Carbon metabolism in the subantarctic Kerguelen cabbage Pringlea antiscorbutica R. Br.: environmental controls over carbohydrates and proline contents and relation to phenology

The subantarctic Brassicaceae Pringlea antiscorbutica R. Br. (Kerguelen cabbage) was used as a model to study the physiological adaptations of higher plants to the subantarctic environment. ¹³C-nuclear magnetic resonance permitted, in combination with biochemical methods, the identification and quantification of the major solutes in leaves, stem and roots. As characterized in many Brassicaceae, proline was a major solute in all organs of the plants, and its accumulation was mainly controlled by salt stress rather than temperature. Glucose was the major soluble sugar in the leaves, whilst sucrose and starch accumulated in stems and roots. Over a period of 1 year we found strong correlations between (i) glucose content in leaves and irradiance, and (ii) starch content in non-photosynthetic organs and air temperature. The pattern of carbohydrate accumulation indirectly indicated that photosynthetis was sustained throughout the year, even during cold days when the temperature remained near 0 °C. This is consistent with the direct gas exchange measurements showing that photosynthetic capacity is mainly influenced by irradiance and weakly by temperature. Taken together, these characteristics demonstrated that the growth and development cycle occurs without a period of dormancy.     

Accumulation of organic and inorganic solutes in the subantarctic cruciferous species Pringlea antiscorbutica in response to saline and cold stresses

Pringlea antiscorbutica R. Br., a subantarctic endemic cruciferous species, is endangered in its natural sites by several ecological changes. This species is tolerant to salinity and a permanent cold temperature on Kerguelen and Crozet Islands. We attempted the investigation of regulating mechanisms of osmotic adjustment in this species. ¹³C NMR analyses of water-soluble compounds from leaves collected from the field revealed glucose and proline to be the main accumulated organic solutes. Colorimetric determinations in these samples showed that proline and soluble carbohydrates were present at remarkably high levels. When young plants were cultivated in growth chambers they showed a good resistance to cold and medium resistance to saline conditions. High levels of soluble carbohydrates were present in all situations. Proline was accumulated in response to a saline and a cold treatment. The quantitative variations of the pool of proline in response to saline treatments were rapid and important. The adaptive value of these responses of organic solutes in the tolerance of Pringlea antiscorbutica to various stresses is discussed. Received: 16 June 1997 / Accepted: 5 May 1998     

Amines of the subantarctic crucifer Pringlea antiscorbutica are responsive to temperature conditions

Different polyamine and aromatic amine compositions and contents were observed in the leaves and the roots of plants from the subantarctic crucifer Pringlea antiscorbutica growing in the field and collected during the austral summer in Kerguelen and those grown under controlled conditions mimicking the thermoperiod and photoperiod conditions in summer in Kerguelen. In controlled conditions, the plants grew more slowly than in the field and did not flower. In roots, this was associated with an increase of agmatine (Agm). In contrast, acetylated putrescine (Put) disappeared while dopamine (Dop) and tryptamine (Try) were strongly reduced. In leaves, cultivation under controlled conditions led to an accumulation of Agm, acetylated Put, tyramine (Tyr) and Try. A complete depletion of acetylated spermidine (Spd) and spermine (Spm) and a strong decrease of Dop occurred. Cultivation of plants at constant 25°C was lethal after a few weeks. Before the external symptoms of heat challenge became acute, leaves and roots showed a different amine content and composition. In roots, and to a lesser extent in leaves, heat treatment was associated with an apparent impairment in the ability to accumulate Agm, acetylated Put and Try. Strong increases in Put, Spd and Tyr and accumulation of hydroxycinnamoyl amines as feruloylputrescine, feruloylspermidine and feruloyltyramine were observed in leaves. Aminoguanidine, a potent and specific inhibitor of diamine oxidase activities, caused phenotypic alterations and changes in amine composition and content in roots and leaves of Pringlea plants similar to those observed under the heat treatment. Our results highlight possible roles of amine catabolism, acetylated polyamines and hydroxycinnamoyl amines in plant responses to external conditions.     

