination. In addition to a set of major seed storage proteins and a large set of constitutively synthesized proteins, we distinguish two sets of proteins that showed different patterns of synthesis in relation to germination. One was synthesized at high rates during germination, and the second set during post-germinative seedling development. Twenty-two proteins were differentially synthesized in the bud-induced versus the germinating embryos. Interestingly, all 22 belonged to either the germination phase-abundant or the seedling protein sets, whereas the constitutively synthesized proteins were unaffected by the treatment. Proteins synthesized exclusively in bud-induced embryos were not found. In total, the bud-induction treatment caused a maintenance of a protein synthesis pattern typical for the germination phase in the nontreated embryos, and the de novo formation of buds was not preceded by a major change in gene expression in the tissue.

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PubMed1990

Quantitation of Gibberellins A(1), A(3), A(4), A(9) and a Putative A(9)-Conjugate in Grafts of Sitka Spruce (Picea sitchensis) during the Period of Shoot Elongation.

The levels of endogenous gibberellin A(1) (GA(1)), GA(3), GA(4), GA(9), and a cellulase hydrolyzable GA(9) conjugate in needles and shoot stems of mature grafts of Sitka spruce (Picea sitchensis [Bong.] Carr.) grown under environmental conditions that were either inductive, hot, and dry, or noninductive, cool, and wet, for flowering, were estimated by combined gas chromatography-mass spectrometry selected ion monitoring using deuterated [(2)H(2)]GA(1), GA(3), GA(4), and GA(9) as internal standards. The samples were taken when the shoots had elongated about 30, 70, and 95% of the final shoot length and 17 days after elongation had terminated. The concentration of putative GA(9)-conjugate, estimated by GCSIM of GA(9) after cellulase hydrolysis of the highly water soluble fraction, was 33 nanograms per gram fresh weight in the needles of both heat and drought- and cool and wet-treated plants sampled just after bud burst. The concentration gradually decreased to a final value of 13 nanograms per gram fresh weight in the heat and drought-treated grafts and 6 nanograms per gram fresh weight in the cool and wet-treated grafts. The stems contained no detectable putative GA(9) conjugate. Free GA(9) was highest in heat and drought-treated material. For plants subjected to this treatment, GA(9) increased from 22 to 32 nanograms per gram fresh weight in needles and from 1 to 22 nanograms per gram fresh weight in stems during the rapid stem elongation phase. By day 17, after cessation of shoot elongation, GA(9) had decreased to 12 nanograms per gram fresh weight in needles and 9 nanograms per gram fresh weight in the shoot stems. The cool and wet-treated material also showed an increase in GA(9) concentration during shoot elongation. However, the concentration was not as high and was also delayed compared with heat and drought-treated material. By day 17, after cessation of shoot elongation, GA(9) concentration was 9 nanograms per gram fresh weight in needles and 5 nanograms per gram fresh weight in stems for cool and wet treatment plants. The concentration of GA(4) was very low in tissue from both treatments. Fluctuation in concentration of the more polar gibberellins, GA(1) and GA(3), showed the same pattern as fluctuations in the content of GA(9). However, the heat and drought-treated material had lower amounts of GA(1) and GA(3) during the later phases of shoot elongation, than the cool and wet-treated material. These results imply differential metabolism between clones treated with conditions inductive and noninductive for flowering. Higher concentrations of putative GA(9) conjugate and free GA(9) in the hot and dry treatment indicate a higher capacity of synthesizing, for flowering, the physiologically important GA(4) in the heat and drought-treated material. This synthesis does not, however, result in a buildup of the GA(4) pool, probably because of a high turnover rate of GA(4). The cool and wet-treated material had higher amounts of GA(1) and GA(3), indicating that the differentiation was preferentially directed toward vegetative growth.

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PubMed1990

Composition and Properties of Hydrogen Peroxide Decomposing Systems in Extracellular and Total Extracts from Needles of Norway Spruce (Picea abies L., Karst.).

Hydrogen peroxide (H(2)O(2)) scavenging systems of spruce (Picea abies) needles were investigated in both extracts obtained from the extracellular space and extracts of total needles. As assessed by the lack of activity of symplastic marker enzymes, the extracellular washing fluid was free from intracellular contaminations. In the extracellular washing fluid ascorbate, glutathione, cysteine, and high specific activities of guaiacol peroxidases were observed. Guaiacol peroxidases in the extracellular washing fluid and needle homogenates had the same catalytic properties, i.e. temperature optimum at 50 degrees C, pH optimum in the range of pH 5 to 6 and low affinity for guaiacol (apparent K(m) = 40 millimolar) and H(2)O(2) (apparent K(m) = 1-3 millimolar). Needle homogenates contained ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase, glutathione reductase, and catalase, but not glutathione peroxidase activity. None of these activities was detected in the extracellular washing fluid. Ascorbate and glutathione related enzymes were freeze sensitive; ascorbate peroxidase was labile in the absence of ascorbate. The significance of extracellular antioxidants for the detoxification of injurious oxygen species is discussed.

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PubMed1992

Antioxidants and Manganese Deficiency in Needles of Norway Spruce (Picea abies L.) Trees.

Chlorotic and green needles from Norway spruce (Picea abies L.) trees were sampled in the Calcareous Bavarian Alps in winter. The needles were used for analysis of the mineral and pigment contents, the levels of antioxidants (ascorbate, glutathione), and the activities of protective enzymes (superoxide dismutase, catalase, ascorbate peroxidase, monodehydroascorbate radical reductase, dehydroascorbate reductase, glutathione reductase). In addition, the activities of two respiratory enzymes (glucose-6-phosphate dehydrogenase, NAD-malate dehydrogenase), which might provide the NADPH necessary for functioning of the antioxidative system, were determined. We found that chlorotic needles were severely manganese deficient (3 to 6 micrograms Mn per gram dry weight as compared with up to 190 micrograms Mn per gram dry weight in green needles) but had a similar dry weight to fresh weight ratio, had a similar protein content, and showed no evidence for enhanced lipid peroxidation as compared with green needles. In chlorotic needles, the level of total ascorbate and the activities of superoxide dismutase, monodehydroascorbate radical reductase, NAD-malate dehydrogenase, and glucose-6-phosphate dehydrogenase were significantly increased, whereas the levels of ascorbate peroxidase, dehydroascorbate reductase, glutathione reductase, and glutathione were not affected. The ratio of ascorbate to dehydroascorbate was similar in both green and chlorotic needles. These results suggest that in spruce needles monodehydroascorbate radical reductase is the key enzyme involved in maintaining ascorbate in its reduced state. The reductant necessary for this process may have been supplied at the expense of photosynthate.

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