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Antioxidative defense and photoprotection in Pinus halepensis induced by Mediterranean conditions and ozone exposure.

Pigment levels and antioxidative stress-related metabolites were determined over a period of two years in Pinus halepensis to characterize the response of some plant protective processes to both environmental stresses and 03 exposure. Two-year-old-Aleppo pine seedlings were grown in open-top chambers under optimal nutrient and water conditions and exposed to different 03 levels: charcoal filtered air, non-filtered air and nonfiltered air plus 40 ppb O3 10 h/day. In summer an activation of photoprotective systems was observed since a decrease in chlorophyll levels, an increase in the carotenoid/chlorophyll ratio and SOD activation were recorded. Interestingly, the SOD activity in Pinus halepensis was highly related to low-molecular weight compounds and this relationship increased with needle age. Ozone exposure induced alterations in the activity of some antioxidant enzymes along with reductions in pigment concentrations and an activation of the xanthophyll cycle.

Antioxidants↗

Paternally inherited chloroplast polymorphism in Pinus: estimation of diversity and population subdivision, and tests of disequilibrium with a maternally inherited mitochondrial polymorphism.

We have surveyed a chloroplast DNA restriction fragment length polymorphism in 745 individuals, distributed rangewide in eight allopatric natural populations of jack pine (Pinus banksiana Lamb.) and eight allopatric natural populations of lodgepole pine (Pinus contorta Dougl.). The polymorphic region of the chloroplast genome is located near duplicated psbA genes. Fourteen length variants were found in the survey, and these variants distinguished the two species qualitatively. Variant diversities were high in both species (hes = 0.43 in jack pine; hes = 0.44 in lodgepole pine). Population subdivision was weak within and among lodgepole pine subspecies and in jack pine (i.e., theta values were less than 0.05). This weak subdivision is compatible with theoretical predictions for paternally inherited markers in wind-pollinated outcrossers, as well as for polymorphisms with high length mutation rates. If these populations are at a drift-migration equilibrium, the chloroplast DNA restriction fragment data and previous mitochondrial frequency data from the same individuals are consistent with gene flow that is differential through seeds and pollen. The new data have permitted the first empirical tests of disequilibrium between maternally and paternally inherited factors. As expected, these tests failed to detect convincing evidence of nonrandom association between chloroplast and mitochondrial variants.

Chloroplasts↗

Growth, nutrition and response to water stress of Pinus pinaster inoculated with ten dikaryotic strains of Pisolithus sp.

Reconstituted dikaryons of Pisolithus sp. (Pers.) Coker & Couch from South Africa influenced growth parameters (shoot length, shoot/root ratio and leaf area), nutrition and physiological indicators (transpiration rate, stomatal conductance and xylem water potential) of maritime pine (Pinus pinaster Ait.) seedlings during drought and recovery from drought. Seedlings colonized with certain dikaryons were more sensitive to water stress and showed less mycorrhiza formation under water stress than seedlings colonized with other dikaryons. Control (uninoculated) seedlings were significantly smaller than those inoculated with dikaryons. Transpiration rate, stomatal conductance and xylem water potential varied among mycorrhizal treatments during the water stress and recovery periods. After rewatering, the controls and seedlings inoculated with dikaryon 34 x 20 had a weaker recovery in transpiration rate, stomatal conductance and xylem water potential than the other treatments and appeared to have experienced damage due to the water stress. Concentrations of various elements differed in the shoots of Pinus pinaster colonized by the various dikaryons. It is suggested that breeding of ectomycorrhizal fungi could constitute a new tool for improving reforestation success in arid and semi-arid zones.

Journal Article↗

Hydraulic conductance and soil water potential at the soil-root interface of Pinus pinaster seedlings inoculated with different dikaryons of Pisolithus sp.

