Expression of a CO2-permeable aquaporin enhances mesophyll conductance in the C4 species setaria viridis

A fundamental limitation of photosynthetic carbon fixation is the availability of CO2. In C4 plants, primary carboxylation occurs in mesophyll cytosol, and little is known about the role of CO2 diffusion in facilitating C4 photosynthesis. We have examined the expression, localization, and functional role of selected plasma membrane intrinsic aquaporins (PIPs) from Setaria italica (foxtail millet) and discovered that SiPIP2;7 is CO2-permeable. When ectopically expressed in mesophyll cells of S. viridis (green foxtail), SiPIP2;7 was localized to the plasma membrane and caused no marked changes in leaf biochemistry. Gas-exchange and C18O16O discrimination measurements revealed that targeted expression of SiPIP2;7 enhanced the conductance to CO2 diffusion from the intercellular airspace to the mesophyll cytosol. Our results demonstrate that mesophyll conductance limits C4 photosynthesis at low pCO2 and that SiPIP2;7 is a functional CO2 permeable aquaporin that can improve CO2 diffusion at the airspace/mesophyll interface and enhance C4 photosynthesis

Complementary resource use by tree species in a rain forest tree plantation

Mixed-species tree plantations, composed of high-value native rain forest timbers, are potential forestry systems for the subtropics and tropics that can provide ecological and production benefits. Choices of rain forest tree species for mixtures are generally based on the concept that assemblages of fast-growing and light-demanding species are less productive than assemblages of species with different shade tolerances. We examined the hypothesis that mixtures of two fast-growing species compete for resources, while mixtures of shade-tolerant and shade-intolerant species are complementary. Ecophysiological characteristics of young trees were determined and analyzed with a physiology-based canopy model (MAESTRA) to test species interactions. Contrary to predictions, there was evidence for complementary interactions between two fast-growing species with respect to nutrient uptake, nutrient use efficiency, and nutrient cycling. Fast-growing Elaeocarpus angustifolius had maximum demand for soil nutrients in summer, the most efficient internal recycling of N, and low P use efficiency at the leaf and whole-plant level and produced a large amount of nutrient-rich litter. In contrast, fast-growing Grevillea robusta had maximum demand for soil nutrients in spring and highest leaf nutrient use efficiency for N and P and produced low-nutrient litter. Thus, mixtures of fast-growing G. robusta and E. angustifolius or G. robusta and slow-growing, shade-tolerant Castanospermum australe may have similar or even greater productivity than monocultures, as light requirement is just one of several factors affecting performance of mixed-species plantations. We conclude that the knowledge gained here will be useful for designing large-scale experimental mixtures and commercial forestry systems in subtropical Australia and elsewhere

Redesigning photosynthesis to sustainably meet global food and bioenergy demand

The world's crop productivity is stagnating whereas population growth, rising affluence, and mandates for biofuels put increasing demands on agriculture. Meanwhile, demand for increasing cropland competes with equally crucial global sustainability and environmental protection needs. Addressing this looming agricultural crisis will be one of our greatest scientific challenges in the coming decades, and success will require substantial improvements at many levels. We assert that increasing the efficiency and productivity of photosynthesis in crop plants will be essential if this grand challenge is to be met. Here, we explore an array of prospective redesigns of plant systems at various scales, all aimed at increasing crop yields through improved photosynthetic efficiency and performance. Prospects range from straightforward alterations, already supported by preliminary evidence of feasibility, to substantial redesigns that are currently only conceptual, but that may be enabled by new developments in synthetic biology. Although some proposed redesigns are certain to face obstacles that will require alternate routes, the efforts should lead to new discoveries and technical advances with important impacts on the global problem of crop productivity and bioenergy production

Effects of reduced carbonic anhydrase activity on CO₂ assimilation rates in Setaria viridis: a transgenic analysis

In C₄ species, the major β-carbonic anhydrase (β-CA) localized in the mesophyll cytosol catalyses the hydration of CO₂ to HCO₃-, which phosphoenolpyruvate carboxylase uses in the first step of C₄ photosynthesis. To address the role of CA in C₄ photosynthesis, we generated transgenic Setaria viridis depleted in β-CA. Independent lines were identified with as little as 13% of wild-type CA. No photosynthetic defect was observed in the transformed lines at ambient CO₂ partial pressure (pCO₂). At low pCO₂, a strong correlation between CO₂ assimilation rates and CA hydration rates was observed. C18O16O isotope discrimination was used to estimate the mesophyll conductance to CO₂ diffusion from the intercellular air space to the mesophyll cytosol (gm) in control plants, which allowed us to calculate CA activities in the mesophyll cytosol (Cm). This revealed a strong relationship between the initial slope of the response of the CO₂ assimilation rate to cytosolic pCO₂ (ACm) and cytosolic CA activity. However, the relationship between the initial slope of the response of CO₂ assimilation to intercellular pCO₂ (ACi) and cytosolic CA activity was curvilinear. This indicated that in S. viridis, mesophyll conductance may be a contributing limiting factor alongside CA activity to CO₂ assimilation rates at low pCO₂.This research was supported by the Bill and Melinda Gates Foundation’s funding for the C₄ Rice consortium and by the Australian Research Council Centre of Excellence for Translational Photosynthesis (CE140100015). RES is funded by ARC DECRA (DE130101760)

Transgenic maize phosphoenolpyruvate carboxylase alters leaf-atmosphere CO2 and 13CO2 exchanges in Oryza sativa.

