The importance of species-specific and temperature-sensitive parameterisation of A/Ci models: A case study using cotton (Gossypium hirsutum L.) and the automated 'OptiFitACi' R-package.

Leaf gas exchange measurements are an important tool for inferring a plant's photosynthetic biochemistry. In most cases, the responses of photosynthetic CO2 assimilation to variable intercellular CO2 concentrations (A/Ci response curves) are used to model the maximum (potential) rate of carboxylation by ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco, Vcmax) and the rate of photosynthetic electron transport at a given incident photosynthetically active radiation flux density (PAR; JPAR). The standard Farquhar-von Caemmerer-Berry model is often used with default parameters of Rubisco kinetic values and mesophyll conductance to CO2 (gm) derived from tobacco that may be inapplicable across species. To study the significance of using such parameters for other species, here we measured the temperature responses of key in vitro Rubisco catalytic properties and gm in cotton (Gossypium hirsutum cv. Sicot 71) and derived Vcmax and J2000 (JPAR at 2000 µmol m-2 s-1 PAR) from cotton A/Ci curves incrementally measured at 15°C-40°C using cotton and other species-specific sets of input parameters with our new automated fitting R package 'OptiFitACi'. Notably, parameterisation by a set of tobacco parameters produced unrealistic J2000:Vcmax ratio of <1 at 25°C, two- to three-fold higher estimates of Vcmax above 15°C, up to 2.3-fold higher estimates of J2000 and more variable estimates of Vcmax and J2000, for our cotton data compared to model parameterisation with cotton-derived values. We determined that errors arise when using a gm,25 of 2.3 mol m-2 s-1 MPa-1 or less and Rubisco CO2-affinities in 21% O2 (KC 21%O2) at 25°C outside the range of 46-63 Pa to model A/Ci responses in cotton. We show how the A/Ci modelling capabilities of 'OptiFitACi' serves as a robust, user-friendly, and flexible extension of 'plantecophys' by providing simplified temperature-sensitivity and species-specificity parameterisation capabilities to reduce variability when modelling Vcmax and J2000.

Estimating leaf day respiration from conventional gas exchange measurements.

Leaf day respiration (Rd ) strongly influences carbon-use efficiencies of whole plants and the global terrestrial biosphere. It has long been thought that Rd is slower than respiration in the dark at a given temperature, but measuring Rd by gas exchange remains a challenge because leaves in the light are also photosynthesizing. The Kok method and the Laisk method are widely used to estimate Rd . We highlight theoretical limitations of these popular methods, and recent progress toward their improvement by using additional information from chlorophyll fluorescence and by accounting for the photosynthetic reassimilation of respired CO2 . The latest evidence for daytime CO2 and energy release from the oxidative pentose phosphate pathway in chloroplasts appears to be important to understanding Rd .

Running the numbers on plant synthetic biology solutions to global problems.

Synthetic biology and metabolic engineering promise to deliver sustainable solutions to global problems such as phasing out fossil fuels and replacing industrial nitrogen fixation. While this promise is real, scale matters, and so do knock-on effects of implementing solutions. Both scale and knock-on effects can be estimated by 'Fermi calculations' (aka 'back-of-envelope calculations') that use uncontroversial input data plus simple arithmetic to reach rough but reliable conclusions. Here, we illustrate how this is done and how informative it can be using two cases: oilcane (sugarcane engineered to accumulate triglycerides instead of sugar) as a source of bio-jet fuel, and nitrogen fixation by bacteria in mucilage secreted by maize aerial roots. We estimate that oilcane could meet no more than about 1% of today's U.S. jet fuel demand if grown on all current U.S. sugarcane land and that, if cane land were expanded to meet two-thirds of this demand, the fertilizer and refinery requirements would create a large carbon footprint. Conversely, we estimate that nitrogen fixation in aerial-root mucilage could replace up to 10% of the fertilizer nitrogen applied to U.S. maize, that 2% of plant carbon income used for growth would suffice to fuel the fixation, and that this extra carbon consumption would likely reduce grain yield only slightly.

