Elevated CO2 induces physiological, biochemical and structural changes in leaves of Arabidopsis thaliana

Leaves of Arabidopsis thaliana grown under elevated or ambient CO2 (700 or 370 micromol mol(-1), respectively) were examined for physiological, biochemical and structural changes. Stomatal characters, carbohydrate and mineral nutrient concentrations, leaf ultrastructure and plant hormone content were investigated using atomic absorption spectrophotometry, transmission electron microscopy and enzyme-linked immunosorbent assay (ELISA). Elevated CO2 reduced the stomatal density and stomatal index of leaves, and also reduced stomatal conductance and transpiration rate. Elevated CO2 increased chloroplast number, width and profile area, and starch grain size and number, but reduced the number of grana thylakoid membranes. Under elevated CO2, the concentrations of carbohydrates and plant hormones, with the exception of abscisic acid, increased whereas mineral nutrient concentrations declined. These results suggest that the changes in chloroplast ultrastructure may primarily be a consequence of increased starch accumulation. Accelerated A. thaliana growth and development in elevated CO2 could in part be attributed to increased foliar concentrations of plant hormones. The reductions in mineral nutrient concentrations may be a result of dilution by increased concentrations of carbohydrates and also of decreases in stomatal conductance and transpiration rate.     

Elevated CO2 enhances stomatal responses to osmotic stress and abscisic acid in Arabidopsis thaliana

A , carbon assimilation rate
ABA, abscisic acid
Ci , intercellular space CO₂ concentration
g , leaf conductance
WUE, water use efficiency

Carbon dioxide and abscisic acid (ABA) are two major signals triggering stomatal closure. Their putative interaction in stomatal regulation was investigated in well-watered air-grown or double CO₂-grown Arabidopsis thaliana plants, using gas exchange and epidermal strip experiments. With plants grown in normal air, a doubling of the CO₂ concentration resulted in a rapid and transient drop in leaf conductance followed by recovery to the pre-treatment level after about two photoperiods. Despite the fact that plants placed in air or in double CO₂ for 2 d exhibited similar levels of leaf conductance, their stomatal responses to an osmotic stress (0·16–0·24 MPa) were different. The decrease in leaf conductance in response to the osmotic stress was strongly enhanced at elevated CO₂. Similarly, the drop in leaf conductance triggered by 1 μ M ABA applied at the root level was stronger at double CO₂. Identical experiments were performed with plants fully grown at double CO₂. Levels of leaf conductance and carbon assimilation rate measured at double CO₂ were similar for air-grown and elevated CO₂-grown plants. An enhanced response to ABA was still observed at high CO₂ in pre-conditioned plants. It is concluded that: (i) in the absence of stress, elevated CO₂ slightly affects leaf conductance in A. thaliana ; (ii) there is a strong interaction in stomatal responses to CO₂ and ABA which is not modified by growth at elevated CO₂.     

Structural Adaptation of the Leaf Mesophyll to Shading

Structural characteristics of the mesophyll were studied in five boreal grass species experiencing a wide range of light and water supply conditions. Quantitative indices of the palisade and spongy mesophyll tissues (cell and chloroplast sizes, the number of chloroplasts per cell, the total cell and chloroplast surface area per unit leaf surface area) were determined in leaves of each of the species. The cell surface area and the cell volume in spongy mesophyll were determined with a novel method based on stereological analysis of cell projections. An important role of spongy parenchyma in the photosynthetic apparatus was demonstrated. In leaves of the species studied, the spongy parenchyma constituted about 50% of the total volume and 40% of the total surface area of mesophyll cells. The proportion of the palisade to spongy mesophyll tissues varied with plant species and growth conditions. In a xerophyte Genista tinctoria, the total cell volume, cell abundance, and the total surface area of cells and chloroplasts were 30–40% larger in the palisade than in the spongy mesophyll. In contrast, in a shade-loving species Veronica chamaedris, the spongy mesophyll was 1.5–2 times more developed than the palisade mesophyll. In mesophyte species grown under high light conditions, the cell abundance and the total cell surface area were 10–20% greater in the palisade mesophyll than in the spongy parenchyma. In shaded habitats, these indices were similar in the palisade and spongy mesophyll or were 10–20% lower in the palisade mesophyll. In mesophytes, CO₂ conductance of the spongy mesophyll accounted for about 50% of the total mesophyll conductance, as calculated from the structural characteristics, with the mesophyll CO₂ conductance increasing with leaf shading.     