Temperature-dependent changes of amine levels during early seedling development of the cold-adapted subantarctic crucifer Pringlea antiscorbutica

Pringlea antiscorbutica R. Br., an endemic crucifer from the Kerguelen Archipelago in the subantarctic, has been previously shown to be unable to acclimatize to 25°C when transferred after several months cultivation under cold conditions. Furthermore, the polyamine composition was greatly modified in such high-temperature-treated plants. The development of seedlings of this species was investigated under a regime mimicking the subantarctic summer thermoperiod (5/10°C night/day) and a regime with high temperatures (22/25°C night/day). In parallel, the associated changes in polyamine composition that occurred during the first 6 days of seedling life were determined. Marked acceleration of seedling growth and intense cotyledon greening were observed at day 4 in 5/10°C-grown seedlings but not in 22/25°C-grown seedlings. Seedlings grown at high temperature accumulated agmatine and putrescine, whereas cold-cultivated seedlings maintained high levels of spermidine. Cold-cultivated seedlings accumulated the uncommon long-chain polyamines norspermidine and homospermidine. These seedlings also accumulated free 1,3-diaminopropane, cadaverine, N¹-acetylspermidine, N¹-acetylspermine and bound polyamines, whereas seedlings under high temperature accumulated N¹-acetylputrescine. Aromatic amine metabolism also appeared to be very responsive to temperature: seedlings under a cold regime accumulated free dopamine and bound phenylethylamine and tyramine, whereas seedlings grown at high temperature accumulated free tyramine. The possible relationships between the observed amine patterns and seedling growth under low and high temperature are discussed.     

Phylogeny and colonization history of Pringlea antiscorbutica (Brassicaceae), an emblematic endemic from the South Indian Ocean Province

The origins and evolution of sub-Antarctic island floras are not well understood. In particular there is uncertainty about the ages of the contemporary floras and the ultimate origins of the lineages they contain. Pringlea R. Br. (Brassicaceae) is a monotypic genus endemic to four sub-Antarctic island groups in the southern Indian Ocean. Here we used sequences from both the chloroplast and nuclear genomes to examine the phylogenetic position of this enigmatic genus. Our analyses confirm that Pringlea falls within the tribe Thelypodieae and provide a preliminary view of its relationships within the group. Divergence time estimates and ancestral area reconstructions imply Pringlea diverged from a South American ancestor ～5Myr ago. It remains unclear whether the ancestor of Pringlea dispersed directly to the South Indian Ocean Province (SIOP) or used Antarctica as a stepping-stone; what is clear, however, is that following arrival in the SIOP several additional long-distance dispersal events must be inferred to explain the current distribution of this species. Our analyses also suggest that although Pringlea is likely to have inherited cold tolerance from its closest relatives, the distinctive morphology of this species evolved only after it split from the South American lineage. More generally, our results lend support to the hypothesis that angiosperms persisted on the sub-Antarctic islands throughout the Pliocene and Pleistocene. Taken together with evidence from other sub-Antarctic island plant groups, they suggest the extant flora of sub-Antarctic is likely to have been assembled over a broad time period and from lineages with distinctive biogeographic histories.     

Carbon balance of a whole tomato plant and the contribution of source leaves to sink growth using the 14CO2 steady-state feeding method

To clarify the entire carbon balance of a young tomato plant and the contribution of each leaf to sink growth, the carbon balance of each leaf was quantitatively measured using the 14CO₂ steady-state feeding method to quantitatively measure the photosynthesis, translocation, distribution and respiration of newly fixed 14C. The entire carbon balance of the whole plant was calculated by adding the data of each leaf. The total amount of carbon fixed by all source leaves was 70.2 mg. Of this amount, in a 24-h period, 29% was accumulated in the sinks, 28% remained in the source leaves and 42% was respired. Of the total amount of carbon accumulated in the sinks, the proportion accumulated in the shoot apex, stem and roots were 27, 40 and 29%, respectively. The third and fourth leaves contributed about 30–40% of the total growth of the main sinks. The distribution pattern of each leaf to the shoot apex of the plant was greatest in the first leaf and decreased with decreasing leaf age, whereas it showed an opposite trend in the roots.     