The engineering process of C4 photosynthesis into C3 plants requires an increased activity of phosphoenolpyruvate carboxylase (PEPC) in the cytosol of leaf mesophyll cells. The literature varies on the physiological effect of transgenic maize (Zea mays) PEPC (ZmPEPC) leaf expression in Oryza sativa (rice). Therefore, to address this issue, leaf-atmosphere CO2 and 13CO2 exchanges were measured, both in the light (at atmospheric O2 partial pressure of 1.84 kPa and at different CO2 levels) and in the dark, in transgenic rice expressing ZmPEPC and wild-type (WT) plants. The in vitro PEPC activity was 25 times higher in the PEPC overexpressing (PEPC-OE) plants (~20% of maize) compared to the negligible activity in WT. In the PEPC-OE plants, the estimated fraction of carboxylation by PEPC (β) was ~6% and leaf net biochemical discrimination against 13CO2[Formula: see text] was ~ 2‰ lower than in WT. However, there were no differences in leaf net CO2 assimilation rates (A) between genotypes, while the leaf dark respiration rates (Rd) over three hours after light-dark transition were enhanced (~ 30%) and with a higher 13C composition [Formula: see text] in the PEPC-OE plants compared to WT. These data indicate that ZmPEPC in the PEPC-OE rice plants contributes to leaf carbon metabolism in both the light and in the dark. However, there are some factors, potentially posttranslational regulation and PEP availability, which reduce ZmPEPC activity in vivo

A sorghum (Sorghum bicolor) mutant with altered carbon isotope ratio

Recent efforts to engineer C4 photosynthetic traits into C3 plants such as rice demand an understanding of the genetic elements that enable C4 plants to outperform C3 plants. As a part of the C4 Rice Consortium’s efforts to identify genes needed to support C4 photosynthesis, EMS mutagenized sorghum populations were generated and screened to identify genes that cause a loss of C4 function. Stable carbon isotope ratio (δ13C) of leaf dry matter has been used to distinguishspecies with C3 and C4 photosynthetic pathways. Here, we report the identification of a sorghum (Sorghum bicolor) mutant with a low δ13C characteristic. A mutant (named Mut33) with a pale phenotype and stunted growth was identified from an EMS treated sorghum M2 population. The stable carbon isotope analysis of the mutants showed a decrease of 13C uptake capacity. The noise of random mutation was reduced by crossing the mutant and its wildtype (WT). The back-cross (BC1F1) progenies were like the WT parent in terms of 13C values and plant phenotypes. All the BC1F2 plants with low δ13C died before they produced their 6th leaf. Gas exchange measurements of the low δ13C sorghum mutants showed a higher CO2 compensation point (25.24 μmol CO2.mol-1air) and the maximum rate of photosynthesis was less than 5μmol.m-2.s-1. To identify the genetic determinant of this trait, four DNA pools were isolated; two each from normal and low δ13C BC1F2 mutant plants. These were sequenced using an Illumina platform. Comparison of allele frequency of the single nucleotide polymorphisms (SNPs) between the pools with contrasting phenotype showed that a locus in Chromosome 10 between 57,941,104 and 59,985,708 bps had an allele frequency of 1. There were 211 mutations and 37 genes in the locus, out of which mutations in 9 genes showed non-synonymous changes. This finding is expected to contribute to future research on the identification of the causal factor differentiating C4 from C3 species that can be used in the transformation of C3 to C4 plants

Rieske FeS overexpression in tobacco provides increased abundance and activity of Cytochrome b₆f

Photosynthesis is fundamental for plant growth and yield. The Cytochrome b₆ f complex catalyses a rate-limiting step in thylakoid electron transport and therefore represents an important point of regulation of photosynthesis. Here we show that overexpression of a single core subunit of Cytochrome bA ) and plastoquinone pool and faster electron transport from the plastoquinone pool to Photosystem I upon changes in irradiance, compared to control plants. A faster establishment of qE , the energy-dependent component of non-photochemical quenching, in transgenic plants suggests a more rapid build-up of the transmembrane proton gradient, also supporting the increased in vivo Cytochrome b₆ f activity. However, there was no consistent increase in steady-state rates of electron transport or CO₂ assimilation in plants overexpressing Rieske FeS grown in either laboratory conditions or field trials, suggesting that the in vivo activity of the complex was only transiently increased upon changes in irradiance. Our results show that overexpression of Rieske FeS in tobacco enhances the abundance of functional Cytochrome b₆ f and may have the potential to increase plant productivity if combined with other traits