Engineering Strategies to Boost Crop Productivity by Cutting Respiratory Carbon Loss.

Roughly half the carbon that crop plants fix by photosynthesis is subsequently lost by respiration. Nonessential respiratory activity leading to unnecessary CO2 release is unlikely to have been minimized by natural selection or crop breeding, and cutting this large loss could complement and reinforce the currently dominant yield-enhancement strategy of increasing carbon fixation. Until now, however, respiratory carbon losses have generally been overlooked by metabolic engineers and synthetic biologists because specific target genes have been elusive. We argue that recent advances are at last pinpointing individual enzyme and transporter genes that can be engineered to (1) slow unnecessary protein turnover, (2) replace, relocate, or reschedule metabolic activities, (3) suppress futile cycles, and (4) make ion transport more efficient, all of which can reduce respiratory costs. We identify a set of engineering strategies to reduce respiratory carbon loss that are now feasible and model how implementing these strategies singly or in tandem could lead to substantial gains in crop productivity.

Redesigning thiamin synthesis: Prospects and potential payoffs.

Thiamin is essential for plant growth but is short-lived in vivo and energetically very costly to produce - a combination that makes thiamin biosynthesis a prime target for improvement by redesign. Thiamin consists of thiazole and pyrimidine moieties. Its high biosynthetic cost stems from use of the suicide enzyme THI4 to form the thiazole and the near-suicide enzyme THIC to form the pyrimidine. These energetic costs lower biomass yield potential and are likely compounded by environmental stresses that destroy thiamin and hence increase the rate at which it must be made. The energy costs could be slashed by refactoring the thiamin biosynthesis pathway to eliminate the suicidal THI4 and THIC reactions. To substantiate this design concept, we first document the energetic costs of the THI4 and THIC steps in the pathway and explain how cutting these costs could substantially increase crop biomass and grain yields. We then show that a refactored pathway must produce thiamin itself rather than a stripped-down analog because the thiamin molecule cannot be simplified without losing biological activity. Lastly, we consider possible energy-efficient alternatives to the inefficient natural THI4- and THIC-mediated steps.

Root growth and anchorage by transplanted 'Tifgreen' (Cynodon dactylon x C. transvaalensis) turfgrass.

Field experiments quantified factors affecting root growth and anchorage by transplanted 'Tifgreen' (Cynodon dactylon (L.) Pers.×Cynodon transvaalensis Burtt Davy) sod, a globally important warm-season C4 turfgrass. Vertical force required to detach recently transplanted sod from underlying soil was the measure of root anchoring strength. In early spring, date of sod harvest and transplantation was important to root growth and anchorage measured 30 days after transplantation. Delaying sod harvest/transplantation by about a month after the end of the winter shoot dormancy period increased root anchoring strength 200% and root dry mass 640% during the 30 days after sodding. The strong effect of early-spring sodding date on root anchorage was related to cumulative thermal time before sod harvesting. Root anchoring strength was directly proportional to the number, but not mass, of roots produced by transplanted sod. In late spring, anchoring of sod to very firm traffic-compacted clay was 87% greater than to loamy sand, measured 14 days after sodding. N-P-K fertilisation did not affect late-spring sod anchorage to loamy sand soil, measured 18 days after sodding, but did enhance shoot density and colour. Sod root penetration into a silt loam soil was unaffected by an initially dry surface layer when sufficient irrigation was used. Overall, root anchorage by transplanted Tifgreen sod was similar to, or greater than, values reported for cool-season C3 turfgrasses in similar circumstances.

Variation in mesophyll conductance among Australian wheat genotypes.