Growth and gibberellin relations of the extreme dwarf dx tomato mutant

The extreme dwarf d ^x tomato ( Lycopersicon esculentum Mill.) mutant has very short internodes which were found to contain shorter and fewer epidermal cells. The leaves are highly abnormal. The mutant showed a substantial stem growth response to GA₃, without approaching normal stature or morphology. The active gibberellin GA₁ and its precursors GA₁₉ and GA₂₀ were identified by coupled gas chromatography-mass spectrometry (GC/MS) in d ^x shoots. Quantitative GC/MS revealed that GA₂₀ accumulated to far higher levels than normal in stems and leaves of the mutant.     

Physiological adaptation to subantarctic climate by the Kerguelen cabbage,Pringlea antiscorbutica R. Br.

Summary The Kerguelen cabbage, Pringlea antiscorbutica, is an endemic species restricted to some Subantarctic Islands, Up to now, all long term acclimatation assays of Kerguelen cabbage in the temperate zones remained unsuccessful. In the field, the Pringlea leaf relative water content (RWC) never decreased below 83%. At the same time the leaf diffusion resistance (LDR) remained low since the water flux was not limited in the plant as in these islands the soil water content is permanently high. Severe water deprivation was necessary to induce stomatal closure with excised leaves. In parallel in vitro experiments, irreversible damage to photosynthesis and respiration were observed in leaf slices under osmotic stress. These results sustain the hypothesis that Kerguelen cabbage can not support water deprivation and is probably specifically adapted to the subantarctic climate.Abbreviations DMF N, N-Dimethylformamide - LDR leaf diffusion resistance - PPFD photosynthetic photon flux density - RWC relative water content     

Effect of plant density on the uptake and distribution of 14C in the field bean, Vicia faba

Whole bean plants, ev. Cockfield, grown in pots crowded or well-spaced (50 or 10 plants m², respectively) were treated with ¹⁴CO₂ at the pod-fill stage (25 modes) and the radioactivity in each leaf was determined after 30 min. With spaced plants the uptake was greatest in the mid-stem leaves and was proportional to leaf area. In contrast, 70% of the total assimilation took place in the upper six leaves of crowded plants and there was a steady decrease down the stem.
When ¹⁴CO₂ was fed to single leaves of similar crowded plants the resultant distribution of labelled assimilates varied with the position of the treated leaf. After 6 h, 67% of the ¹⁴C fixed by a mid-stem leaf (node 13) was recovered from the beans, whereas 76% of that from an upper leaf (node 23) had accumulated along the stem. Due to the shading of mid-stem leaves at the higher planting densities, seed yield becomes increasingly dependent upon re-distribution of assimilates from stem to beans.     

Sources of proline-nitrogen in water-stressed soybean (Glycine max). II. Fate of 15N-labelled protein

The absorption of nitrate, protein metabolism and the source of nitrogen for proline synthesis were studied in soybean ( Glycine max L. cv. Akisengoku) with ¹⁵N tracer technique under water stress conditions. The absorption of nitrate was sensitive to water stress and the flow of nitrate into the leaves completely ceased under severe stress conditions. Net protein loss from the water-stressed leaves was attributable to both a decrease in synthetic activity and a stimulation of protein degradation. Proline and asparagine accumulated extensively in the severely water-stressed plant tissues, especially in the younger green leaves. Fifty four % of the loss of leaf protein-¹⁵N during the stress period was balanced by a gain in ¹⁵N in the free amino acids, 41% being found in proline and asparagine. The increase in ¹⁵N content of the free proline was 3 times greater than the decrease in ¹⁵N content of the protein-bound proline in the leaf. The results indicate that the accumulation of proline in response to water stress was caused by enhanced synthesis and that the nitrogen source for this proline is the leaf protein. The possible association of these findings with stress tolerance is discussed.     

