Reactions to cadmium stress in a cadmium-tolerant variety of cabbage (Brassica oleracea L.): is cadmium tolerance necessarily desirable in food crops?

Cadmium is a cumulative, chronic toxicant in humans for which the main exposure pathway is via plant foods. Cadmium-tolerant plants may be used to create healthier food products, provided that the tolerance is associated with the exclusion of Cd from the edible portion of the plant. An earlier study identified the cabbage (Brassica oleracea L.) variety, Pluto, as relatively Cd tolerant. We exposed the roots of intact, 4-week-old seedlings of Pluto to Cd (control ∼1 mg L(-1) treatment 500 µg L(-1)) for 4 weeks in flowing nutrient solutions and observed plant responses. Exposure began when leaf 3 started to emerge, plants were harvested after 4 weeks of Cd exposure and the high Cd treatment affected all measured parameters. The elongation rate of leaves 4-8, but not the duration of elongation was reduced; consequently, individual leaf area was also reduced (P < 0.001) and total leaf area and dry weight were approximately halved. A/C i curves immediately before harvest showed that Cd depressed the photosynthetic capacity of the last fully expanded leaf (leaf 5). Despite such large impairments of the source and sink capacities, specific leaf weight and the partitioning of photosynthate between roots, stems and leaves were unaffected (P > 0.1). Phytochelatins (PCs) and glutathione (GSH) were present in the roots even at the lowest Cd concentration in the nutrient medium, i.e. ∼1 µg Cd L(-1), which would not be considered contaminated if it were a soil solution. The Cd concentration in these roots was unexpectedly high (5 mg kg(-1) DW) and the molar ratio of -SH (in PCs plus GSH) to Cd was large (>100:1). In these control plants, the Cd concentration in the leaves was 1.1 mg kg(-1) DW, and PCs were undetectable. For the high Cd treatment, the concentration of Cd in roots exceeded 680 mg kg(-1) DW and the molar -SH to Cd ratio fell to ∼1.5:1. For these plants, Cd flooded into the leaves (107 mg kg(-1) DW) where it probably induced synthesis of PCs, and the molar -SH to Cd ratio was ∼3:1. Nonetheless, this was insufficient to sequester all the Cd, as evidenced by the toxic effects on photosynthesis and growth noted above. Lastly, Cd accumulation in the leaves was associated with lowered concentrations of some trace elements, such as Zn, a combination of traits that is highly undesirable in food plants.

Photosynthetic responses of two eucalypts to industrial-age changes in atmospheric [CO2] and temperature.

The unabated rise in atmospheric [CO(2)] is associated with increased air temperature. Yet, few CO(2)-enrichment studies have considered pre-industrial [CO(2)] or warming. Consequently, we quantified the interactive effects of growth [CO(2)] and temperature on photosynthesis of faster-growing Eucalyptus saligna and slower-growing E. sideroxylon. Well-watered and -fertilized tree seedlings were grown in a glasshouse at three atmospheric [CO(2)] (290, 400, and 650 µL L(-1)), and ambient (26/18 °C, day/night) and high (ambient + 4 °C) air temperature. Despite differences in growth rate, both eucalypts responded similarly to [CO(2)] and temperature treatments with few interactive effects. Light-saturated photosynthesis (A(sat)) and light- and [CO(2)]-saturated photosynthesis (A(max) ) increased by ∼ 50% and ∼ 10%, respectively, with each step-increase in growth [CO(2)], underpinned by a corresponding 6-11% up-regulation of maximal electron transport rate (J(max)). Maximal carboxylation rate (V(cmax)) was not affected by growth [CO(2)]. Thermal photosynthetic acclimation occurred such that A(sat) and A(max) were similar in ambient- and high-temperature-grown plants. At high temperature, the thermal optimum of A(sat) increased by 2-7 °C across [CO(2)] treatments. These results are the first to suggest that photosynthesis of well-watered and -fertilized eucalypt seedlings will remain strongly responsive to increasing atmospheric [CO(2)] in a future, warmer climate.

The sensitivity of photosynthesis to phosphorus deficiency differs between C3 and C4 tropical grasses.

