David T. Hanson

Evolution and diversity of CO2 concentrating mechanisms in cyanobacteria

Cyanobacteria have developed an effective photosynthetic CO2 concentrating mechanism (CCM) for improving the efficiency of carboxylation by a relatively inefficient Rubisco. The development of this CCM was presumably in response to the decline in atmospheric CO2 levels and rising O2, both of which were triggered by the development of oxygenic photosynthesis by cyanobacteria themselves. In the past few years there has been a rapid expansion in our understanding of the mechanism and genes responsible for the CCM. In addition, there has been a recent expansion in the availability of complete cyanobacterial genomes, thus increasing our potential to examine questions regarding both the evolution and diversity of components of the CCM across cyanobacteria. This paper considers various CCM and photosynthesis gene components across eight cyanobacteria where significant genomic information is available. Significant conclusions from our analysis of the distribution of various genes indicated the following. Firstly, cyanobacteria have developed with two types of carboxysomes, and this is correlated with the form of Rubisco present. We have coined the terms α-cyanobacteria to refer to cyanobacteria containing Form 1A Rubisco and α-carboxysomes, and β-cyanobacteria having Form 1B Rubisco and β-carboxysomes. Secondly, there are two NDH-1 CO2 uptake systems distributed variably, withProchlorococcus marinus species appearing to lack this CO2 uptake system. There are at least two HCO3- transport systems distributed variably, with some α-cyanobacteria having an absence of systems identified in β-cyanobacteria. Finally, there are multiple forms of carbonic anhydrases (CAs), but with only β-carboxysomal CA having a clearly shown role at present. The α-cyanobacteria appear to lack a clearly identifiable carboxysomal CA. A pathway for the evolution of cyanobacterial CCMs is proposed. The acquisition of carboxysomes triggered by the rapid decline of atmospheric CO2 in the Phanerozoic is argued to be the initial step. This would then be followed by the development of NDH-1 CO2-uptake systems, followed by the development of low-and high-affinity HCO3- transporters. An intriguing question is, were carboxysomes developed first in cyanobacteria, or did they originate by the lateral transfer of pre-existing proteobacterial bacterial microcompartment genes? The potentially late evolution of the CCM genes in cyanobacteria argues for a polyphyletic and separate evolution of CCMs in cyanobacteria, algae, and higher plants.

Function of Nicotiana tabacum Aquaporins as Chloroplast Gas Pores Challenges the Concept of Membrane CO2 Permeability

Photosynthesis is often limited by the rate of CO2 diffusion from the atmosphere to the chloroplast. The primary resistances for CO2 diffusion are thought to be at the stomata and at photosynthesizing cells via a combination resulting from resistances of aqueous solution as well as the plasma membrane and both outer and inner chloroplast membranes. In contrast with stomatal resistance, the resistance of biological membranes to gas transport is not widely recognized as a limiting factor for metabolic function. We show that the tobacco (Nicotiana tabacum) plasma membrane and inner chloroplast membranes contain the aquaporin Nt AQP1. RNA interference–mediated decreases in Nt AQP1 expression lowered the CO2 permeability of the inner chloroplast membrane. In vivo data show that the reduced amount of Nt AQP1 caused a 20% change in CO2 conductance within leaves. Our discovery of CO2 aquaporin function in the chloroplast membrane opens new opportunities for mechanistic examination of leaf internal CO2 conductance regulation.

The Arabidopsis thaliana aquaporin AtPIP1;2 is a physiologically relevant CO2 transport facilitator

Cellular exchange of carbon dioxide (CO₂) is of extraordinary importance for life. Despite this significance, its molecular mechanisms are still unclear and a matter of controversy. In contrast to other living organisms, plants are physiologically limited by the availability of CO₂. In most plants, net photosynthesis is directly dependent on CO₂ diffusion from the atmosphere to the chloroplast. Thus, it is important to analyze CO₂ transport with regards to its effect on photosynthesis. A mutation of the Arabidopsis thaliana AtPIP1;2 gene, which was characterized as a non-water transporting but CO₂ transport-facilitating aquaporin in heterologous expression systems, correlated with a reduction in photosynthesis under a wide range of atmospheric CO₂ concentrations. Here, we could demonstrate that the effect was caused by reduced CO₂ conductivity in leaf tissue. It is concluded that the AtPIP1;2 gene product limits CO₂ diffusion and photosynthesis in leaves.

