[Show abstract][Hide abstract] ABSTRACT: Pineapple (Ananas comosus (L.) Merr.) is the most economically valuable crop possessing crassulacean acid metabolism (CAM), a photosynthetic carbon assimilation pathway with high water-use efficiency, and the second most important tropical fruit. We sequenced the genomes of pineapple varieties F153 and MD2 and a wild pineapple relative, Ananas bracteatus accession CB5. The pineapple genome has one fewer ancient whole-genome duplication event than sequenced grass genomes and a conserved karyotype with seven chromosomes from before the ρ duplication event. The pineapple lineage has transitioned from C3 photosynthesis to CAM, with CAM-related genes exhibiting a diel expression pattern in photosynthetic tissues. CAM pathway genes were enriched with cis-regulatory elements associated with the regulation of circadian clock genes, providing the first cis-regulatory link between CAM and circadian clock regulation. Pineapple CAM photosynthesis evolved by the reconfiguration of pathways in C3 plants, through the regulatory neofunctionalization of preexisting genes and not through the acquisition of neofunctionalized genes via whole-genome or tandem gene duplication.
[Show abstract][Hide abstract] ABSTRACT: Global climate change threatens the sustainability of agriculture and agroforestry worldwide through increased heat, drought, surface evaporation, and associated soil drying. Exposure of crops and forests to warmer and drier environments will increase leaf:air water vapor-pressure deficits (VPD), and result in increased drought susceptibility and reduced productivity, particularly in arid regions, but also in tropical regions with seasonal dry periods. Fast-growing, short-rotation forestry (SRF) bioenergy crops such, as poplar (Populus spp.) and willow (Salix spp.), are particularly susceptible to hydraulic failure following drought stress due to their isohydric nature and relatively high stomatal conductance. One approach to sustaining plant productivity is to improve water-use efficiency (WUE) by engineering crassulacean acid metabolism (CAM) into C3 crops. CAM improves WUE by shifting stomatal opening and primary CO2 uptake and fixation to the nighttime when leaf:air VPD is low. CAM members of the tree genus Clusia exemplify the compatibility of CAM performance within tree species and highlight CAM as a mechanism to conserve water and maintain carbon uptake during drought conditions. The introduction of bioengineered CAM into SRF bioenergy trees is a potentially viable path to sustaining agroforestry production systems in the face of a globally changing climate.
No preview · Article · Sep 2015 · Plant Cell and Environment
[Show abstract][Hide abstract] ABSTRACT: Sustainable production of lignocellulosic biofuels requires a sufficient supply of biomass feedstocks. Agave and Opuntia represent highly water-use efficient bioenergy crops that are suitable for expanding feedstock production into semi-arid marginal lands. These feedstocks have garnered interest as dedicated biofuel feedstocks because of their high water- and fertilizer-use efficiency and not competing with major food crops or conventional biofuel feedstocks. To better understand the potential of these feedstocks, the biomass composition of Agave tequilana and Opuntia ficus-indica was analyzed. Previous extraction procedures and analytical methods have led to variable estimates of the chemical compositions of the biomass of these species. Therefore, National Renewable Energy Laboratory (NREL) standard methods were used in the present study. A. tequilana showed higher mass fractions of water-soluble constituents, structural carbohydrates, cellulose, hemicellulose, and lignin than O. ficus-indica. In contrast, O. ficus-indica had higher protein, water, and ash mass fractions than A. tequilana. Both species had lower lignin mass fractions, thus yielding lower heating values, but had higher water and ash mass fractions than most woody biomass feedstocks. The high water mass fractions of these species (85–94%) could prove advantageous for biomass deconstruction and aqueous phase catalytic conversion processes as less exogenous water inputs would be needed. Lastly, solid-state NMR analysis revealed that both A. tequilana and O. ficus-indica had high amorphous and para-crystalline cellulose mass fractions (>80%), indicating that these biomass feedstocks would be far less recalcitrant to deconstruction than traditional lignocellulosic biomass feedstocks.
