ALBERT

Notes: Thermotolerance induced by isoprene has been assessed during heat bursts but there is little information on the ability of endogenous isoprene to confer thermotolerance under naturally elevated temperature, on the interaction between isoprene-induced thermotolerance and light stress, and on the persistence of this protection in leaves recovering at lower temperatures. Moderately high temperature treatment (38 °C for 1.5 h) reduced photosynthesis, stomatal conductance, and photochemical efficiency of photosystem II in isoprene-emitting, but to a significantly lower extent than in isoprene-inhibited Phragmites australis leaves. Isoprene inhibition and high temperature independently, as well as together, induced lipid peroxidation, increased level of H2O2, and increased catalase and peroxidase activities. However, leaves in which isoprene emission was previously inhibited developed stronger oxidative stress under high temperature with respect to isoprene-emitting leaves. The heaviest photosynthetic stress was observed in isoprene-inhibited leaves exposed to the brightest illumination (1500 µmol m−2 s−1) and, in general, there was also a clear additive effect of light excess on the formation of reactive oxygen species, antioxidant enzymes, and membrane damage. The increased thermotolerance capability of isoprene-emitting leaves may be due to isoprene ability to stabilize membranes or to scavenge reactive oxygen species. Irrespective of the mechanism by which isoprene reduces thermal stress, isoprene-emitting leaves are able to quickly recover after the stress. This may be an important feature for plants coping with frequent and transient temperature changes in nature.

Notes: Olive (Olea europea L) is one of the most valuable and widespread fruit trees in the Mediterranean area. To breed olive for resistance to salinity, an environmental constraint typical of the Mediterranean, is an important goal. The photosynthetic limitations associated with salt stress caused by irrigation with saline (200 mm) water were assessed with simultaneous gas-exchange and fluorescence field measurements in six olive cultivars. Cultivars were found to possess inherently different photosynthesis when non-stressed. When exposed to salt stress, cultivars with inherently high photosynthesis showed the highest photosynthetic reductions. There was no relationship between salt accumulation and photosynthesis reduction in either young or old leaves. Thus photosynthetic sensitivity to salt did not depend on salt exclusion or compartmentalization in the old leaves of the olive cultivars investigated. Salt reduced the photochemical efficiency, but this reduction was also not associated with photosynthesis reduction. Salt caused a reduction of stomatal and mesophyll conductance, especially in cultivars with inherently high photosynthesis. Mesophyll conductance was generally strongly associated with photosynthesis, but not in salt-stressed leaves with a mesophyll conductance higher than 50 mmol m−2 s−1. The combined reduction of stomatal and mesophyll conductances in salt-stressed leaves increased the CO2 draw-down between ambient air and the chloroplasts. The CO2 draw-down was strongly associated with photosynthesis reduction of salt-stressed leaves but also with the variable photosynthesis of controls. The relationship between photosynthesis and CO2 draw-down remained unchanged in most of the cultivars, suggesting no or small changes in Rubisco activity of salt-stressed leaves. The present results indicate that the low chloroplast CO2 concentration set by both low stomatal and mesophyll conductances were the main limitations of photosynthesis in salt-stressed olive as well as in cultivars with inherently low photosynthesis. It is consequently suggested that, independently of the apparent sensitivity of photosynthesis to salt, this effect may be relieved if conductances to CO2 diffusion are restored.

Notes: Leaves of the monoterpene emitter Quercus ilex were exposed to a temperature ramp with 5 °C steps from 30 to 55 °C while maintained under conditions in which endogenous emission of monoterpenes was allowed or suppressed, or under fumigation with selected exogenous monoterpenes. Fumigation with monoterpenes reduced the decline of photosynthesis, photorespiration and monoterpene emission found in non-fumigated leaves exposed to high temperatures. It also substantially increased respiration when photosynthesis and photorespiration were inhibited by low O2 and CO2-free air. These results indicate that, as previously reported for isoprene, monoterpenes may help plants cope with heat stress. Monoterpenes may enhance membrane stability, thus providing a rather non-specific protection of photosynthetic and respiratory processes. Monoterpene emission was maximal at a temperature of 35 °C and was inhibited at higher temperatures. This is likely to be the result of the temperature dependency of the enzymes involved in monoterpene synthesis. In contrast to other monoterpenes, cis- and trans-β-ocimene did not respond to exposure to high temperatures. Cis-β-ocimene also did not respond to low O2 or to fumigation. These results indicate that cis and trans-β-ocimene may have a different pathway of formation that probably does not involve enzymatic synthesis.