Soil water deficits decrease the internal conductance to CO2 transfer but atmospheric water deficits do not

The internal conductance to CO₂ supply from substomatal cavitiesto sites of carboxylation poses a large limitation to photosynthesis.It is known that internal conductance is decreased by soil waterdeficits, but it is not known if it is affected by atmosphericwater deficits (i.e. leaf to air vapour pressure deficit, VPD).The aim of this paper was to examine the responses of internalconductance to atmospheric and soil water deficits in seedlingsof the evergreen perennial Eucalyptus regnans F. Muell and theherbaceous plants Solanum lycopersicum (formerly Lycopersiconesculentum) Mill. and Phaseolus vulgaris L. Internal conductancewas estimated with the variable J method from concurrent measurementsof gas exchange and fluorescence. In all three species steady-statestomatal conductance decreased by 30% as VPD increased from1 kPa to 2 kPa. In no species was internal conductance affectedby VPD despite large effects on stomatal conductance. In contrast,soil water deficits decreased stomatal conductance and internalconductance of all three species. Decreases in stomatal andinternal conductance under water deficit were proportional,but this proportionality differed among species, and thus therelationship between stomatal and internal conductance differedamong species. These findings indicate that soil water deficitsaffect internal conductance while atmospheric water deficitsdo not. The reasons for this distinction are unknown but areconsistent with soil and atmospheric water deficits having differingeffects on leaf physiology and/or root–shoot communication. Key words: Carbon dioxide, drought, internal conductance, mesophyll conductance, photosynthesis, stomatal conductance, transfer conductance, vapour pressure deficit, water deficit Received 11 October 2007; Revised 9 November 2007 Accepted 15 November 2007     

Elevated atmospheric CO(2) concentration and diurnal cycle induce changes in lipid composition in Arabidopsis thaliana

Few studies regarding the effects of elevated atmospheric CO(2) concentrations on plant lipid metabolism have been carried out. Here, the effects of elevated CO(2) concentration on lipid composition in mature seeds and in leaves during the diurnal cycle of Arabidopsis thaliana were investigated. Plants were grown in controlled climate chambers at elevated (800 ppm) and ambient CO(2) concentrations. Lipids were extracted and characterized using thin layer chromatography (TLC) and gas liquid chromatography. The fatty acid profile of total leaf lipids showed large diurnal variations. However, the elevated CO(2) concentration did not induce any significant differences in the diurnal pattern compared with the ambient concentration. The major chloroplast lipids monogalactosyldiacylglycerol (MGDG) and phosphatidylglycerol (PG) were decreased at elevated CO(2) in favour of phosphatidylcholine (PC) and phosphatidylethanolamine (PE). Elevated CO(2) produced a 25% lower ratio of 16:1trans to 16:0 in PG compared with the ambient concentration. With good nutrient supply, growth at elevated CO(2) did not significantly affect single seed weight, total seed mass, oil yield per seed, or the fatty acid profile of the seeds. This study has shown that elevated CO(2) induced changes in leaf lipid composition in A. thaliana, whereas seed lipids were unaffected.     

Effects of low and elevated CO2 partial pressure on growth and reproduction of Arabidopsis thaliana from different elevations

Atmospheric CO₂ partial pressure may have been as low as 18 Pa during the Pleistocene and is expected to increase from 35 to 70 Pa before the end of the next century. Low CO₂ reduces the growth and reproduction of C₃ plants, whereas elevated CO₂ often increases growth and reproduction. Plants at high elevation are exposed to reduced CO₂ partial pressure and may be better adapted to the low CO₂ of the Pleistocene. We examined genotypes of Arabidopsis thaliana from different elevations for variation in growth and reproduction at the CO₂ levels of the Pleistocene, the present and the future. Genotypes exhibited limited genetic variation in the response of the production of biomass to changes in CO₂, but showed significant variation in reproductive characters. We found evidence that plants from high elevations may be better adapted to low CO₂ when considering seed number, which is an important component of fitness. Genotypes showed greater variation in the response of seed number between 35 and 20 Pa CO₂ compared to 35 and 70 Pa CO₂. We conclude that present-day C₃ annuals may have greater potential for evolution in response to the low CO₂ of the Pleistocene relative to the elevated CO₂ predicted for the future.     