Seedlings of maritime pine (Pinus pinaster Ait.) were inoculated with different dikaryons of Pisolithus sp. from South Africa to determine the influence of extension of the extramatrical phase and diameter of the mycelial strands on water relations parameters including xylem water potential (Psi(x)), soil water potential at the soil-root interface (Psi(s)) and hydraulic conductance (L(p)) during and after a period of water stress. Seedlings inoculated with dikaryons having an extensive extramatrical phase and large diameter mycelial strands showed higher Psi(s) (-2 MPa) during severe water stress than seedlings inoculated with dikaryons producing fine hyphae and sparse extramatrical phases (-3.8 MPa). Seedlings inoculated with strand-forming dikaryons recovered faster from water stress than did non-inoculated seedlings or seedlings inoculated with non-strand-forming dikaryons. Architectural aspects of the extramatrical phase, including the presence of large diameter mycelial strands or fine hyphae, influenced the soil-root contact and the water relations of an inoculated host plant. When water stress was not limiting, the architecture of the extramatrical phase did not have a large effect on Psi(s). It is suggested that the architecture of the extramatrical phase influences the resistance to water flow through the soil-root interface and that large mycelial strands increase the water flow by bridging the gap between the soil and the root. These changes in physiology indicate that dikaryons can improve the survival of Pinus pinaster under dry conditions.

Journal Article↗

High temperature and drought stress effects on survival of Pinus ponderosa seedlings.

We studied the effects of high temperature and drought on the survival, growth and water relations of seedlings of Pinus ponderosa (Dougl.) Lawson, one of few coniferous tree species that can successfully colonize drought-prone sites with high soil surface temperatures. Temperature profiles were measured with 0.07-mm thermocouples in a sparse ponderosa pine forest in northern Idaho. The soil surface and the adjacent 5 mm of air reached maximum temperatures exceeding 75 degrees C, well above the lethal temperature threshold for most plants. Air temperatures 50 mm above the soil surface (seedling needle height) rarely exceeded 45 degrees C. Pinus ponderosa seedlings that survived maintained basal stem temperatures as much as 15 degrees C lower than the surrounding air. The apparent threshold temperature at the seedling stem surface resulting in death was approximately 63 degrees C for less than 1 min. No correlation between seedling mortality and needle temperature was found, although some needles reached temperatures as high as 60 degrees C for periods of </= 1 min. Surviving seedlings had significantly higher stomatal conductance than seedlings that did not survive until fall. Transpiration rates, calculated from measured needle temperatures, stomatal conductance and evaporative demand, were high (up to 16.7 mmol m(-2) s(-1)), indicating that water transport through seedling stems may have acted as a heat transfer mechanism, cooling the stem below the lethal threshold temperature. Heat exchange calculations showed that rapid water flow through seedling stems can absorb sufficient energy to reduce stem temperature by 30 degrees C during peak sunlight hours.

Journal Article↗

Control of longitudinal and cambial growth by gibberellins and indole-3-acetic acid in current-year shoots of Pinus sylvestris.

We investigated the involvement of gibberellins (GAs) and indole-3-acetic acid (IAA) in the control of longitudinal and cambial growth in current-year shoots of Pinus sylvestris L. Elongating terminal shoots, located at the apex of previous-year (1-year-old) branches in the uppermost whorl on the main stem, were variously decapitated (apical 5 to 10 mm removed), defoliated (all developing needle fascicles removed) and treated with endogenous GA(4/7) or IAA, or both. Shoot length and the radial widths of xylem and phloem were measured, and the concentrations of GA(1), GA(3), GA(4), GA(9) and IAA in the stem were determined by combined gas chromatography-mass spectrometry with deuterated GAs and [(13)C(6)]-IAA as internal standards. Decapitation decreased the production of xylem and phloem and the IAA concentration, but did not alter either longitudinal growth or the concentrations of GAs. Defoliation markedly inhibited shoot elongation, as well as cambial growth, and reduced the concentrations of GA(1), GA(3), GA(4), GA(9) and IAA. Application of GA(4/7) to defoliated shoots promoted longitudinal growth and phloem production, without affecting xylem production or IAA concentration. Application of GA(4/7) and IAA together to decapitated + defoliated shoots increased shoot elongation, xylem and phloem production and IAA concentration, whereas applying either substance alone had a smaller effect or none at all. We conclude that, for elongating current-year shoots of Pinus sylvestris, (1) both the shoot apex and the developing needle fascicles are major sources of the IAA present in the stem, whereas stem GAs originate primarily in the needle fascicles, (2) GAs and IAA are required for both shoot elongation and cambial growth, and (3) GAs act directly in the control of shoot growth, rather than indirectly through affecting the IAA concentration.