Phenology and autumnal accumulation of N reserves in belowground organs of wetland helophytes Phragmites australis and Glyceria maxima affected by nutrient surplus

Two co-occurring dominant wetland helophytes and potential competitors, Phragmites australis and Glyceria maxima, were cultivated under N, P availabilities simulating the trophic status of wetlands with different fertility (oligo- and eutrophic). The long-term outdoor cultivation was performed with the goal to characterise the extent to which the nutrient enrichment affects plant growth, phenology, and particularly, the accumulation of N storage compounds in belowground organs of wetland rhizomatous plants prior to the onset of winter dormancy. In the present study, both species responded similarly to nutrient surplus. The enhanced growth, delayed shoot senescence, and delayed retranslocation of N into belowground organs were found in both species in eutrophic treatment. Furthermore, N levels remaining in dry leaves were proportionally related to those in living ones, being significantly higher in eutrophic treatment. The efficiency of N retranslocation from senescing leaves varied around 60% in both species and treatments. The formation of N reserves was, however, not disrupted in either species. Although plants in eutrophic treatments accumulated N in their belowground organs significantly later in the season (in the September–December period), the amount of accumulated N was sufficient to reach high belowground N standing stock. Considering formation of N reserves, the differences in species response to treatments were negligible. Phragmites and Glyceria accumulated similar belowground N standing stock prior to the winter. Glyceria may, however, additionally profit from N standing stock of over-wintering green leaves and from the potential of growth and N assimilation during a mild winter period, which is not possible in fully dormant Phragmites.     

Breeding system of the subantarctic plant species Pringlea antiscorbutica R. Br. and search for potential insect pollinators in the Kerguelen Islands

The reproductive biology of Pringlea antiscorbutica R. Br. (Brassicaceae), an endemic species from the southern Indian Ocean islands, is investigated here. Controlled crosses were performed between plants grown in a common garden on the Kerguelen Islands. Searching for potential insect pollinators, we investigated the flying ability of all insects known from the Kerguelen Islands. We showed evidence for self-compatibility, low pollen limitation in the absence of a biotic pollen vector and slight selfing depression in P. antiscorbutica. The insects present on the islands are mostly wingless and are not likely to be efficient pollinators of this species. Since P. antiscorbutica shows morphological adaptations to insect pollination despite the absence of pollinators in its present range, we conclude that this species recently evolved from insect pollination to wind and/or autonomous pollination.     

Quinolizidine alkaloids in petteria ramentacea and the infesting aphids, Aphis cytisorum

Alkaloid extracts of Petteria ramentacea (leaves, fruits, stem) contained cytisine, N-methylcytisine, anagyrine as major alkaloids, and lupanine, 5,6-dehydrolupanine and rhombifoline as minor constituents. Aphids (Aphis cytisorum) feeding on this plant accumulated cytisine in a concentration of 21 mmol/kg fresh weight (0.4 %).     

A metal-accumulator mutant of Arabidopsis thaliana.

A mutation designated man1 (for manganese accumulator) was found to cause Arabidopsis thaliana seedlings to accumulate a range of metals. The man1 mutation segregated as a single recessive locus located on chromosome 3. When grown on soil, mutant seedlings accumulated Mn (7.5 times greater than wild type), Cu (4.6 times greater than wild type), Zn (2.8 times greater than wild type), and Mg (1.8 times greater than wild type) in leaves. In addition to these metals, the man1 mutant accumulated 2.7-fold more S in leaves, primarily in the oxidized form, than wild-type seedlings. Analysis of seedlings grown by hydroponic culture showed a similar accumulation of metals in leaves of man1 mutants. Roots of man1 mutants also accumulated metals, but unlike leaves they accumulated 10-fold more total Fe (symplasmic and apoplasmic combined) than wild-type roots. Roots of man1 mutants possessed greater (from 1.8- to 20-fold) ferric-chelate reductase activity than wild-type seedings, and this activity was not responsive to changes of Mn nutrition in either genotype. Taken together, these results suggest that the man1 mutation disrupts the regulation of metal-ion uptake or homeostasis in Arabidopsis.     