Full-text · Article · May 2015 · Biomass and Bioenergy
[Show abstract][Hide abstract] ABSTRACT: Microalgae can serve as useful feedstocks for biofuel production as they can be grown with fresh, brackish, or salt water and their lipid and starch contents can be manipulated to create customized feedstocks for different classes of biofuels. Continuous buoyant density gradient centrifugation (CBDGC) was used to perform reiterative, transgressive selection to isolate wildtype and ethyl methanesulfonate-mutagenized Dunaliella salina cells with enhanced lipid and starch production. Sixty rounds of transgressive selection resulted in the isolation of cell populations with significantly lower or higher buoyant densities. Lipid content in the low-density populations was enhanced by 1.2- to 2.9-fold in wildtype cells and 1.3- to 2.3-fold in mutagenized cells as measured by Nile Red dye staining, but the lipid content differences were not significant when quantified by liquid chromatography–tandem mass spectroscopy possibly due to the composition of the lipid pools measured by these contrasting techniques. In contrast, starch content in the high-density populations was increased by 2-fold in wild type cells and 1.4- to 1.6-fold in mutagenized cells, respectively. The observed alterations in lipid and starch contents appeared to be stable after more than 70 weeks (392 cell generations). CBDGC-based selection provides a useful and accessible technological alternative to genetic engineering approaches for the customization of microbial biofuel feedstocks.
[Show abstract][Hide abstract] ABSTRACT: Microalgae offer great potential as a third-generation biofuel feedstock, especially when grown on wastewater, as they have the dual application for wastewater treatment and as a biomass feedstock for biofuel production. The potential for growth on wastewater centrate was evaluated for forty microalgae strains from fresh (11), brackish (11), or saltwater (18) genera. Generally, freshwater strains were able to grow at high concentrations of centrate, with two strains, Neochloris pseudostigmata and N. conjuncta, demonstrating growth at up to 40% v/v centrate. Fourteen of eighteen salt water Dunaliella strains also demonstrated growth in centrate concentrations at or above 40% v/v. Lipid profiles of freshwater strains with high-centrate tolerance were determined using gas chromatography-mass spectrometry (GC-MS) and compared against those obtained on cells grown on defined maintenance media. The major lipid compounds were found to be palmitic (16:0), oleic (18:1), and linoleic (18:2) acids for all freshwater strains grown on either centrate or their respective maintenance medium. These results demonstrate the highly concentrated wastewater can be used to grow microalgae, which limits the need to dilute wastewater prior to algal production. In addition, the algae produced generate lipids suitable for biodiesel or green diesel production.
Preview · Article · May 2015 · Frontiers in Energy Research
[Show abstract][Hide abstract] ABSTRACT: Global climate change is predicted to increase heat, drought, and soil-drying conditions, and thereby increase crop sensitivity
to water vapour pressure deficit, resulting in productivity losses. Increasing competition between agricultural freshwater
use and municipal or industrial uses suggest that crops with greater heat and drought durability and greater water-use efficiency
will be crucial for sustainable biomass production systems in the future. Agave (Agavaceae) and Opuntia (Cactaceae) represent highly water-use efficient bioenergy crops that could diversify bioenergy feedstock supply yet preserve
or expand feedstock production into semi-arid, abandoned, or degraded agricultural lands, and reclaim drylands. Agave and Opuntia are crassulacean acid metabolism species that can achieve high water-use efficiencies and grow in water-limited areas with
insufficient precipitation to support traditional C3 or C4 bioenergy crops. Both Agave and Opuntia have the potential to produce above-ground biomass rivalling that of C3 and C4 crops under optimal growing conditions. The low lignin and high amorphous cellulose contents of Agave and Opuntia lignocellulosic biomass will be less recalcitrant to deconstruction than traditional feedstocks, as confirmed by pretreatments
that improve saccharification of Agave. Refined environmental productivity indices and geographical information systems modelling have provided estimates of Agave and Opuntia biomass productivity and terrestrial sequestration of atmospheric CO2; however, the accuracy of such modelling efforts can be improved through the expansion of field trials in diverse geographical
settings. Lastly, life cycle analysis indicates that Agave would have productivity, life cycle energy, and greenhouse gas balances comparable or superior to those of traditional bioenergy
feedstocks, but would be far more water-use efficient.