CO2 diffusion from substomatal intercellular cavities to sites of carboxylation in chloroplasts (mesophyll conductance; gm) limits photosynthetic rate and influences leaf intrinsic water-use efficiency (A/gsw). We investigated genotypic variability of gm and effects of gm on A/gsw among eleven wheat (Triticum aestivum L.) genotypes under light-saturated conditions and at either 2 or 21% O2. Significant variation in gm and A/gsw was found between genotypes at both O2 concentrations, but there was no significant effect of O2 concentration on gm. Further, gm was correlated with photosynthetic rate among the 11 genotypes, but was unrelated to stomatal conductance. The effect of leaf age differed between genotypes, with gm being lower in older leaves for one genotype but not another. This study demonstrates a high level of variation in gm between wheat genotypes; 0.5 to 1.0µmolm-2s-1 bar-1. Further, leaf age effects indicate that great care must be taken to choose suitable leaves in studies of genotypic variation in gm and water-use efficiency.

Carbon dynamics of eucalypt seedlings exposed to progressive drought in elevated [CO2] and elevated temperature.

Elevated [CO2] and temperature may alter the drought responses of tree seedling growth, photosynthesis, respiration and total non-structural carbohydrate (TNC) status depending on drought intensity and duration. Few studies have addressed these important climatic interactions or their consequences. We grew Eucalyptus globulus Labill. seedlings in two [CO2] concentrations (400 and 640 µl l(-1)) and two temperatures (28/17 and 32/21 °C) (day/night) in a sun-lit glasshouse, and grew them in well-watered conditions or exposed them to two drought treatments having undergone different previous water conditions (i.e., rewatered drought and sustained drought). Progressive drought in both drought treatments led to similar limitations in growth, photosynthesis and respiration, but reductions in TNC concentration were not observed. Elevated [CO2] ameliorated the impact of the drought during the moderate drought phase (i.e., Day 63 to Day 79) by increasing photosynthesis and enhancing leaf and whole-plant TNC content. In contrast, elevated temperature exacerbated the impact of the drought during the moderate drought phase by reducing photosynthesis, increasing leaf respiration and decreasing whole-plant TNC content. Extreme drought (i.e., Day 79 to Day 103) eliminated [CO2] and temperature effects on plant growth, photosynthesis and respiration. The combined effects of elevated [CO2] and elevated temperature on moderate drought stressed seedlings were reduced with progressive drought, with no sustained effects on growth despite greater whole-plant TNC content.

Sensitivity of plants to changing atmospheric CO2 concentration: from the geological past to the next century.

The rate of CO(2) assimilation by plants is directly influenced by the concentration of CO(2) in the atmosphere, c(a). As an environmental variable, c(a) also has a unique global and historic significance. Although relatively stable and uniform in the short term, global c(a) has varied substantially on the timescale of thousands to millions of years, and currently is increasing at seemingly an unprecedented rate. This may exert profound impacts on both climate and plant function. Here we utilise extensive datasets and models to develop an integrated, multi-scale assessment of the impact of changing c(a) on plant carbon dioxide uptake and water use. We find that, overall, the sensitivity of plants to rising or falling c(a) is qualitatively similar across all scales considered. It is characterised by an adaptive feedback response that tends to maintain 1 - c(i)/c(a), the relative gradient for CO(2) diffusion into the leaf, relatively constant. This is achieved through predictable adjustments to stomatal anatomy and chloroplast biochemistry. Importantly, the long-term response to changing c(a) can be described by simple equations rooted in the formulation of more commonly studied short-term responses.

From sunlight to phytomass: on the potential efficiency of converting solar radiation to phyto-energy.