Notes: Abstract. Isoprene (2-methyl 1, 3-butadiene) is emitted from many plants, especially trees. We tested the effect of growth at high CO2 partial pressure and sun versus shade conditions on the capacity of Quercus rubra L. (red oak) and Populus tremuloides Michx. (quaking aspen) leaves to make isoprene. Oak leaves grown at high CO2 partial pressure (65 Pa) had twice the rate of isoprene emission as leaves grown at 40Pa CO2. However, aspen leaves behaved oppositely, with high CO2-grown leaves having just 60-70% the rate of isoprene emission as leaves grown in 40 Pa CO2. Similar responses were observed from 25 to 35 °C leaf temperature during assay. The stimulation of isoprene emission by growth at high CO2 and the stimulation in high temperature resulted in isoprene emission consuming over 15% of the carbon fixed during photosynthesis in high-CO2 grown oak leaves assayed at 35 °C. Leaves from the south (sunny) sides of trees growing in natural conditions had rates of isoprene emission double those of leaves growing in shaded locations on the same trees. This effect was similar in both aspen and oak. The leaves used for these experiments had significantly different chlorophyll a/b ratios indicating they were functionally sun (from the sunny locations) or shade leaves (from the protected locations). Because the metabolic pathway of isoprene synthesis is unknown, we are unable to speculate about how or why these effects occur. However, these effects are more consistent with metabolic control of isoprene release rather than a metabolic leak of isoprene from metabolism. The results are also important for large scale modelling of isoprene emission and for predicting the effect of future increases in atmospheric CO2 level on isoprene emission from vegetation.

Notes: In this study it has been shown that increased diffusional resistances caused by salt stress may be fully overcome by exposing attached leaves to very low [CO2] (∼ 50 µmol  mol−1), and, thus a non-destructive-in vivo method to correctly estimate photosynthetic capacity in stressed plants is reported. Diffusional (i.e. stomatal conductance, gs, and mesophyll conductance to CO2, gm) and biochemical limitations to photosynthesis (A) were measured in two 1-year-old Greek olive cultivars (Chalkidikis and Kerkiras) subjected to salt stress by adding 200 mm NaCl to the irrigation water. Two sets of A–Ci curves were measured. A first set of standard A–Ci curves (i.e. without pre-conditioning plants at low [CO2]), were generated for salt-stressed plants. A second set of A–Ci curves were measured, on both control and salt-stressed plants, after pre-conditioning leaves at [CO2] of ∼ 50 µmol mol−1 for about 1.5 h to force stomatal opening. This forced stomata to be wide open, and gs increased to similar values in control and salt-stressed plants of both cultivars. After gs had approached the maximum value, the A–Ci response was again measured. The analysis of the photosynthetic capacity of the salt-stressed plants based on the standard A–Ci curves, showed low values of the Jmax (maximum rate of electron transport) to Vcmax (RuBP-saturated rate of Rubisco) ratio (1.06), that would implicate a reduced rate of RuBP regeneration, and, thus, a metabolic impairment. However, the analysis of the A–Ci curves made on pre-conditioned leaves, showed that the estimates of the photosynthetic capacity parameters were much higher than in the standard A–Ci responses. Moreover, these values were similar in magnitude to the average values reported by Wullschleger (Journal of Experimental Botany 44, 907–920, 1993) in a survey of 109 C3 species. These findings clearly indicates that: (1) salt stress did affect gs and gm but not the biochemical capacity to assimilate CO2 and therefore, in these conditions, the sum of the diffusional resistances set the limit to photosynthesis rates; (2) there was a linear relationship (r2 = 0.68) between gm and gs, and, thus, changes of gm can be as fast as those of gs; (3) the estimates of photosynthetic capacity based on A–Ci curves made without removing diffusional limitations are artificially low and lead to incorrect interpretations of the actual limitations of photosynthesis; and (4) the analysis of the photosynthetic properties in terms of stomatal and non-stomatal limitations should be replaced by the analysis of diffusional and non-diffusional limitations of photosynthesis. Finally, the C3 photosynthesis model parameterization using in vitro-measured and in vivo-measured kinetics parameters was compared. Applying the in vivo-measured Rubisco kinetics parameters resulted in a better parameterization of the photosynthesis model.