No preview · Article · Apr 2015 · Journal of Experimental Botany
[Show abstract][Hide abstract] ABSTRACT: Crassulacean acid metabolism (CAM) is an elaboration of C3 photosynthesis wherein carbon
assimilation occurs at night to reduce daytime water losses through a temporal separation of primary
C4 and secondary C3 carboxylation reactions. The circadian clock controls the temporal separation of
these potentially competing reactions. However, the underlying orchestration of transcriptional
network modules of CAM is poorly understood. Comparative RNA-seq analysis was performed in
well-watered (C3 performing) and water-deficit-stressed (CAM performing) common ice plants
(Mesembryanthemum crystallinum L.), a facultative CAM model species. Leaves were collected every
4 h over a 72 h time course under both 24 h light/dark and 48 h light/light conditions to characterize
the circadian clock-controlled transcriptome in both the C3 photosynthesis and CAM states. Under
water-deficit conditions that induce CAM, greater numbers of transcripts become rhythmic indicating
that the stress-adaptive and CAM transcriptional machinery is directly under circadian clock control.
Weighted co-expression network analyses of differentially expressed genes upon CAM induction (log2
fold-change in CAM performing compared to C3 photosynthesis performing) revealed both persistent
and induced circadian clock-controlled transcriptional network modules. Together these results
tentatively identified CAM-specific network modules and provide insights into the circadian clock controlled
transcriptional expression of CAM.
[Show abstract][Hide abstract] ABSTRACT: Phosphoenolpyruvate carboxylase (PEPC) catalyses the initial fixation of atmospheric CO2 into oxaloacetate and subsequently malate. Nocturnal accumulation of malic acid within the vacuole of photosynthetic cells
is a typical feature of plants that perform crassulacean acid metabolism (CAM). PEPC is a ubiquitous plant enzyme encoded
by a small gene family, and each member encodes an isoform with specialized function. CAM-specific PEPC isoforms probably
evolved from ancestral non-photosynthetic isoforms by gene duplication events and subsequent acquisition of transcriptional
control elements that mediate increased leaf-specific or photosynthetic-tissue-specific mRNA expression. To understand the
patterns of functional diversification related to the expression of CAM, ppc gene families and photosynthetic patterns were characterized in 11 closely related orchid species from the subtribe Oncidiinae
with a range of photosynthetic pathways from C3 photosynthesis (Oncidium cheirophorum, Oncidium maduroi, Rossioglossum krameri, and Oncidium sotoanum) to weak CAM (Oncidium panamense, Oncidium sphacelatum, Gomesa flexuosa and Rossioglossum insleayi) and strong CAM (Rossioglossum ampliatum, Trichocentrum nanum, and Trichocentrum carthagenense). Phylogenetic analysis revealed the existence of two main ppc lineages in flowering plants, two main ppc lineages within the eudicots, and three ppc lineages within the Orchidaceae. Our results indicate that ppc gene family expansion within the Orchidaceae is likely to be the result of gene duplication events followed by adaptive sequence
divergence. CAM-associated PEPC isoforms in the Orchidaceae probably evolved from several independent origins.
Full-text · Article · Jun 2014 · Journal of Experimental Botany
[Show abstract][Hide abstract] ABSTRACT: To meet future food and energy security needs, which are amplified by increasing population growth and reduced natural resource availability, metabolic engineering efforts have moved from manipulating single genes/proteins to introducing multiple genes and novel pathways to improve photosynthetic efficiency in a more comprehensive manner. Biochemical carbon-concentrating mechanisms such as crassulacean acid metabolism (CAM), which improves photosynthetic, water-use, and possibly nutrient-use efficiency, represent a strategic target for synthetic biology to engineer more productive C3 crops for a warmer and drier world. One key challenge for introducing multigene traits like CAM onto a background of C3 photosynthesis is to gain a better understanding of the dynamic spatial and temporal regulatory events that underpin photosynthetic metabolism. With the aid of systems and computational biology, vast amounts of experimental data encompassing transcriptomics, proteomics, and metabolomics can be related in a network to create dynamic models. Such models can undergo simulations to discover key regulatory elements in metabolism and suggest strategic substitution or augmentation by synthetic components to improve photosynthetic performance and water-use efficiency in C3 crops. Another key challenge in the application of synthetic biology to photosynthesis research is to develop efficient systems for multigene assembly and stacking. Here, we review recent progress in computational modelling as applied to plant photosynthesis, with attention to the requirements for CAM, and recent advances in synthetic biology tool development. Lastly, we discuss possible options for multigene pathway construction in plants with an emphasis on CAM-into-C3 engineering.