The relationship between solar radiation capture and potential plant growth is of theoretical and practical importance. The key processes constraining the transduction of solar radiation into phyto-energy (i.e. free energy in phytomass) were reviewed to estimate potential solar-energy-use efficiency. Specifically, the out-put:input stoichiometries of photosynthesis and photorespiration in C(3) and C(4) systems, mobilization and translocation of photosynthate, and biosynthesis of major plant biochemical constituents were evaluated. The maintenance requirement, an area of important uncertainty, was also considered. For a hypothetical C(3) grain crop with a full canopy at 30°C and 350 ppm atmospheric [CO(2) ], theoretically potential efficiencies (based on extant plant metabolic reactions and pathways) were estimated at c. 0.041 J J(-1) incident total solar radiation, and c. 0.092 J J(-1) absorbed photosynthetically active radiation (PAR). At 20°C, the calculated potential efficiencies increased to 0.053 and 0.118 J J(-1) (incident total radiation and absorbed PAR, respectively). Estimates for a hypothetical C(4) cereal were c. 0.051 and c. 0.114 J J(-1), respectively. These values, which cannot be considered as precise, are less than some previous estimates, and the reasons for the differences are considered. Field-based data indicate that exceptional crops may attain a significant fraction of potential efficiency.

Efficiency of lignin biosynthesis: a quantitative analysis.

Lignin is derived mainly from three alcohol monomers: p-coumaryl alcohol, coniferyl alcohol and sinapyl alcohol. Biochemical reactions probably responsible for synthesizing these three monomers from sucrose, and then polymerizing the monomers into lignin, were analysed to estimate the amount of sucrose required to produce a unit of lignin. Included in the calculations were amounts of respiration required to provide NADPH (from NADP(+)) and ATP (from ADP) for lignin biosynthesis. Two pathways in the middle stage of monomer biosynthesis were considered: one via tyrosine (found in monocots) and the other via phenylalanine (found in all plants). If lignin biosynthesis proceeds with high efficiency via tyrosine, 76.9, 70.4 and 64.3 % of the carbon in sucrose can be retained in the fraction of lignin derived from p-coumaryl alcohol, coniferyl alcohol and sinapyl alcohol, respectively. The corresponding carbon retention values for lignin biosynthesis via phenylalanine are less, at 73.2, 65.7 and 60.7 %, respectively. Energy (i.e. heat of combustion) retention during lignin biosynthesis via tyrosine could be as high as 81.6, 74.5 and 67.8 % for lignin derived from p-coumaryl alcohol, coniferyl alcohol and sinapyl alcohol, respectively, with the corresponding potential energy retention values for lignin biosynthesis via phenylalanine being less, at 77.7, 69.5 and 63.9 %, respectively. Whether maximum efficiency occurs in situ is unclear, but these values are targets that can be considered in: (1) plant breeding programmes aimed at maximizing carbon or energy retention from photosynthate; (2) analyses of (minimum) metabolic costs of responding to environmental change or pest attack involving increased lignin biosynthesis; (3) understanding costs of lignification in older tissues; and (4) interpreting carbon balance measurements of organs and plants with large lignin concentrations.

Direct effect of elevated CO(2) on nocturnal in situ leaf respiration in nine temperate deciduous tree species is small.

Direct (i.e., short-term) effects of elevated CO(2) on nocturnal in situ leaf respiration rate were measured in nine deciduous tree species (seven genera) in 20 3.5-4.0-h experiments. During the experiments, CO(2) concentration was alternated between 400 and 800 ppm (approximately 40 and 80 Pa of CO(2)). Data analysis accounted for effects on respiration rate of the normal decline in temperature with time after sunset. The median response to a 40-Pa increase in CO(2) was a 1.5% decrease in respiration rate, with responses ranging from a 5.6% inhibition to a 0.4% stimulation. Direct effects of elevated CO(2) on respiration were similar among the species. Thus, the response of nocturnal leaf respiration rate to a short-term CO(2) increase was small, and of little practical importance to the accuracy of measurements of respiration involving similar changes in CO(2) concentration during measurement. These direct respiratory responses of leaves to elevated CO(2) would translate into only slight, if any, effects on the carbon balance of temperate deciduous forests in a future atmosphere containing as much as 80 Pa CO(2).