Phosphorus (P) is an important determinant of plant productivity, particularly in the tropical grasslands of Australia, which contain both C3 and C4 species. Few studies have compared the responses of such species to P deficiency. Previous work led us to hypothesise that C3 photosynthesis and the three subtypes of C4 photosynthesis have different sensitivities to P deficiency. To examine their dynamic response to P deficiency in more detail, four taxonomically related tropical grasses (Panicum laxum (C3) and Panicum coloratum, Cenchrus ciliaris and Panicum maximum belonging to the C4 subtypes NAD-ME, NADP-ME and PCK, respectively) were grown under contrasting P supplies, including P withdrawal from the growing medium. Changes in photosynthesis and growth were compared with leaf carbohydrate contents and metabolic fingerprints obtained using high-resolution proton nuclear magnetic resonance (1H-NMR). The response of CO2 assimilation rates to leaf contents of inorganic phosphate ([Pi]) was linear in the C3 grass, but asymptotic for the three C4 grasses. Relative growth rate was affected most by low P in the C3 species and was correlated with the leaf content of glucose 6-phosphate more than with carbohydrates. Principal component analysis of the 1H-NMR spectra revealed distinctive profiles of carbohydrates and amino acids for the four species. Overall, the data showed that photosynthesis of the three C4 subtypes behaved similarly. Compared with the C3 counterpart, photosynthesis of the three C4 grasses had a higher P use efficiency and lower Pi requirement, and responded to a narrower range of [Pi]. Although each of the four grass species showed distinctive 1H-NMR fingerprints, there were no differences in response that could be attributed to the C4 subtypes.

Canopy development and hydraulic function in Eucalyptus tereticornis grown in drought in CO2-enriched atmospheres.

We report on the relationship between growth, partitioning of shoot biomass and hydraulic development of Eucalyptus tereticornis Sm. grown in glasshouses for six months. Close coordination of stem vascular capacity and shoot architecture is vital for survival of eucalypts, especially as developing trees are increasingly subjected to spasmodic droughts and rising atmospheric CO2 levels. Trees were exposed to constant soil moisture deficits in 45 L pots (30-50% below field capacity), while atmospheric CO2 was raised to 700 µL CO2 L-1 in matched glasshouses using a hierarchical, multi-factorial design. Enrichment with CO2 stimulated shoot growth rates for 12-15 weeks in well-watered trees but after six months of CO2 enrichment, shoot biomasses were not significantly heavier (30% stimulation) in ambient conditions. By contrast, constant drought arrested shoot growth after 20 weeks under ambient conditions, whereas elevated CO2 sustained growth in drought and ultimately doubled the shoot biomass relative to ambient conditions. These growth responses were achieved through an enhancement of lateral branching up to 8-fold due to CO2 enrichment. In spite of larger transpiring canopies, CO2 enrichment also improved the daytime water status of leaves of droughted trees. Stem xylem development was highly regulated, with vessels per unit area and cross sectional area of xylem vessels in stems correlated inversely across all treatments. Furthermore, vessel numbers related to the numbers of leaves on lateral branches, broadly supporting predictions arising from Pipe Model Theory that the area of conducting tissue should correlate with leaf area. Diminished water use of trees in drought coincided with a population of narrower xylem vessels, constraining hydraulic capacity of stems. Commensurate with the positive effects of elevated CO2 on growth, development and leaf water relations of droughted trees, the capacity for long-distance water transport also increased.

Phosphorus deficiency inhibits growth in parallel with photosynthesis in a C3 (Panicum laxum) but not two C4 (P. coloratum and Cenchrus ciliaris) grasses.

This study compared the growth and photosynthetic responses of one C3 (Panicum laxum L.) and two C4 grasses (Panicum coloratum L. and Cenchrus ciliaris L.) to changes in soil phosphorus (P) nutrition. Plants were grown in potted soil amended with six different concentrations of P. One week before harvest, leaf elongation and photosynthetic rates and the contents of carbohydrate, P and inorganic phosphate (Pi) were measured. Five weeks after germination, plants were harvested to estimate biomass accumulation. At each soil P supply, leaf P contents were lower in the C3 (0.6-2.6 mmol P m-2) than in the two C4 grasses (0.8-4.1 mmol P m-2), and Pi constituted ~40-65% of total leaf P. The P deficiency reduced leaf growth, tillering and plant dry mass to a similar extent in all three grasses. In contrast, P deficiency suppressed photosynthetic rates to a greater extent in the C3 (50%) than the C4 grasses (25%). The foliar contents of non-structural carbohydrates were affected only slightly by soil P supply in all three species. Leaf mass per area decreased at low P in the two C4 grasses only, and biomass partitioning changed little with soil P supply. The percentage changes in assimilation rates and plant dry mass were linearly related in the C3 but not the C4 plants. Thus, P deficiency reduced growth in parallel with reductions of photosynthesis in the C3 grass, and independently of photosynthesis in the two C4 grasses. We propose that this may be related to a greater Pi requirement of C4 relative to C3 photosynthesis. Photosynthetic P use efficiency was greater and increased more with P deficiency in the C4 relative to the C3 species. The opposite was observed for whole-plant P-use efficiency. Hence, the greater P-use efficiency of C4 photosynthesis was not transferred to the whole-plant level, mainly as a result of the larger and constant leaf P fraction in the two C4 grasses.