Preview · Article · Feb 2014 · Journal of Experimental Botany
[Show abstract][Hide abstract] ABSTRACT: Climatic extremes threaten agricultural sustainability worldwide. One approach to increase plant water- use efficiency (WUE) is to introduce crassulacean acid metabolism (CAM) into C3 crops. Such a task requires comprehensive systems-level understanding of the enzymatic and regulatory pathways underpinning this temporal CO2 pump. Here we review the progress that has been made in achieving this goal. Given that CAM arose through multiple independent evolutionary ori- gins, comparative transcriptomics and genomics of taxonomically diverse CAM species are being used to define the genetic ‘parts list’ required to operate the core CAM functional modules of nocturnal carboxylation, diurnal decarboxylation, and inverse stomatal regulation. Engi- neered CAM offers the potential to sustain plant productivity for food, feed, fiber, and biofuel production in hotter and drier climates.
No preview · Article · Feb 2014 · Trends in Plant Science
[Show abstract][Hide abstract] ABSTRACT: Crassulacean acid metabolism (CAM) is an elaboration of C3 photosynthetic that limits atmospheric CO2 fixation to all or part of the nighttime thereby reducing transpiration during the day and improving water-use efficiency. The common or crystalline ice plant (Mesembryanthemum crystallinum L.) is a facultative CAM and halophytic model species that can switch from C3 photosynthesis to CAM following high salinity or water-deficit stress treatments. The estimated genome size of ice plant is 390 Mb in 9 chromosomes (2N=18). To reveal the molecular basis of halophytism and CAM, the ice plant genome is being sequenced and analyzed through hybrid assembly approaches. Results from three different sequencing platforms, Illumina Mate Pair (MP), Illumina Paired End (PE) and PacBio (PB) long reads, that are being used to sequence the genome will be discussed. Two strategies will be applied for hybrid assembly of the genome. Strategy 1 will build contigs from PB reads after error-correction by PE data followed by scaffolding using MP reads. Strategy 2 will rely on the assembly of contigs build from PE reads combined with scaffold assembly using MP reads, followed by gap filling with PB reads. The completed ice plant genome sequence will facilitate our understanding of the genomic basis of salt tolerance and the water-use efficiency that permits CAM species to inhabit semi-arid and arid environments.
[Show abstract][Hide abstract] ABSTRACT: Crassulacean acid metabolism (CAM) is a water-use efficient photosynthetic adaptation found in more than 6% of vascular plant species. This adaptation is distinguished by the temporal separation of primary C4 and secondary C3 carboxylation reactions to reduce daytime water loss. To investigate the underlying orchestration of transcriptional networks controlling the temporal separation of these competing carboxylation reactions and their associated metabolic demands, transcriptomic and metabolomic analyses were conducted on the facultative CAM ice plant, Mesembryanthemum crystallinum. These omic technologies revealed diel and circadian changes in transcript and metabolite abundance patterns in ice plants performing C3 and CAM (induced by water-deficit stress) over a 72 h time course. Key differences in transcripts and metabolites were identified between the C3 and CAM states that are involved in CAM adaptation and possibly their role in the regulation of CAM by the circadian clock. Other pivotal omic results and candidate CAM transcription factors to elucidate clock-controlled regulatory networks operating in mesophyll cells will be discussed.