Evolutionary significance of isopreneemission from mosses

Isoprene emission has been documented and characterized from species in all major groups of vascular plants. We report in our survey that isoprene emission is much more common in mosses and ferns than later divergent land plants but is absent in liverworts and hornworts. The light and temperature responses of isoprene emission from Sphagnum capillifolium (Ehrh.) Hedw. are similar to those of other land plants. Isoprene increases thermotolerance of S. capillifolium to the same extent seen in higher plants as measured by chlorophyll fluorescence. Sphagnum species in a northern Wisconsin bog experienced large temperature fluctuations similar to those reported in tree canopies. Since isoprene has been shown to help plants cope with large, rapid temperature fluctuations, we hypothesize the thermal and correlated dessication stress experienced by early land plants provided the selective pressure for the evolution of light-dependent isoprene emission in the ancestors of modern mosses. As plants radiated into different habitats, this capacity was lost multiple times in favor of other thermal protective mechanisms.

The Chlamydomonas reinhardtii cia3 mutant lacking a thylakoid lumen-localized carbonic anhydrase is limited by CO2 supply to rubisco and not photosystem II function in vivo.

The Chlamydomonas reinhardtii cia3 mutant has a phenotype indicating that it requires high-CO2 levels for effective photosynthesis and growth. It was initially proposed that this mutant was defective in a carbonic anhydrase (CA) that was a key component of the photosynthetic CO2-concentrating mechanism (CCM). However, more recent identification of the genetic lesion as a defect in a lumenal CA associated with photosystem II (PSII) has raised questions about the role of this CA in either the CCM or PSII function. To resolve the role of this lumenal CA, we re-examined the physiology of the cia3 mutant. We confirmed and extended previous gas exchange analyses by using membrane-inlet mass spectrometry to monitor16O2,18O2, and CO2 fluxes in vivo. The results demonstrate that PSII electron transport is not limited in the cia3 mutant at low inorganic carbon (Ci). We also measured metabolite pools sizes and showed that the RuBP pool does not fall to abnormally low levels at low Ci as might be expected by a photosynthetic electron transport or ATP generation limitation. Overall, the results demonstrate that under low Ci conditions, the mutant lacks the ability to supply Rubisco with adequate CO2 for effective CO2 fixation and is not limited directly by any aspect of PSII function. We conclude that the thylakoid CA is primarily required for the proper functioning of the CCM at low Ci by providing an ample supply of CO2 for Rubisco.

Do bryophyte shoot systems function like vascular plant leaves or canopies? Functional trait relationships in Sphagnum mosses (Sphagnaceae)

Vascular plant leaf traits that influence photosynthetic function form the basis of mechanistic models of carbon exchange. Given their unique tissue organization, bryophytes may not express similar patterns. We investigated relationships among tissue, shoot, and canopy traits, and their associations with photosynthetic characteristics in 10 Sphagnum species. Trait relationships were organized around a primary dimension accounting for 43% of variation in 12 traits. There was no significant relationship between nitrogen content of shoot systems and maximum photosynthesis expressed on mass (A(mass)) or area (A(area)) bases due to nitrogen sequestration and storage within the canopy interior. This pattern differs from the distribution of nitrogen in vascular plant canopies. Thus, nitrogen and its relationship to carbon uptake in Sphagnum shoots does not conform to patterns of either vascular plant leaves or canopies. Species that concentrate biomass and nitrogen in the capitulum have enhanced rates of A(mass) and A(area). Consequently, A(area) was positively associated with N(area) of the capitulum only. Overall, water content and carotenoid concentration were the strongest predictors of both A(mass) and A(area) and these were expressed as inverse relationships. The relationships of plant traits in Sphagnum defines a principal trade-off between species that tolerate environmental stress and those that maximize carbon assimilation.