Faster Rubisco is the key to superior nitrogen-use efficiency in NADP-malic enzyme relative to NAD-malic enzyme C4 grasses.

In 27 C4 grasses grown under adequate or deficient nitrogen (N) supplies, N-use efficiency at the photosynthetic (assimilation rate per unit leaf N) and whole-plant (dry mass per total leaf N) level was greater in NADP-malic enzyme (ME) than NAD-ME species. This was due to lower N content in NADP-ME than NAD-ME leaves because neither assimilation rates nor plant dry mass differed significantly between the two C4 subtypes. Relative to NAD-ME, NADP-ME leaves had greater in vivo (assimilation rate per Rubisco catalytic sites) and in vitro Rubisco turnover rates (k(cat); 3.8 versus 5.7 s(-1) at 25 degrees C). The two parameters were linearly related. In 2 NAD-ME (Panicum miliaceum and Panicum coloratum) and 2 NADP-ME (Sorghum bicolor and Cenchrus ciliaris) grasses, 30% of leaf N was allocated to thylakoids and 5% to 9% to amino acids and nitrate. Soluble protein represented a smaller fraction of leaf N in NADP-ME (41%) than in NAD-ME (53%) leaves, of which Rubisco accounted for one-seventh. Soluble protein averaged 7 and 10 g (mmol chlorophyll)(-1) in NADP-ME and NAD-ME leaves, respectively. The majority (65%) of leaf N and chlorophyll was found in the mesophyll of NADP-ME and bundle sheath of NAD-ME leaves. The mesophyll-bundle sheath distribution of functional thylakoid complexes (photosystems I and II and cytochrome f) varied among species, with a tendency to be mostly located in the mesophyll. In conclusion, superior N-use efficiency of NADP-ME relative to NAD-ME grasses was achieved with less leaf N, soluble protein, and Rubisco having a faster k(cat).

Wood density and anatomy of water-limited eucalypts.

We hypothesized that seedlings grown under water-limited conditions would develop denser wood than seedlings grown under well-watered conditions. Three Eucalyptus species (E. grandis Hill (ex Maiden), E. sideroxylon Cunn. (ex Woolls) and E. occidentalis Endl.) were grown in a temperature-controlled greenhouse for 19 weeks with watering treatments (well-watered and water-limited) applied at six weeks. The water-limitation treatment consisted of four drought cycles. Wood density increased by between 4 and 13% in the water-limited seedlings, but this increase was mainly due to extractive compounds embedded in the cell wall matrix. Once these compounds were removed, the increase was 0-9% and was significant for E. grandis only. Water-limitation significantly reduced mean vessel lumen area; however, this was balanced by a trend toward greater vessel frequency in water-limited plants, and consequently there was no difference in the proportion of stem area allocated to vessels. Conduit efficiency value was lowest in the water-limited plants, indicating that there was a cost in terms of stem hydraulic conductivity for decreasing vessel lumen area. Wood density was negatively correlated with vessel lumen fraction in well-watered plants, but this relationship broke down in the water-limited plants, possibly because of the significantly larger proportion of the stem taken up by pith in water-limited seedlings. Diurnal variation in leaf water potential was positively correlated with wood density in well-watered plants. This relationship did not hold in the water-limited plants owing to the collapse of the pressure gradient between soil and leaf. We conclude that drought periods of greater than 1 month are required to increase wood density in these species and that increases in wood density appear to result in diminished capacity to supply water to leaves.

