Summary 221

I. Introduction 221

1. The role of sucrose in plants 221

2. C4photosynthesis 222

II. Pathway of sucrose synthesis in C4plants 223

III. Control of sucrose synthesis in C4plants 225

1. The diurnal cycle in C3plants 225

2. The diurnal cycle in C4plants 227

3. Abiotic stress and sucrose synthesis 230

4. Integration of sucrose synthesis with other metabolic pathways 232

IV. Future prospects 233

Acknowledgements 234

References 234

Sucrose has a central role in the metabolism of all higher plants. Here we discuss the location of sucrose synthesis in the leaves of C4 plants, the control of flux through the pathway and its response to environmental conditions, and finally some of the prospects for the genetic manipulation of sucrose metabolism in C4 plants. Much of our knowledge about sucrose synthesis in plants comes from C3 species; it is evident, from those studies of C4 species that are available, that there are many similarities between sucrose synthesis in C3 and C4 plants. However, some of the basic regulatory mechanisms common to both C3 and C4 plants seem to have been adapted for the specialized photosynthetic metabolism of C4 plants. There are also some other important differences: for example, some C4 species possess photosynthetic cells with little or no capacity for sucrose synthesis. For these reasons it is not always appropriate to extrapolate directly from C3 to C4 plants. Even where data are available from one C4 species, usually maize, it should not be assumed that all C4 plants are necessarily the same in every respect. Although C4 plants constitute a small fraction of the plant kingdom as a whole, they include a disproportionate number of agriculturally important species. C4 plants also have some advantages over C3 plants for studies of the regulation of cell-specific gene expression and the control of metabolism. For these reasons we suggest that they are worthy of more attention by researchers.

The aim of this study was to investigate whether genetically different pollen donors (Betula pendula clones) differed in pollen-tube growth rate across 11 maternal plants and in vitro, and whether the differences between the donors were consistent across the recipients. To compare the seed-siring success of competing pollen donors, a two-donor hand-pollination experiment with six donors and six recipients was conducted. The experiments were performed at a plastic-house seed orchard. The donors showed significant variation in pollen-tube growth rate on all the 11 recipients. The rankings of the pollen donors were statistically consistent across different maternal plants. A significant positive correlation between pollen tube growth in vivo and in vitro was found. The seed-siring success of two competing pollen donors was unequal in 20 of 29 cases and there was a significant positive correlation between seed-siring success and pollen-tube growth rate in vivo and in vitro. The results show that fertilizations are not random and pollen competition operates in a B. pendula seed orchard population.

Exposure of Chlorella vulgaris to elevated concentrations of copper, chromium, nickel and zinc led to intracellular accumulation of high concentrations of these metals. Concomitantly, accumulation of free proline occurred, depending on the concentration of metals in the external medium or in the cell. The greater the toxicity or accumulation of a metal, the greater the amount of intracellular proline in algal cells. However, higher concentrations of copper and chromium were inhibitory to proline accumulation by the test organism. The accumulation of proline was triggered within a few hours of metal treatment. Test metals also induced lipid peroxidation; copper was the most efficient inducer whereas zinc was the least. Pretreatment of C. vulgaris with proline counteracted metal-induced lipid peroxidation and potassium ion efflux. Thus the present work shows a protective effect of proline on metal toxicity through inhibition of lipid peroxidation.

Feeding dynamics in the spittlebug Philaenus spumarius were analysed to determine the food source. Evidence from previous reports indicates that this species and similar insects feed from the xylem. However, it is not clear whether they extract sap from the main transpiration stream, or from some subsidiary compartment of xylem, such as immature or cavitated vessels. In feeding from such subsidiary compartments, the insects could avoid the large hydraulic tensions thought to prevail in xylem on the main transpiration stream. Philaenus, for example, is shown here to initiate feeding, and to feed rapidly, from plants in which the full xylem tension is estimated at −10 bar. From measurements of feeding rates and feeding patterns in stressed and unstressed plants, it is shown that Philaenus cannot feed from subsidiary xylem compartments. It is concluded that this insect feeds directly from the main transpiration stream, and at the full hydraulic tension which prevails there. The anatomy of the feeding pump in Philaenus appears consistent with this conclusion. Spittlebugs therefore offer an experimental tool for quasi- non-invasive sampling of the transpiration stream.