Thermoperiodic growth control by gibberellin does not involve changes in photosynthetic or respiratory capacities in pea

Active gibberellin (GA1) is an important mediator of thermoperiodic growth in pea. Plants grown under lower day than night temperature (negative DIF) elongate less and have reduced levels of GA1 compared with plants grown at higher day than night temperature (positive DIF). By comparing the wild type (WT) and the elongated DELLA mutant la crys, this study has examined the effect of impaired GA signalling on thermoperiodic growth, photosynthesis, and respiration in pea. In the WT a negative DIF treatment reduced stem mass ratio and increased both root mass ratio and leaf mass ratio (dry weight of specific tissue related to total plant dry weight). Leaf, root and stem mass ratios of la crys were not affected by DIF. Under negative DIF, specific leaf area (projected leaf area per unit leaf dry mass), biomass, and chlorophyll content of WT and la crys plants were reduced. Young, expanding leaves of plants grown under negative DIF had reduced leaf area-based photosynthetic capacity. However, the highest photosynthetic electron transport rate was found in fully expanded leaves of WT plants grown under negative DIF. Negative DIF increased night respiration and was similar for both genotypes. It is concluded that GA signalling is not a major determinant of leaf area-based photosynthesis or respiration and that reduced dry weight of plants grown under negative DIF is caused by a GA-mediated reduction of photosynthetic stem and leaf tissue, reduced photosynthesis of young, expanding leaves, and reduced growth caused by low temperature in the photoperiod

Temperature responses of Rubisco from Paniceae grasses provide opportunities for improving C3 photosynthesis.

Enhancing the catalytic properties of the CO2-fixing enzyme Rubisco is a target for improving agricultural crop productivity. Here, we reveal extensive diversity in the kinetic response between 10 and 37 °C by Rubisco from C3 and C4 species within the grass tribe Paniceae. The CO2 fixation rate (kcatc) for Rubisco from the C4 grasses with nicotinamide adenine dinucleotide (NAD) phosphate malic enzyme (NADP-ME) and phosphoenolpyruvate carboxykinase (PCK) photosynthetic pathways was twofold greater than the kcatc of Rubisco from NAD-ME species across all temperatures. The declining response of CO2/O2 specificity with increasing temperature was less pronounced for PCK and NADP-ME Rubisco, which would be advantageous in warmer climates relative to the NAD-ME grasses. Modelled variation in the temperature kinetics of Paniceae C3 Rubisco and PCK Rubisco differentially stimulated C3 photosynthesis relative to tobacco above and below 25 °C under current and elevated CO2. Amino acid substitutions in the large subunit that could account for the catalytic variation among Paniceae Rubisco are identified; however, incompatibilities with Paniceae Rubisco biogenesis in tobacco hindered their mutagenic testing by chloroplast transformation. Circumventing these bioengineering limitations is critical to tailoring the properties of crop Rubisco to suit future climates

A cross-scale analysis to understand and quantify the effects of photosynthetic enhancement on crop growth and yield across environments

Abstract Photosynthetic manipulation provides new opportunities for enhancing crop yield. However, understanding and quantifying the importance of individual and multiple manipulations on the seasonal biomass growth and yield performance of target crops across variable production environments is limited. Using a state-of-the-art cross-scale model in the APSIM platform we predicted the impact of altering photosynthesis on the enzyme-limited (Ac) and electron transport-limited (Aj) rates, seasonal dynamics in canopy photosynthesis, biomass growth, and yield formation via large multiyear-by-location crop growth simulations. A broad list of promising strategies to improve photosynthesis for C3 wheat and C4 sorghum were simulated. In the top decile of seasonal outcomes, yield gains were predicted to be modest, ranging between 0% and 8%, depending on the manipulation and crop type. We report how photosynthetic enhancement can affect the timing and severity of water and nitrogen stress on the growing crop, resulting in nonintuitive seasonal crop dynamics and yield outcomes. We predicted that strategies enhancing Ac alone generate more consistent but smaller yield gains across all water and nitrogen environments, Aj enhancement alone generates larger gains but is undesirable in more marginal environments. Large increases in both Ac and Aj generate the highest gains across all environments. Yield outcomes of the tested manipulation strategies were predicted and compared for realistic Australian wheat and sorghum production. This study uniquely unpacks complex cross-scale interactions between photosynthesis and seasonal crop dynamics and improves understanding and quantification of the potential impact of photosynthesis traits (or lack of it) for crop improvement research