Notes: The long-term effect of elevated atmospheric CO2 on isoprenoid emissions from adult trees of two Mediterranean oak species (the monoterpene-emitting Quercus ilex L. and the isoprene-emitting Quercus pubescens Willd.) native to a high-CO2 environment was investigated. During two consecutive years, isoprenoid emission was monitored both at branch level, measuring the actual emissions under natural conditions, and at leaf level, measuring the basal emissions under the standard conditions of 30 °C and at light intensity of 1000 µmol m−2 s−1. Long-term exposure to high atmospheric levels of CO2 did not significantly affect the actual isoprenoid emissions. However, when leaves of plants grown in the control site were exposed for a short period to an elevated CO2 level by rapidly switching the CO2 concentration in the gas-exchange cuvette, both isoprene and monoterpene basal emissions were clearly inhibited. These results generally confirm the inhibitory effect of elevated CO2 on isoprenoid emission. The absence of a CO2 effect on actual emissions might indicate higher leaf temperature at elevated CO2, or an interaction with multiple stresses some of which (e.g. recurrent droughts) may compensate for the CO2 effect in Mediterranean ecosystems. Under elevated CO2, isoprene emission by Q. pubescens was also uncoupled from the previous day's air temperature. In addition, pronounced daily and seasonal variations of basal emission were observed under elevated CO2 underlining that correction factors may be necessary to improve the realistic estimation of isoprene emissions with empirical algorithms in the future. A positive linear correlation of isoprenoid emission with the photosynthetic electron transport and in particular with its calculated fraction used for isoprenoid synthesis was found. The slope of this relationship was different for isoprene and monoterpenes, but did not change when plants were grown in either ambient or elevated CO2. This suggests that physiological algorithms may usefully predict isoprenoid emission also under rising CO2 levels.

Notes: Isoprene basal emission (the emission of isoprene from leaves exposed to a light intensity of 1000 µmol m−2 s−1 and maintained at a temperature of 30 °C) was measured in Phragmites australis plants growing under elevated CO2 in the Bossoleto CO2 spring at Rapolano Terme, Italy, and under ambient CO2 at a nearby control site. Gas exchange and biochemical measurements were concurrently taken. Isoprene emission was lower in the plants growing at elevated CO2 than in those growing at ambient CO2. Isoprene emission and isoprene synthase activity (IsoS) were very low in plants growing at the bottom of the spring under very rich CO2 and increased at increasing distance from the spring (and decreasing CO2 concentration). Distance from the spring did not significantly affect photosynthesis making it therefore unlikely that there is carbon limitation to isoprene formation. The isoprene emission rate was very quickly reduced after rapid switches from elevated to ambient CO2 in the gas-exchange cuvette, whereas it increased when switching from ambient to elevated CO2. The rapidity of the response may be consistent with post-translational modifications of enzymes in the biosynthetic pathway of isoprene formation. Reduction of IsoS activity is interpreted as a long-term response. Basal emission of isoprene was not constant over the day but showed a diurnal course opposite to photosynthesis, with a peak during the hottest hours of the day, independent of stomatal conductance and probably dependent on external air temperature or temporary reduction of CO2 concentration. The present experiments show that basal emission rate of isoprene is likely to be reduced under future elevated CO2 levels and allow improvement in the modelling of future isoprene emission rates.