A model of seed germination is proposed which uses a variable with the units of an osmotic potential (virtual osmotic potential) to integrate the effect of a constant or a varying water potential. This differs from existing models that describe the effects of fixed water potentials on germination, or the effects of fixed priming water potentials on the subsequent germination at a fixed water potential. When a seed is sown, the virtual osmotic potential is assumed to fall at a rate that depends on the ambient water potential, and on the difference between its current and a minimum value. Radicle growth is assumed to initiate when the difference between the ambient water potential and the virtual osmotic potential exceeds a threshold. The germination of carrot and onion seeds at various fixed potentials below 0 MPa was well described by the virtual osmotic potential model. The model was also used to simulate the results of experiments in which seeds were given a single step change in water potential.

The facultative hemiparasite Rhinanthus minor was grown alone, or with either Hordeum vulgare or Trifolium alpestre as a host. All plants were fed 5 mM nitrate. In the parasites, successful attachment led to dramatic increases in growth (particularly with barley as host) and in tissue concentrations of total N, NO3−, H2PO4− and K+, as well as moderate increases in Cl− and SO42−. Mg2+ and Ca2+ concentrations either remained unchanged, increased or decreased, depending on the host. Xylem sap was collected from leaf veins (barley) or the stem (Rhinanthus and clover), by applying pneumatic pressure to the rooted soil to raise the soil water potential so that in vivo xylem sap exuded from small incisions once the applied pressure balanced the tension exerted by transpiration. High balancing pressures were needed for unattached Rhinanthus, whereas simultaneous collection of xylem sap from attached Rhinanthus and its host, at a much lower balancing pressure, was possible only when the parasite was enclosed in polyethylene film and high transpiration abolished. From plots of xylem sap volume flow versus applied pressure, the hydraulic conductivity Lp and the overall hydraulic resistance r were obtained. The value of r was high in unattached Rhinanthus and dramatically decreased by attachment to a host, highlighting the improved access to water gained by Rhinanthus when parasitizing a host. For the parasites, attachment resulted in greatly increased concentrations and solute flow rates in xylem of K+, NO3−, H2PO4− and amino acids, and decreased concentrations of Ca2+ and Mg2+. Estimates of NO3− reduction in the parasite showed that 99% was reduced in unattached plants, 85% in those attached to clover and 52% in those attached to barley. In the parasitized hosts, ion concentrations in xylem sap were somewhat increased relative to unparasitized controls. Conversely, xylem sap amino-acid concentrations in infected hosts were decreased. Glutamine (Gln) was the principal amino acid in xylem sap of unattached Rhinanthus, but after attachment to both hosts asparagine (Asn) predominated. In clover, Asn was the major transport amino acid, suggesting massive transfer of Asn from host to parasite. In unparasitized barley, however, Gln was the major xylem sap amino acid, but unexpectedly, in this species Asn was induced as the principal amino acid when parasitized. The results are discussed in terms of mutual host–parasite interactions.

A centrifugal method for extracting apoplastic sap from roots of lupin (Lupinus angustifolius) and pea (Pisum sativum) plants, and a method to analyse malic dehydrogenase in the sap using capillary electrophoresis, are described. Osmolality of apoplastic sap was relatively constant at relative centrifugal forces (RCFs) between 600 and 3000 g for lupin, and between 600 and 4000 g for pea. Electropherograms of a marker enzyme (malic dehydrogenase) and other components in apoplastic and symplastic saps revealed that contamination occurred at 7000 g. It is suggested that apoplastic sap expelled from plant roots at RCF between 600 and 3000 g is free from symplastic contamination, and is regarded as being of apoplastic origin. The proposed method was used to measure apoplastic pH changes in the plant roots in response to external pH, ammonium, nitrate and vanadate.