Study of mechanisms for plant growth promotion elicited by rhizobacteria in Arabidopsis thaliana

Plant growth-promoting rhizobacteria (PGPR) colonize plant roots and exert beneficial effects on plant health and development. We are investigating the mechanisms by which PGPR elicit plant growth promotion from the viewpoint of signal transduction pathways within plants. We report here our first study to determine if well-characterized PGPR strains, which previously demonstrated growth promotion of various other plants, also enhance plant growth in Arabidopsis thaliana. Eight different PGPR strains, including Bacillus subtilis GB03, B. amyloliquefaciens IN937a, B. pumilus SE-34, B. pumilus T4, B. pasteurii C9, Paenibacillus polymyxa E681, Pseudomonas fluorescens 89B-61, and Serratia marcescens 90-166, were evaluated for elicitation of growth promotion of wild-type and mutant Arabidopsis in vitro and in vivo. In vitro testing on MS medium indicated that all eight PGPR strains increased foliar fresh weight of Arabidopsis at distances of 2, 4, and 6 cm from the site of bacterial inoculation. Among the eight strains, IN937a and GB03 inhibited growth of Arabidopsis plants when the bacteria were inoculated 2 cm from the plants, while they significantly increased plant growth when inoculated 6 cm from the plants, suggesting that a bacterial metabolite that diffused into the agar accounted for growth promotion with this strain. In vivo, eight PGPR strains promoted foliar fresh weight under greenhouse conditions 4 weeks after sowing. To define signal transduction pathways associated with growth promotion elicited by PGPR, various plant-hormone mutants of Arabidopsis were evaluated in vitro and in vivo. Elicitation of growth promotion by PGPR strains in vitro involved signaling of brassinosteroid, IAA, salicylic acid, and gibberellins. In vivo testing indicated that ethylene signaling was involved in growth promotion. Results suggest that elicitation of growth promotion by PGPR in Arabidopsis is associated with several different signal transduction pathways and that such signaling may be different for plants grown in vitro vs. in vivo.     

Salmonella enterica Flagellin Is Recognized via FLS2 and Activates PAMP-Triggered Immunity in Arabidopsis thaliana

Infections with Salmonella enterica belong to the most prominent causes of food poisoning and infected fruits and vegetables represent important vectors for salmonellosis. Recent evidence indicates that plants recognize S. enterica and raise defense responses. Nonetheless, the molecular mechanisms controlling the interaction of S. enterica with plants are still largely unclear. Here, we show that flagellin from S. enterica represents a prominent pathogenassociated molecular pattern （PAMP） in Arabidopsis thaliana, which induces PAMP-triggered immunity （PTI） via the recognition of the fig22 domain by the receptor kinase FLS2. The Arabidopsis fls2 mutant shows reduced though not abolished PTI activation, indicating that plants rely also on recognition of other S. enterica PAMPs. Interestingly, the S. enterica type III secretion system （T3SS） mutant prgH- induced stronger defense gene expression than wild-type bacteria in Arabidopsis, suggesting that T3SS effectors are involved in defense suppression. Furthermore, we observe that S. enterica strains show variation in the fig22 epitope, which results in proteins with reduced PTI-inducing activity. Altogether, these results show that S. enterica activates PTI in Arabidopsis and suggest that, in order to accomplish plant colonization, S. enterica evolved strategies to avoid or suppress PTI.     

叶肉导度的组成、大小及其对环境因素的响应

植物光合作用过程中,大气中的CO2需要克服气孔和叶肉细胞等阻力传输到羧化位点。CO2从气孔下腔传输到羧化位点的阻力称为叶肉阻力,其倒数即为叶肉导度。近十年内,叶肉导度已经成为光合作用研究领域的一个重要方面。本文首先系统地阐述了叶肉导度的组成及各部分所占的比重;然后通过与气孔导度的比较,分析叶肉导度的大小及其对光合作用的影响;最后阐述了叶肉导度对环境变化的响应,并分析了其中可能的原因。     

Strong ecological but weak evolutionary effects of elevated CO2 on a recombinant inbred population of Arabidopsis thaliana

Increases in atmospheric CO2 concentration have an impact on plant communities by influencing plant growth and morphology, species interactions, and ecosystem processes. These ecological effects may be accompanied by evolutionary change if elevated CO2 (eCO2) alters patterns of natural selection or expression of genetic variation. Here, a statistically powerful quantitative genetic experiment and manipulations of CO2 concentrations in a field setting were used to investigate how eCO2 impacts patterns of selection on ecologically important traits in Arabidopsis thaliana; heritabilities, which influence the rate of response to selection; and genetic covariances between traits, which may constrain responses to selection. CO2 had strong phenotypic effects; plants grown in eCO2 were taller and produced more biomass and fruits. Also, significant directional selection was observed on many traits and significant genetic variation was observed for all traits. However, no evolutionary effect of eCO2 was detected; patterns of selection, heritabilities and genetic correlations corresponded closely in ambient and elevated CO2 environments. The data suggest that patterns of natural selection and the quantitative genetic parameters of this A. thaliana population are robust to increases in CO2 concentration and that responses to eCO2 will be primarily ecological.     