Notes: Poplar (Populus × euroamericana) saplings were grown in the field to study the changes of photosynthesis and isoprene emission with leaf ontogeny in response to free air carbon dioxide enrichment (FACE) and soil nutrient availability. Plants growing in elevated [CO2] produced more leaves than those in ambient [CO2]. The rate of leaf expansion was measured by comparing leaves along the plant profile. Leaf expansion and nitrogen concentration per unit of leaf area was similar between nutrient treatment, and this led to similar source–sink functional balance. Consequently, soil nutrient availability did not cause downward acclimation of photosynthetic capacity in elevated [CO2] and did not affect isoprene synthesis. Photosynthesis assessed in growth [CO2] was higher in plants growing in elevated than in ambient [CO2]. After normalizing for the different number of leaves over the profile, maximal photosynthesis was reached and started to decline earlier in elevated than in ambient [CO2]. This may indicate a [CO2]-driven acceleration of leaf maturity and senescence. Isoprene emission was adversely affected by elevated [CO2]. When measured on the different leaves of the profile, isoprene peak emission was higher and was reached earlier in ambient than in elevated [CO2]. However, a larger number of leaves was emitting isoprene in plant growing in elevated [CO2]. When integrating over the plant profile, emissions in the two [CO2] levels were not different. Normalization as for photosynthesis showed that profiles of isoprene emission were remarkably similar in the two [CO2] levels, with peak emissions at the centre of the profile. Only the rate of increase of the emission of young leaves may have been faster in elevated than in ambient [CO2]. Our results indicate that elevated [CO2] may overall have a limited effect on isoprene emission from young seedlings and that plants generally regulate the emission to reach the maximum at the centre of the leaf profile, irrespective of the total leaf number. In comparison with leaf expansion and photosynthesis, isoprene showed marked and repeatable differences among leaves of the profile and may therefore be a useful trait to accurately monitor changes of leaf ontogeny as a consequence of elevated [CO2].

Notes: Isoprene is the primary biogenic hydrocarbon emitted from temperate deciduous forest ecosystems. The effects of varying photon flux density (PFD) and nitrogen growth regimes on rates of isoprene emission and net photosynthesis in potted aspen and white oak trees are reported. In both aspen and oak trees, whether rates were expressed on a leaf area or dry mass basis, (1) growth at higher PFD resulted in significantly higher rates of isoprene emission, than growth at lower PFD, (2) there is a significant positive relationship between isoprene emission rate and leaf nitrogen concentration in both sun and shade trees, and (3) there is a significant positive correlation between isoprene emission rate and photosynthetic rate in both sun and shade trees. The greater capacity for isoprene emission in sun leaves was due to both higher leaf mass per unit area and differences in the biochemical and/or physiological properties that influence isoprene emission. Positive correlations between isoprene emission rate and leaf nitrogen concentration support the existence of mechanisms that link leaf nitrogen status to isoprene synthase activity. Positive correlations between isoprene emission rate and photosynthesis rate support previous hypotheses that isoprene emission plays a role in protecting photosynthetic mechanisms during stress.

Notes: α-Pinene is formed in and emitted by Quercus ilex leaves. The carbon emitted as α-pinene is rapidly and totally labelled by 13C in CO2 in air, but α-pinene contained in the leaf shows a fraction of completely unlabelled carbon even after long exposures to air containing only 13CO2. When the labelled leaf is darkened, α-pinene emission drops but is still measurable for about 10 h, and carbon becomes partially unlabelled. After an 11 h darkening the α-pinene content is still as high as in the light but the carbon is mostly unlabelled. If the leaf is re-illuminated but photosynthesis is inhibited by removing CO2 and lowering O2, a burst of emission occurs and the content of α-pinene is depleted. Our experiments suggest that a pool of α-pinene which is not directly generated by photosynthesis intermediates exists. Since this pool does not contribute relevantly to the emission in the light, we hypothesize that it is inhibited in the light and spatially located in a different compartment from chloroplasts. As we discuss, glycolysis in the cytoplasm and leucine catabolism in the mitochondria are both possible extra-chloroplasts sources of carbon for isoprenoids.