Both sensitive and tolerant clones of aspen (Populus tremuloides) were exposed to ozone using four different exposure regimes under controlled environmental conditions. Based on data on ambient ozone from 10 cities in the USA, three treatments of 4-wk exposure to the same SUM06 (an accumulation of hourly O3 concentrations greater than 0.06 ml l−1) were constructed. The regimes allowed us to investigate: (a) the importance of long (3 wk, treatment 1) versus short (1 wk, treatment 2) duration of regimes with high peaks; (b) the effect of treatments with variable peak occurrence (treatments 1 and 2) versus uniform peak occurrence (treatment 3) during the exposure period. Nonfumigated control plants were maintained at ozone concentrations <10 nl l−1. Bifacial black necrosis, a typical symptom of ozone injury on aspen leaves, occurred on both clones after 2 wk exposure. Up to 60% of the leaves on the sensitive clone were injured, with an average of 6% of total leaf area injured. In the tolerant clone only 10% of the leaves were injured, with less than 1% of the total leaf area symptomatic. The severity of injury was consistently greatest in treatment 2, followed by treatments 1 and 3, respectively. The interval between peak exposures was less important than the occurrence of peaks versus a stable maximum concentration. Premature leaf abscission occurred in the sensitive clone. Measures of gas exchange demonstrated reduced photosynthesis under ozone fumigation, but exposure regime was not a significant factor. Concentrations of two antioxidants, ascorbic acid and glutathione, were almost always greater in the resistant than in the sensitive clone, but the differences were not statistically significant. The levels of these antioxidants in aspen leaves did not change with ozone fumigation or leaf age.

Potted subterranean clover (Trifolium subterraneum) plants of different ages were exposed to 70 nl l−1 ozone for 6 h, either during the light or during the dark period in a laboratory-based climate chamber. There was limited visible leaf injury on plants which were 14–20 and 28–34 d old and no significant decrease in biomass after daytime ozone exposure. The oldest leaves of 22–26 d old plants exhibited severe visible injury, which was associated with a significant reduction in biomass in 24–26 d old plants. Thus, ozone-induced visible injury of different magnitude developed in all plants, but was associated with biomass reduction only during a limited period of the plant's life-span. Apart from modifying ozone uptake by plants, climatic conditions are important as growth modifiers. It is suggested that subterranean clover plants of defined developmental stages should be used in bioindication of ozone. Night ozone exposure injured significantly fewer leaves than day exposure. However, some leaves developed visible injury even after night ozone exposure. Night uptake of ozone may be of more importance in northern than in central and southern Europe, because summer nights are short and, for a certain period, never completely dark.

The responses to CO2 of perennial grasses (Danthonia richardsonii and Phalaris aquatica) and legumes (Lotus pedunculatus and Trifolium repens) were compared under controlled conditions for isolated plants, monoculture stands and mixed-species stands along a N gradient to test whether: plant–plant interactions between species in mixed stands changed with concentration of CO2; responses to CO2 of species in mixtures could be related to their responses as single stands; responses of mixtures to CO2 could be related to the responses of individual species to CO2 and to competition. Plants were grown for 60 d in sand, using nutrient solutions (six nitrate concentrations from 0.25 to 16 mM NO3), at ambient (c. 357 μl l−1) or elevated CO2 (c. 712 μl l−1). Species dominance in the mixtures depended more on the range of N than of CO2 concentration provided: T. repens and L. pedunculatus dominated at low concentrations of N; L. pedunculatus and P. aquatica performed better at high concentrations. Responses of species in mixtures to CO2 were related to their responses in monocultures but not to those of isolated plants. Species biomass proportions in mixtures under ambient CO2 determined the outcome of mixture responses to CO2 more than of individual species responses to CO2. These results emphasize the influence of plant–plant interactions on community responses to CO2, since mixture behaviour under elevated CO2 could not be scaled-up from responses by isolated plants in this experiment.