Promoter strength and tissue specificity effects on growth of tomato plants transformed with maize sucrose-phosphate synthase

Abstract. When sucrose-phosphate synthase (SPS; EC 2.4.1.14) is expressed in tomato (Lycopersicon esculentum Mill.) from a ribulose-1,5-bisphosphate carboxylase-oxygenase (Rubisco) small subunit (rbcS) promoter, yields are often unchanged but when SPS is expressed from a Cauliflower Mosaic Virus 35S promoter, yield is enhanced up to 80%. Two explanations for this phenomenon are (i) that expression of SPS in tissues other than leaves accounts for the increased yield or (ii) that the lower level of expression directed by the 35S promoter is more beneficial than the high level of expression directed by the rbcS promoter. To test the first hypothesis, we conducted a reciprocal graft experiment, which showed that root SPS activity did not substantially affect growth. To test the second hypothesis, we conducted a field trial using a backcrossed, segregating, population of SPS-transformed plants derived from 35S and rbcS lines. The optimal dose of SPS activity for growth was approximately twice that of the wild type regardless of which promoter was used. The effect of SPS on growth was the result of a shift in partitioning of carbon among starch, sucrose, and ionic compounds (primarily amino acids), rather than of an increase in net photosynthesis. Excessive SPS activity resulted in a decreased rate of amino acid synthesis, which could explain the non-linear response of plant growth to the level of SPS expression.

The physiological basis for genetic variation in water use efficiency and carbon isotope composition in Arabidopsis thaliana

Ecologists and physiologists have documented extensive variation in water use efficiency (WUE) in Arabidopsis thaliana, as well as association of WUE with climatic variation. Here, we demonstrate correlations of whole-plant transpiration efficiency and carbon isotope composition (δ13C) among life history classes of A. thaliana. We also use a whole-plant cuvette to examine patterns of co-variation in component traits of WUE and δ13C. We find that stomatal conductance (gs) explains more variation in WUE than does A. Overall, there was a strong genetic correlation between A and gs, consistent with selection acting on the ratio of these traits. At a more detailed level, genetic variation in A was due to underlying variation in both maximal rate of carboxylation (Vcmax) and maximum electron transport rate (Jmax). We also found strong effects of leaf anatomy, where lines with lower WUE had higher leaf water content (LWC) and specific leaf area (SLA), suggesting a role for mesophyll conductance (gm) in variation of WUE. We hypothesize that this is due to an effect through gm, and test this hypothesis using the abi4 mutant. We show that mutants of ABI4 have higher SLA, LWC, and gm than wild-type, consistent with variation in leaf anatomy causing variation in gm and δ13C. These functional data also add further support to the central, integrative role of ABI4 in simultaneously altering ABA sensitivity, sugar signaling, and CO2 assimilation. Together our results highlight the need for a more holistic approach in functional studies, both for more accurate annotation of gene function and to understand co-limitations to plant growth and productivity.

Perspectives on plant photorespiratory metabolism

Being intimately intertwined with (C3) photosynthesis, photorespiration is an incredibly high flux-bearing pathway. Traditionally, the photorespiratory cycle was viewed as closed pathway to refill the Calvin-Benson cycle with organic carbon. However, given the network nature of metabolism, it hence follows that photorespiration will interact with many other pathways. In this article, we review current understanding of these interactions and attempt to define key priorities for future research, which will allow us greater fundamental comprehension of general metabolic and developmental consequences of perturbation of this crucial metabolic process.

Engineering photorespiration: current state and future possibilities

Reduction of flux through photorespiration has been viewed as a major way to improve crop carbon fixation and yield since the energy-consuming reactions associated with this pathway were discovered. This view has been supported by the biomasses increases observed in model species that expressed artificial bypass reactions to photorespiration. Here, we present an overview about the major current attempts to reduce photorespiratory losses in crop species and provide suggestions for future research priorities.