基于光子学技术研究弱光胁迫早期拟南芥光合损伤机理

弱光限制植物的光合作用,降低了光合作用效率,造成农业产量下降.本文主要研究了弱光处理早期,拟南芥光合作用相关指标的变化.研究中发现在弱光处理的早期,植株生长表型和最大光化学效率（Fv/Fm）没有明显变化,实际光化学效率Y（Ⅱ）以及光系统电子传递效率（ETR）下降较明显.此外,弱光处理原生质体,利用2 ＇,7＇-二氯二氢荧光素二乙酯（dichlorofluorescin diacetate,H2DCF-DA）染色,共聚焦显微镜观察,发现细胞中有较明显的活性氧（ROS）合成,且定位于叶绿体.该研究结果为植物弱光耐受性的研究提供理论依据.     

Antisense inhibition of protein phosphatase 2C accelerates cold acclimation in Arabidopsis thaliana

Two related protein phosphatases 2C, ABI1 and AtPP2CA have been implicated as negative regulators of ABA signalling. In this study we characterized the role of AtPP2CA in cold acclimation. The pattern of expression of AtPP2CA and ABI1 was studied in different tissues and in response to abiotic stresses. The expression of both AtPP2CA and ABI1 was induced by low temperature, drought, high salt and ABA. The cold and drought-induced expression of these genes was ABA-dependent, but divergent in various ABA signalling mutants. In addition, the two PP2C genes exhibited differences in their tissue-specific expression as well as in temporal induction in response to low temperature. To elucidate the function of AtPP2CA in cold acclimation further, the corresponding gene was silenced by antisense inhibition. Transgenic antisense plants exhibited clearly accelerated development of freezing tolerance. Both exposure to low temperature and application of ABA resulted in enhanced freezing tolerance in antisense plants. These plants displayed increased sensitivity to ABA both during development of frost tolerance and during seed germination, but not in their drought responses. Furthermore, the expression of cold-and ABA-induced genes was enhanced in transgenic antisense plants. Our results suggest that AtPP2CA is a negative regulator of ABA responses during cold acclimation.     

A non-destructive selection method for resistance to fusicoccin in Arabidopsis thaliana

A mutagenised population of seeds of Arabidopsis thaliana was allowed to germinate in the presence of the positively charged aminoglycoside hygromycin (4 μg/ml) and the fungal toxin fusicoccin (5×10^–6 m). This hygromycin concentration, which is non-toxic by itself, becomes toxic when used together with fusicoccin, which stimulates cation uptake. Seeds that had germinated after 3–5 days and produced seedlings with green cotyledons were potentially resistant to fusicoccin and were therefore transferred into sterile Magenta vessels containing 1/2-strength Murashige and Skoog medium. This selection procedure is non-destructive, i.e. it allows the recovery of viable seedlings and their growth into adult plants thus permitting direct physiological characterisation. Received: 16 February 1998 / Revision received: 11 August 1998 / Accepted: 13 August 1998     

Efficient regeneration from cotyledon protoplasts in Arabidopsis thaliana

Summary. An efficient and fast regeneration system from cotyledon protoplasts was established for Arabidopsis thaliana accessions C24, Columbia, and Wassilewskija. Culture conditions and media compositions were optimised for the development of protoplasts embedded in thin alginate layers. Unexpectedly, the absence of cytokinins had a positive effect on cell development. Moreover, combined adjustment of -naphthylacetic acid and dicamba concentrations resulted in high plating efficiencies of up to 30%, followed by shoot regeneration within only 19 days after protoplast isolation. The protocol is reproducible, efficient, extremely fast, and regenerated plants are fertile. Thus, this cotyledon-based system could prove useful for studying plant cell and molecular biology in A. thaliana.Correspondence and reprints: Zentrum für angewandte Biowissenschaften, Sonnenstrasse 5, 79104 Freiburg, Federal Republic of Germany.Received December 9, 2002; accepted April 13, 2003; published online September 23, 2003     