Journal Article↗

Influence of rate of change in stomatal conductance to fluctuating irradiance on estimates of daily water use by Pinus taeda leaves.

Pinus taeda L. stomata respond slowly to changes in irradiance. Because incident irradiance on a leaf varies constantly, the rate of change in stomatal conductance to fluctuating irradiance may have a large effect on plant water use. We estimated total daily water use of Pinus taeda foliage for 10 days with very different irradiance patterns, assuming that rates of stomatal opening and closing were similar. To determine how the most extreme imbalance in rates of stomatal opening and closing affects estimates of water use, we also estimated total daily water use assuming instantaneous stomatal opening and a realistic rate of stomatal closing. Total daily water use was calculated by summing estimates of transpiration based on irradiance and vapor pressure deficit measured every minute at locations atop and within a canopy. Estimates of total daily water use calculated on the basis of realistic rates of stomatal opening and closing were similar to estimates calculated assuming instantaneous stomatal change (mean difference between methods of calculation was less than 0.2%). Estimates of total daily water use assuming instantaneous stomatal opening and a realistic rate of closing differed from estimates of total daily water use based on similar rates of stomatal opening and closing. The discrepancy was greater within the canopy (mean difference 6%) than at the top of the canopy (mean difference 1%). Calculation of mean daily conductance from mean daily irradiance, without accounting for minute-by-minute variations in irradiance, resulted in overestimations of daily stomatal conductance (13% mean error) and the magnitude of the error was directly related to the variation in irradiance for that day. We conclude that, provided variation in irradiance is accounted for, rates of stomatal opening and closing have little effect on estimates of daily water use.

Journal Article↗

Mineral Nutrient Requirements of a Loblolly Pine (Pinus taeda) Cell Suspension Culture : Evaluation of a Medium Formulated from Seed Composition Data.

The mineral nutrient requirements of Pinus taeda cells were explored using quantitative cell culture growth measurements. An appraisal was thereby made of the critical features of a novel and successful medium which was developed specifically for this gymnosperm using chemical composition data for developing seeds, and characterized by generally high concentration of all micronutrients, high magnesium, and low calcium. The high magnesium concentration was found not to be detrimental and possibly beneficial whereas the calcium level bordered on a deficiency threshold. Within the microelements high iodide was found to be essential, as was a higher borate level than is present in media developed for angiosperms. High zinc concentrations were also beneficial, with normal levels permitting slower but nevertheless healthy growth. An improved medium was thereby formulated which was stress-free and exhibited broader genotype specificity. This new formulation has proved very successful in maintaining long-term growth of highly uniform and apparently meristematic suspension cultures of Pinus radiata.

Journal Article↗

In vivo effects of octanoate and oleate on glucose-6-phosphate dehydrogenase during germination of Pinus pinea seeds.

During our investigation on the effect of some fatty acids on the germination of Pinus pinea seeds (Vincenzini et al., 1973), we noted a marked decrease of G6PDH activity when octanoate or oleate were added to the culture medium. In an attempt to provide biochemical information on the effect of free fatty acids on G6PDH activity, in vivo, during Pinus pinea seeds germination, we tested different concentrations of octanoate and oleate added to the culture medium. Moreover other enzymes of fundamental metabolic pathways were also considered, particularly 6PGDH, the second enzyme of HMS.

Caprylates↗

[Review on genetic diversity in Pinus].

The ways of probing genetic diversity of pines involve many aspects, such as morphology, chromosome, isozyme, DNA, etc. The phenotypic characteristics in pines vary widely and the differences of inheritability(h2) are obvious among characteristics and among species. Up to now,isozyme is still the most common means to measure genetic diversity of pines. Generally,there are high allozyme diversity within populations and low differentiation coefficient among populations, but differences exist between species in Pinus. The variations of chromosome among pines are very low and the karyotypes of pines are consentaneous,but the genomes of pines in cell nucleus are much larger than that of broadleaves. Diversity of pines are abundant at nucleus DNA level but are poor at plastid DNA level, such as ctDNA. There are many factors that will affect genetic diversity of pines, in which mating system and environment are two main factors. Finally,we reviewed the research on origin of Pinus and conservation strategy of genetic diversity,etc.