Leaf water use efficiency differs between Eucalyptus seedlings from contrasting rainfall environments.

This study investigates the putative role of thicker leaves in enhancing photosynthetic capacity and water-use efficiency (WUE) of Eucalyptus species native to xeric environments. Three Eucalyptus species, Eucalyptus grandis Hill. (ex Maiden), E. sideroxylon Cunn. (ex Woolls) and E. occidentalis (Endl.), were grown under well-watered or water-limited conditions in a single compartment of a temperature-controlled glasshouse. Eucalyptus grandis is native to a mesic environment while E. sideroxylon and E. occidentalis are native to xeric environments. Leaves of E. sideroxylon and E. occidentalis were thicker and contained more nitrogen (N) on a leaf-area basis than E. grandis. Leaf gas-exchange measurements indicated that the photosynthetic capacity of E. sideroxylon and E.occidentalis was greater than E. grandis and that stomatal conductance and WUE were negatively correlated. Whole-plant, gas-exchange and carbon-isotope measurements showed that E. sideroxylon and E. occidentalis had lower WUE than E. grandis under both well-watered and water-limited conditions. However, there was no difference in N-use efficiency between species. We suggest that stomatal conductance and leaf N content are functionally linked in these seedlings and conclude that thick leaves can, in some conditions, result in low WUE.

Genetic modification of photosynthesis with E. coli genes for trehalose synthesis.

Improvement in photosynthesis per unit leaf area has been difficult to alter by breeding or genetic modification. We report large changes in photosynthesis in Nicotiana tabacum transformed with E. coli genes for the trehalose pathway. Significantly, photosynthetic capacity (CO2 assimilation at varying light and CO2, and quantum yield of PSII electron transport) per unit leaf area and per leaf dry weight were increased in lines of N. tabacum transformed with the E. coli gene otsA, which encodes trehalose phosphate synthase. In contrast, transformation with otsB, which encodes trehalose phosphate phosphatase or Trec, encoding trehalose phosphate hydrolase, produced the opposite effect. Changes in CO2 assimilation per unit leaf area were closely related to the amount and activity of Rubisco, but not to the maximum activities of other Calvin cycle enzymes. Alterations in photosynthesis were associated with trehalose 6-phosphate content rather than trehalose. When growth parameters were determined, a greater photosynthetic capacity did not translate into greater relative growth rate or biomass. This was because photosynthetic capacity was negatively related to leaf area and leaf area ratio. In contrast, relative growth rate and biomass were positively related to leaf area. These results demonstrate a novel means of modifying Rubisco content and photosynthesis, and the complexities of regulation of photosynthesis at the whole plant level, with potential benefits to biomass production through improved leaf area.

Nonstomatal limitations are responsible for drought-induced photosynthetic inhibition in four C4 grasses.

• Here, the contribution of stomatal and nonstomatal factors to photosynthetic inhibition under water stress in four tropical C4 grasses was investigated (Panicum coloratum, Bothriochloa bladhii, Cenchrus ciliaris and Astrebla lappacea). • Plants were grown in well watered soil, and then the effects of soil drying were measured on leaf gas exchange, chlorophyll a fluorescence and water relations. • During the drying cycle, leaf water potential (Ψleaf ) and relative water content (RWC) decreased from c. -0.4 to -2.8 MPa and 100-40%, respectively. The CO2 assimilation rates (A) and quantum yield of PSII (ΦPSII ) of all four grasses decreased rapidly with declining RWC. High CO2 concentration (2500 µl l-1 ) had no effect on A or ΦPSII at any stage of the drying cycle. Electron transport capacity and dark respiration rates were unaltered by drought. The CO2 compensation concentrations of P. coloratum and C. ciliaris rose sharply when leaf RWC fell below 70%. In P. coloratum, 5% CO2 did not prevent the decline of O2 evolution rates under water stress. • We conclude that inhibition of photosynthesis in the four C4 grasses under water stress is dependent mainly on biochemical limitations.

The effect of drought on plant water use efficiency of nine NAD-ME and nine NADP-ME Australian C4 grasses.