The apoplastic pH of intact green leaves of Bromus erectus was measured non-invasively by inserting blunt microelectrodes through stomatal openings. After making electrical contact, the recorded signal was stable for hours, yielding a pH of 4.67±0.10. The leaves responded to ‘light-off’ with an initial transient acidification and subsequent sustained alkalinization of 0.2–0.3 pH; ‘light-on’ caused the opposite response. Flushing the leaves with 280 nmol NH3 mol−1 air within 18±6 s alkalinized the apoplast by 0.22±0.07 pH, followed by a slower pH increase to reach a steady-state alkalinization of 0.53±0.14 after 19±7 min. This pH shift was persistent as long as the NH3 was flushed, and readily returned to its initial value after replacing the NH3 with clean air. The resultant [NH4+] increase within the apoplast was measured with a NH4+-selective microelectrode. In the presence of 280 nmol NH3 mol−1 air, apoplastic NH4+ initially increased within 15±10 s to 1.53±0.41 mM, to reach a steady state of 1.62±0.16 mM after 27±7 min. An apoplastic buffer capacity of 6 mM pH−1 unit was calculated from the initial changes of pH and [NH4+ ], whereas the steady-state values yielded 2.7 mM pH−1. Infiltrated leaves responded to NH4+ with concentration-dependent depolarizations, the maxima of which yielded saturation kinetics indicating carrier-mediated NH4+ uptake into adjacent cells, as well as a linear component indicating non-specific transport. We infer that the initial alkalinization is due to rapid conversion of NH3 to NH4+, whereas the slower pH increase would be caused by regulatory processes involving both membrane transport, and (mainly) NH4+ assimilation. Possible consequences of the NH3-induced pH shift for the development of plants growing in polluted areas are discussed.

Several physiological processes controlling tree phenology remain poorly understood and in particular bud dormancy. Many studies have emphasised the action of chilling temperatures in breaking dormancy. However, the effect of the preceding summer temperatures has rarely been investigated although there is some evidence that they may be involved in the settlement and intensity of dormancy as well as cold acclimation. In this paper, thermal time to budburst in relation to the duration of chilling outdoors, preceding summer temperatures and forcing temperatures was studied by outdoors experiments in seedlings of Platanus acerifolia, Vitis vinifera, Quercus pubescens and Castanea sativa. Results showed that temperatures of the preceding summer had no significant effect on the timing of budburst, P. acerifolia and Q. pubescens showed a very weak response to the duration of chilling, and the phenological characteristics of each species were found to be adapted to the climate conditions of its own geographical area. The phenological model used in this study explained 82–100% of the variance of the data without taking into account summer temperatures. Thus, although summer temperatures may be well involved in the intensity of dormancy and cold hardiness, they do not significantly affect budburst and therefore may not need to be considered in phenological models for predicting budburst.

Leaf anatomical parameters such as leaf mass per area (LMA) and biochemical composition can be used as indicators of leaf photosynthetic capacity. The aims of this study are to evaluate the potential of reflectance spectroscopy of fresh leaves for assessing and predicting various parameters, anatomical (LMA and tissue thickness) and biochemical (nitrogen concentration). This paper describes results obtained with fresh leaves of holm oak (Quercus ilex), an evergreen oak that is widely distributed from mesic to xeric habitats in the Mediterranean. Fresh leaves (560) were collected over 3 yr at six different sites, from the top to the bottom of the canopy. The reflectance of each leaf was obtained within 1 h of sampling with an NIRSystems 6500 spectrophotometer over the range 400–2500 nm. LMA was determined for all samples; biochemical and anatomical measurements were conducted over representative subsample populations of 92 and 87 leaves, respectively. Stepwise regression calibrations and partial least squares (PLS) calibrations were developed and compared with different spectral regions and mathematical treatments. Calibration equations had high coefficients of determination (r2 ranging from 0.94 for nitrogen to 0.98 for LMA and tissue thickness). The PLS regressions gave better results than stepwise regressions for all parameters studied. Compared with regressions calculated on raw spectral data, calculations on second derivatives of spectra improved results in all cases. The use of scatter corrections also improved results. These results show that visible and near-infra red reflectance can be used for accurately predicting anatomical parameters and the nitrogen concentration of fresh holm oak leaves. The results support the suggestion that high spectral resolution imaging spectrometry can be a useful tool for assessing functional processes in forest ecosystems.