English Abstract↗

[Regularities of posthyperthermic recovery in haploid and diploid yeasts Saccharomyces cerevisiae and Pichia pinus].

Isogenic diploid cells of Saccharomyces cerevisiae are more sensitive to hyperthermic treatment (50 degrees C) than haploid ones, the posthyperthermical recovery efficiency being the same for both the cell types. In contrast to this, thermosensitivity of haploid and diploid cells of Pichia pinus does not practically differ, the posthyperthermic recovery efficiency for both the cell types being also the same. It is shown that diploid cells of P. pinus in the logarithmic phase of their growth are incapable of recovering after hyperthermal treatment, which is largely the reason for their higher sensitivity to such a treatment as compared with cells in the stationary phase of growth.

Diploidy↗

Occurrence of cyclic AMP and related enzymes during germination of Pinus pinea seeds.

The occurrence of cAMP, adenylate cyclase and cAMP phosphodiesterase has been tested in Pinus pinea seed during germination. The study has been carried out on dormant and imbibed seeds, seedlings, endospermic residues, roots and cotyledons. cAMP has been detected by the protein binding method and its occurrence has been verified by HPLC detections. cAMP phosphodiesterase shows a very high activity at acidic pH, while being completely inactive at pH 7.4. At this pH value, well detectable levels of adenylate cyclase have been observed. Therefore, the classical pathway of synthesis and breakdown of cAMP, already accepted for animal and bacterial cells, seems to be operating in Pinus pinea plant too.

3',5'-Cyclic-AMP Phosphodiesterases↗

Induction of antimicrobial activity by antitumor substances from pine cone extract of Pinus parviflora Sieb. et Zucc.

Pretreatment with two distinct antitumor substances extracted from pine cone of Pinus parviflora Sieb. et Zucc. protected mice from the lethal effects of E. coli infection. Intraperitoneal administration of these fractions transiently induced differentiation-inducing factor (DIF) with a peak at 1-2hr. The rapid decay of DIF activity from the peritoneal cavity was followed by polymorphonuclear cell (PMN) accumulation and enhancement of superoxide generation (assayed with luminol-dependent chemiluminescence (LDCL] by peritoneal exudate cells. The superoxide generation by adherent cells was similarly enhanced by pretreatment, but was only 10-20% of that of the peritoneal exudate cells. Fractions that showed comparable antitumor/antimicrobial activity were also obtained from seed shells and cones of other pine trees of Japanese and foreign origin. On the other hand, a neutral polysaccharide fraction from Pinus parviflora Sieb. et Zucc. that lacked any of these activities did not induce PMN accumulation, DIF activity or LDCL generation. The results suggest a significant role in PMN activation for the expression of antimicrobial activity induced by pine cone extracts.

Animals↗

[Comparative study of the spectra and activities of isoenzymes in haploid and autodiploid strains of Pichia pinus].

The spectra and activities of isozymes of hexokinase and alcohol dehydrogenase (enzymes that catalyse glucose and ethanol assimilation), glucose-6-phosphate dehydrogenase (the key enzyme of the pentose phosphate shunt) and malate dehydrogenase (an enzyme of the tricarboxylic acid cycle) were comparatively studied in Pichia pinus haploid (MH4) and autodiploid (D4) strains. Differences in the qualitative composition of the isoenzyme spectra and activities suggest that the intensity and the role of the studied metabolic pathways and cycles differ between the haploid and autodiploid strains of Pichia pinus.

Alcohol Dehydrogenase↗

[Lactate as competitive inhibitor of Pinus pinea isocitrate lyase].

We studied the effect of L-lactate on both the cleavage and the condensation reactions of Pinus pinea isocitrate lyase. This compound is a competitive of Pinus pinea isocitrate lyase towards both isocitrate and glyoxylate, whereas is a mixed type inhibitor towards succinate. Assuming that L-lactate acts as a glyoxylate analogue, our finding agrees with an uni-bi ordered mechanism of isocitrate lyase, with glyoxylate first substrate to enter the active site in the condensation reaction. Results are discussed and compared with those known in the literature about other structurally related metabolites.