We investigated the response to drought of nine NAD-malic enzyme (NAD-ME) and nine NADP-malic enzyme (NADP-ME) C4 grasses. Species were grown from seeds in potted soil in a glasshouse. Seedlings were either watered regularly or exposed to two successive drying cycles of 8-10 d each, after which plants were harvested. Under well-watered conditions, average water use efficiency (WUE; dry mass gain per unit water transpired) was similar for NAD-ME and NADP-ME C4 grasses, and ranged between 6.0 and 8.7 g dry mass kg-1 H2O. Drought enhanced WUE of most species, but to a significantly greater extent in NAD-ME (1.20-fold) than NADP-ME (1.11-fold) grasses. Inhibition of dry matter accumulation (average of 12%) and shoot elongation under drought was similar among the C4 grasses. Leaf dry matter carbon (δ13C) and oxygen (δ18O) isotope compositions were significantly different between the two C4 subtypes. Leaf δ13C averaged -13.3 and -12.2, and leaf δ18O averaged 26.0 and 26.9 in well-watered NAD-ME and NADP-ME grasses, respectively. Drought significantly reduced leaf δ13C in most C4 grasses by an average 0.5. Leaf δ18O was not significantly affected by drought, indicating that leaf δ18O does not reflect drought-induced changes in leaf transpiration of C4 grasses. In the experiment reported here, NAD-ME grasses increased their WUE under drought to a greater extent than their NADP-ME counterparts. Increased WUE of the C4 grasses under drought was primarily related to control of water loss relative to carbon gain at the leaf, rather than the plant, level.

Elevated CO2 increases the leaf temperature of two glasshouse-grown C4 grasses.

This study investigates the effect of elevated CO2 partial pressure (pCO2)-induced stomatal closure on leaf temperature and gas exchange of C4 grasses. Two native Australian C4 grasses, Astrebla lappacea (Lindl.) Domin and Bothriochloa bladhii Kuntze, were grown at three different pCO2 (35, 70 and 120 Pa) in three matched, temperature-controlled glasshouse compartments. The difference between leaf and air temperature (ΔT) was monitored diurnally with thermocouples. ΔT increased with both step-increases of ambient pCO2. Average noon leaf temperature increased by 0.4 and 0.3°C for A. lappacea with the 35-70 and 70-120 Pa steps of pCO2 elevation, respectively. For B. bladhii, the increases were 0.5°C for both pCO2 steps. ΔT was strongly dependent on irradiance, pCO2 and air humidity. Leaf gas exchange was measured at constant temperature and high irradiance at the three growth pCO2. Under these conditions, CO2 assimilation saturated at 70 Pa, while stomatal conductance decreased by the same extent (0.58-fold) with both step-increases in pCO2, suggesting that whole-plant water use efficiency of C4 grasses would increase beyond a doubling of ambient pCO2. The ratio of intercellular to ambient pCO2 was not affected by short- or long-term doubling or near-tripling of pCO2, in either C4 species when measured under standard conditions.

Changes in sourcesink relations during development influence photosynthetic acclimation of rice to free air CO2 enrichment (FACE).

Relationships between photosynthetic acclimation and changes in the balance between source-sink supply and demand of carbon (C) and nitrogen (N) were tested using rice (Oryza sativa L. cv. Akitakomachi). Plants were field-grown in northern Japan at ambient CO2 partial pressure [p(CO2)] or free air CO2 enrichment (FACE; p(CO2) ~ 26-32 Pa above ambient) with low, medium or high N supplies. Leaf CO2 assimilation rates (A) and biochemical parameters were measured at 32-36 (eighth leaf) and 76-80 (flag leaf) d after transplanting, representing stages with a contrasting balance between C and N supply and demand in sources and sinks. Acclimation due to FACE was pronounced in flag leaves at each N supply. This was not fully accounted for by reductions in leaf N concentrations, because A/N and Vcmax/N were lower in FACE-grown flag leaves. Acclimation did not occur in the eighth leaf, and A/N and Vcmax/N was not significantly increased in FACE-grown leaves. Soluble protein / sucrose and amino acid / sucrose concentrations decreased under FACE, whereas sucrose phosphate synthase protein levels increased. At flag leaf stage, there was a discrepancy between the demand and supply of N, which was resolved by enhanced leaf N remobilization, associated with the lower Rubisco concentrations under FACE. In contrast to the early growth stage, enhanced growth of rice plants was accompanied by increased plant N uptake in FACE. We conclude that photosynthetic acclimation in flag leaves occurs under FACE because there is a large demand for N for reproductive development, relative to supply of N from root uptake and remobilization from leaves.