Leaf phenology, photosynthesis, and the persistence of saplings and shrubs in a mature northern hardwood forest.

We quantified leaf phenologies of saplings and overstory trees of sugar maple (Acer saccharum Marsh.) and American beech (Fagus grandifolia Ehrh.), and the shrub hobblebush viburnum (Viburnum alnifolium Marsh.) in a 72-year-old northern hardwood forest. Seasonal changes in irradiance in the shrub layer, and in the leaf CO(2) exchange of viburnum, and sugar maple and beech saplings were also measured. Leaf expansion occurred earlier in the spring and green leaves were retained later in the autumn in saplings and shrubs than in overstory trees. During the spring light phase (before overstory closure), large CO(2) gains by all three shrub-layer species occurred as a result of a combination of relatively large leaf area, high photosynthetic capacity, and high irradiance. Throughout the summer shade phase, photosynthetic capacity at a given irradiance remained relatively constant, but CO(2) gain was typically limited by low irradiances. Even though irradiance in the shrub layer increased during the autumn light phase as the overstory opened, CO(2) gains were modest compared to springtime values because of declining leaf area and photosynthetic capacity in all three species. The CO(2) gains during the spring light phase, and to a lesser extent during the autumn light phase, may be important to the carbon balance and long-term persistence of saplings and shrubs in the usually light-limited shrub layer of a northern hardwood forest. Therefore, for some late-successional species, leaf phenology may be an important characteristic that permits their long-term persistence in the shrub layer of mature northern hardwood forests.

Energy content, construction cost and phytomass accumulation of Glycine max (L.) Merr. and Sorghum bicolor (L.) Moench grown in elevated CO2 in the field.

Grain sorghum [Sorghum bicolor (L.) Moench, a C4 crop] and soybean [Glycine max (L.) Merr. cv. Stonewall, a C3 crop] plants were grown in ambient (c. 360µl 1-1 ) and twice-ambient (c. 720 µl 1-1 ) CO2 levels in open-top chambers in soil without root constriction. Plant dry mass, energy content, composition and construction cost (i.e. amount of carbohydrate required to synthesize a unit of plant dry mass) were assessed at the end of the growing season. Elevated CO2 (a) increased phytomass accumulation (kg per plant) in both species, (b) had little affect on energy concentration (MJ kg-1 plant) but caused large increases in the amount of plant energy per ground area (MJ m-2 ground), and (c) did not alter specific growth cost (kg carbohydrate kg-1 plant growth) but greatly increased growth cost per ground area (kg carbohydrate m-2 ground) because growth was enhanced. For soybean, twice-ambient CO2 resulted in a 50 % increase in the amount of nitrogen and energy in grain (seed plus pod) per ground area. This response to elevated CO2 has important implications for agricultural productivity during the next century because the rate of human population growth is exceeding the rate of increase of land used for agriculture so that future food demands can only be met by greater production per ground area.

Autumnal leaf conductance and apparent photosynthesis by saplings and sprouts in a recently disturbed northern hardwood forest.

Leaf surface conductance and apparent photosynthesis were measured during late summer and autumn on saplings and sprouts of pin cherry (Prunus pensylvanica), yellow birch (Betula alleghaniensis), American beech (Fagus grandifolia), and sugar maple (Acer saccharum) naturally revegetating a site in the northern hardwood forest 5 years following a commercial whole-tree harvest. Prior to the disturbance (i.e., the harvest) the site was codominated by American beech, sugar maple, and yellow birch, whereas after the disturbance pin cherry was the dominant species. Conductance and photosynthetic rate of pin cherry leaves were comparatively high while those of American beech and sugar maple were low. Pin cherry retained green, physiologically active leaves longer into autumn than American beech and sugar maple. The rates and seasonal duration of leaf gas exchange on the disturbed site were therefore greater than they would have been had the site not become dominated by pin cherry.