Binding, Competitive↗

Abortifacient effects of lodgepole pine (Pinus contorta) and common juniper (Juniperus communis) on cattle.

Lodgepole pine (Pinus contorta) and common juniper (Juniperus communis) contain high levels of isocupressic acid that has been identified as the abortifacient component of ponderosa pine needles in cattle. Therefore, the abortifacient potential of P contorta and J communis needles was tested in feeding trials with pregnant cattle. Cows (2 groups of 2 each) were fed by gavage 4.5-5.5 kg/d ground dry needles from either P contorta or J communis starting on gestation day 250. Isocupressic acid (ICA) levels in P contorta needles and J communis plant material were 0.8 and 2.0% (dry weight) respectively. Cows fed P contorta received a daily dose of 62-78 mg ICA/kg body weight and aborted after 8 and 10 d. The 2 cows fed J communis received a daily dose of 190 and 245 mg ICA/kg body weight and aborted after 3 and 4 days respectively. All cows retained fetal membranes and had classical clinical signs of pine needle-induced abortion. Pinus ponderosa, P contorta, J communis, and Cupressus macrocarpa samples were also analyzed for the presence of myristate and laurate esters of 1,14-tetradecanediol and 1,12-dodecanediol. These lipid like compounds of P ponderosa have potent vasoconstrictive activity in a placentome perfusion assay and are proposed as possible abortifacients in cattle. Concentration of the vasoactive lipids were 0.028% (P ponderosa), 0.023% (P contorta), 0.001% (J communis), and none detected (C macrocarpa). It was concluded that these compounds are not required for the plant material to be abortifacient in cattle.

Abortifacient Agents↗

The role of the megagametophyte in maintaining loblolly pine (Pinus taeda L.) seedling arginase gene expression in vitro.

Following loblolly pine (Pinus taeda L.) seed germination, storage-protein breakdown in the megagametophyte and in the seedling results in a large increase in the seedling's free amino acid pool. A substantial portion of both the storage proteins and the amino acid pool is arginine, a very efficient nitrogen-storage compound. Free arginine is hydrolyzed in the seedling by the enzyme arginase (EC 3.5.3.1), which is under strong developmental control. At present, regulation of arginase in conifers is not well understood. Here we report the utilization of an in vitro culture system to address the separate impacts of the seedling and megagametophyte tissues on arginase enzyme activity, protein levels and patterns of gene expression. We also describe the generation of an anti-arginase antibody prepared from a histidine-tagged loblolly pine arginase fusion protein expressed in Escherichia coli. Our results indicate that arginase gene expression in the seedling is initiated by the seedling itself and then maintained or up-regulated by the megagametophyte. The contribution of storage-protein breakdown and the free amino acid pool, particularly arginine, in this regulation is also addressed.

Arginase↗

Regulation of amino acid uptake by carbon and nitrogen in Pinus sylvestris.

The simultaneous uptake of 13 different amino acids by Scots pine ( Pinus sylvestris L.) was characterized and its regulation investigated after pre-treatments with a range of C and N metabolites. The uptake system exhibited a broad substrate specificity, acquiring all tested amino acids at similar uptake rates. Uptake of all tested amino acids by excised roots was linear over a time period of 150 min and exhibited pH-dependency, showing a peak at pH 4.0-5.0. Uptake was increased following pre-treatments with glucose and sucrose, while ammonium pre-treatments had a negative effect on amino acid acquisition. Pre-treatments with the important Krebs cycle intermediates oxaloacetate or 2-oxoglutarate did not result in altered amino acid uptake. It is speculated that the up-regulation of uptake may be due to an increased flow of glucose through a sensor mechanism, such as hexokinase. The regulation of transport by N and C suggests a function for amino acid uptake in the N nutrition of the plant, thus further highlighting the importance of organic nitrogen for plant N nutrition suggested by an increasing amount of research.

Amino Acids↗