Maintenance of High Photosynthetic Rates during the Accumulation of High Leaf Starch Levels in Sunflower and Soybean

Sunflower (cv. “Mammoth Greystripe”) and soybean (Merr. cv. “Amsoy 71”) leaves were exposed to continuous light for at least 52 hours in an attempt to determine the relationship between leaf starch levels and photosynthetic rates. Immature rapidly expanding and relatively mature slowly expanding sunflower leaves were studied. After 52 hours continuous light, the rapidly expanding leaves accumulated high starch levels (3.3 milligrams per square centimeter, 43% of dry weight) with only about a 10% decline from the initial photosynthetic rate of 42 milligrams CO₂ per square decimeter per hour. Under the same conditions, the slowly expanding leaves accumulated less starch, but the photosynthetic rate declined 30%. Soybean leaves, which were slowly expanding, accumulated less starch than sunflower leaves (2.1 milligrams per square centimeter, 34% of dry weight), and their photosynthetic rates declined only about 10% after 54 hours continuous light.     

六盘山鸡爪大黄茎和叶多糖贮藏分布特征的研究

采用组织化学方法研究了六盘山鸡爪大黄茎和叶中大黄多糖贮藏和分布的特征。结果表明：大黄多糖在茎和叶内的贮藏部位各有特点,在茎中主要分布于皮层、维管束中韧皮部和木质部的薄壁细胞、髓射线和髓,随着茎的成熟大黄多糖有一定程度的增加;叶表皮、叶肉和叶脉中不同程度地分布着较少量的多糖;叶柄维管束外围的基本组织细胞中不同程度地分布着一定数量的大黄多糖,而维管束中分布较少。茎和叶中大黄多糖的贮藏和分布总体较少。     

Involvement of polyamines in root development at low temperature in the subantarctic cruciferous species Pringlea antiscorbutica

Polyamine involvement in root development at low temperature was studied in seedlings of Pringlea antiscorbutica R. Br. This unique endemic cruciferous species from the subantarctic zone is subjected to strong environmental constraints and shows high polyamine contents. In the present study, free polyamine levels were modified by inhibitors of polyamine biosynthesis (D-arginine, difluoromethylornithine, cyclohexylammonium, and methylglyoxal-bis-guanylhydrazone) and variations of the endogenous pools were compared to changes in root growth. The arginine decarboxylase pathway, rather than that of ornithine decarboxylase, seemed to play a major role in polyamine synthesis in Pringlea antiscorbutica seedlings. Root, but not shoot, phenotypes were greatly affected by these treatments, which modified polyamine endogenous levels according to their expected effects. A positive correlation was found between agmatine level and growth rate of the primary root. Spermidine and spermine contents also showed positive correlations with primary root growth whereas the putrescine level showed neutral or negative effects on this trait. Free polyamines were therefore found to be differentially involved in the phenotypic plasticity of root architecture. A comparison of developmental effects and physiological concentrations suggested that agmatine and spermine in particular may play a significant role in the control of root development.     

Accumulation of a 22-kDa protein and its mRNA in the leaves of Raphanus sativus in response to salt stress or water deficit

The response of 10-day-old seedlings of Raphanus sativus L. cv. Fakir to salt stress (100 m M to 200 m M NaCl) was investigated. Three weeks after initiation of salt treatment, the fresh weight of the shoots of salt-treated plants was half that of untreated plants. The salt stress resulted in the accumulation of Na⁺, preferably in the old leaves. The K⁺ level was reduced by as much as 50% in the old leaves of NaCl-treated plants, whereas this reduction was only 20–25% in the young leaves. Free proline accumulated in all aerial organs, and the highest levels were found in the young leaves. Patterns of total proteins extracted from the leaves of control or salt-treated plants were compared. The most obvious change concerned a 22-kDa, pl 7.5 polypeptide, which accumulated after exposure of the plants to NaCl. The appearance of this polypeptide was also mediated by a rapid drought stress, and sequencing indicated that it is related to the Künitz protease inhibitor family. A cDNA clone corresponding to the radish 22-kDa polypeptide was obtained and sequenced. Northern blot analysis showed that salt stress induces a large accumulation of this mRNA in the leaves of radish.