拟南芥花粉细胞质游离钙离子荧光测定法

以拟南芥花粉为材料,利用低温装载法在完整的花粉粒中,成功地载入酯化形式的钙离子荧光探针Fura-2/AM。利用荧光比率分析法对花粉细胞质中游离钙离子的分布特点进行研究并测定了花粉细胞内游离钙离子浓度。结果表明花粉萌发初期细胞质内游离钙离子呈极性分布,萌发沟附近的钙离子浓度明显高于其它部位,萌发孔附近最高,花粉细胞核中最低。花粉粒细胞萌发状态下的[Ca2 ]i=246±38nmol/L,该值与花粉粒细胞萌发状态下游离钙离子浓度用其它方法测得值接近,进一步表明所建立的用Fura-2/AM检测拟南芥花粉粒细胞质游离钙离子的方法是可靠的。     

Plasmodesmata in Arabidopsis thaliana suspension cells

Summary. A current challenge in plant biology is to identify the structural and functional components of plasmodesmata (PDs). The use of plant tissue as a source material for plasmodesmal characterisation has had limited success, so we have explored the frequency and features of PDs occurring in suspension cell cultures of Arabidopsis thaliana. This material has the advantages of homogeneity, quantity, and ease of disruption. Using light and electron microscopy and immunostaining for callose and calreticulin, we showed that suspension cells laid down abundant PDs in division walls, and that vestiges of these structures were retained as half PDs even when the cell-to-cell contacts were disrupted during culture growth. Although callose was a reliable marker for PD distribution, which was deposited in an organised collar around the neck of PDs, it was not abundant in unstressed cells. Calreticulin and the chemical stain 3,3-dihexyloxacarbocyanine iodide also provided useful markers when monitoring PDs in cell wall preparations by light microscopy. Purified cell walls were shown to be virtually free of contamination from cytoplasmic components, except for the presence of small amounts of cortical endoplasmic reticulum attached to PDs. Hence, clean cell walls from A. thaliana suspension cells provide a valuable resource for a proteomic approach to the analysis of plasmodesmal components.Correspondence and reprints: John Innes Centre, Norwich Research Park, Colney, Norwich NR4 7UH, United Kingdom.     

The Arabidopsis thaliana aquaporin AtPIP1;2 is a physiologically relevant CO₂ transport facilitator

Cellular exchange of carbon dioxide (CO₂) is of extraordinary importance for life. Despite this significance, its molecular mechanisms are still unclear and a matter of controversy. In contrast to other living organisms, plants are physiologically limited by the availability of CO₂. In most plants, net photosynthesis is directly dependent on CO₂ diffusion from the atmosphere to the chloroplast. Thus, it is important to analyze CO₂ transport with regards to its effect on photosynthesis. A mutation of the Arabidopsis thaliana AtPIP1;2 gene, which was characterized as a non‐water transporting but CO₂ transport‐facilitating aquaporin in heterologous expression systems, correlated with a reduction in photosynthesis under a wide range of atmospheric CO₂ concentrations. Here, we could demonstrate that the effect was caused by reduced CO₂ conductivity in leaf tissue. It is concluded that the AtPIP1;2 gene product limits CO₂ diffusion and photosynthesis in leaves.     

The metal ion transporter IRT1 is necessary for iron homeostasis and efficient photosynthesis in Arabidopsis thaliana

The mutants irt1-1 and irt1-2 of Arabidopsis thaliana were identified among a collection of T-DNA-tagged lines on the basis of a decrease in the effective quantum yield of photosystem II. The mutations responsible interfere with expression of IRT1, a nuclear gene that encodes the metal ion transporter IRT1. In irt1 mutants, photosensitivity and chlorophyll fluorescence parameters, as well as abundance and composition of the photosynthetic apparatus, are significantly altered. Additional effects of the mutation under greenhouse conditions, including chlorosis and a drastic reduction in growth rate and fertility, are compatible with a deficiency in iron transport. Propagation of irt1 plants on media supplemented with additional quantities of iron salts restores almost all aspects of wild-type behaviour. The irt2-1 mutant, which carries an En insertion in the highly homologous IRT2 gene of Arabidopsis thaliana, was identified by reverse genetics and shows no symptoms of iron deficiency. This, together with the finding that irt1-1 can be complemented by 35S::IRT1 but not by 35S::IRT2, demonstrates that, although the products of the two genes are closely related, only AtIRT1 is required for iron homeostasis under physiological conditions.