Vulnerability of stem xylem to cavitation was measured in 10 species of conifers using high pressure air to induce xylem embolism. Mean values of air pressure required to induce a 50% loss in hydraulic conductivity (φ50) varied enormously between species, ranging from a maximum of 14.2±0.6 MPa (corresponding to a xylem water potential of −14.2 MPa) in the semi-arid species Actinostrobus acuminatus to a minimum of 2.3±0.2 MPa in the rainforest species Dacrycarpus dacrydioides. Mean φ50 was significantly correlated with the mean rainfall of the driest quarter within the distribution of each species. The value of φ50 was also compared with leaf drought tolerance data for these species in order to determine whether xylem dysfunction during drought dictated drought response at the leaf level. Previous data describing the maximum depletion of internal CO2 concentration (ci) in the leaves of these species during artificial drought was strongly correlated with φ50 suggesting a primary role of xylem in effecting leaf drought response. The possibility of a trade-off between xylem conductivity and xylem vulnerability was tested in a sub-sample of four species, but no evidence of an inverse relationship between φ50 and either stem-area specific (Ka) or leaf-area specific conductivity (K1) was found.

Predictions of root architecture have been tested using only a narrow set of environmental conditions and species. There is little information on root architecture of plants growing on semi-arid grasslands where soil nutrient heterogeneity and defoliation might impose particular restrictions on root growth. We conducted a split-root study to evaluate the effect of nonuniform nutrient supply and clipping on root architecture and biomass allocation of three range grasses found in the Intermountain West of the USA: ‘Whitmar’, a cultivar of Pseudoroegneria spicata, Bromus tectorum (cheatgrass), and ‘Hycrest’, a hybrid between Agropyron desertorum and A. cristatum. Root systems of individual seedlings were equally divided between the two divisions of a partitioned 10-l pot, and each of the divisions was independently supplied with ‘high or ‘low’ nutrient solution. Plants were either clipped or not. Characteristics of root architecture (topology and link length), root size (root length and number of root tips), and biomass distribution (specific root length and root mass[ratio ]root volume ratio) were evaluated. Root topology (branching distribution) changed with nutrient treatments and defoliation in all three species. Length of links (a segment of root between two branches, or between a node and a root tip) responded to nutrient treatment only in defoliated plants of ‘Whitmar’, in which relative growth rate was associated with changes in architectural traits (external–external and internal–internal link length), while biomass accumulation and relative growth rate in cheatgrass and ‘Hycrest’ maintained allometric relationships with root length. Our results suggest that large plasticity in root architecture, together with inherently low relative growth rate are adaptations that allow ‘Whitmar’ to grow well in heterogeneous and poor soil-nutrient conditions. Cheatgrass and ‘Hycrest’ exhibited characteristics suggesting capability for rapid exploitation of nutrient patches.

The assumption that the mushroom stem has the ability to undergo autonomic straightening enables a mathematical model to be written that accurately mimics the gravitropic reaction of the stems of Coprinus cinereus. The straightening mechanism is called curvature compensation here, but is equivalent to the ‘autotropism’ that often accompanies the gravitropic reactions of axial organs in plants. In the consequently revised local curvature distribution model, local bending rate is determined by the difference between the ‘bending signal’ (generated by gravitropic signal perception systems) and the ‘straightening signal’ (proportional to the local curvature at the given point). The model describes gravitropic stem bending in the standard assay with great accuracy but has the virtue of operating well outside the experimental data set used in its derivation. It is shown, for example, that the mathematical model can be fitted to the gravitropic reactions of stems treated with metabolic inhibitors by a change of parameters that parallel the independently derived physiological interpretation of inhibitor action. The revised local curvature distribution model promises to be a predictive tool in the further analysis of gravitropism in mushrooms.