[Show abstract][Hide abstract] ABSTRACT: Improved crop water-use efficiency (WUE) is critical for the long-term sustainability of agricultural production systems in the face of predicted future warmer and drier climates. Crassulacean acid metabolism (CAM) is a specialized mode of photosynthesis that enhances WUE through an inverse day/night pattern of stomatal closure/opening and improves photosynthetic efficiency by concentrating CO2 around RUBISCO. CAM has evolved multiple times from C3 photosynthesis and ~6.5% of higher plant species in more than 35 families have acquired CAM via parallel or convergent evolution. There are two fundamental questions to be answered to understand the molecular basis and evolutionary mechanism of CAM: 1) what are the genetic differences between CAM and non-CAM species and 2) what are the common molecular features shared among CAM plants from diverse origins? To address these questions, comparative genomics analysis was performed using multiple plant species including CAM (e.g., Agave, Kalanchoe, Mesembryanthemum), C3 (e.g., Arabidopsis, Oryza, Populus), C4 (e.g., Setaria, Sorghum, and Zea), and non-vascular plant species (e.g., Physcomitrella, Selaginella). Our analysis not only revealed orthologous gene groups shared between CAM and non-CAM species, but also identified genes specific to the CAM species. Also, expanded gene families were identified in CAM species compared with non-CAM species. Gene ontology and gene expression profiles were used to build hypothesis related to divergent gene functions that likely arose during CAM evolution. This research establishes a framework for CAM comparative genomics studies and provides new knowledge to inform genetic improvement in WUE and photosynthetic efficiency in crop plants under water-limiting conditions.
[Show abstract][Hide abstract] ABSTRACT: The development of low water input forages would be useful for improving the
water-use efficiency of livestock production in semi-arid and arid regions. The
desiccation-tolerant (DT) species Sporobolus stapfianus Gandoger and two desic-
cation-sensitive (DS) species, Sporobolus pyramidalis and Sporobolus fimbriatus
(Trin.) Nees. (Poaceae), were evaluated for aerial biomass production and seed
productivity under three different irrigation regimes. Sporobolus stapfianus dis-
played the least biomass production, whereas S. pyramidalis and S. fimbriatus
produced up to 3.8- and 11.2-fold greater dry biomass, respectively, at the highest
irrigation rate of 12 334 l (0.01 acre-feet). Sporobolus fimbriatus and to a lesser
extent S. pyramidalis showed significant increases in biomass production in
response to increased irrigation rates, whereas S. stapfianus did not. Sporobolus
pyramidalis and S. fimbriatus exhibited 3.2- and 6.0-fold greater seed production,
respectively, in response to increased irrigation rates, whereas S. stapfianus
showed only a 1.4-fold increase. All Sporobolus species possessed forage quality
traits (e.g. crude protein, fibre content) comparable to those of timothy, a forage
grass grown widely in the Great Basin in the western United States. Micronutrient
content exceeded the minimum requirements of beef cattle, without surpassing
tolerable limits, with the exception of zinc, which appeared low in all three Sporo-
bolus species. The low water requirements displayed by these species, combined
with their acceptable forage qualities, indicate that these grasses have the poten-
tial to serve farmers and ranchers in semi-arid and arid regions of the western
United States where irrigation resources are limited.
Full-text · Article · Oct 2013 · Journal of Agronomy and Crop Science
[Show abstract][Hide abstract] ABSTRACT: Over 13% of all genes in the Arabidopsis thaliana genome encode for proteins classified as having a completely unknown function, with the function of >30% of the Arabidopsis proteome poorly characterized. Although empirical data in the form of mRNA and proteome profiling experiments suggest that many of these proteins play an important role in different biological processes, their functional characterization remains one of the major challenges in modern biology. To expand the annotation of genes with unknown function involved in the response of Arabidopsis to different environmental stress conditions, we selected 1007 such genes and tested the response of their corresponding homozygous T-DNA insertional mutants to salinity, oxidative, osmotic, heat, cold and hypoxia stresses. Depending on the specific abiotic stresses tested, 12-31% of mutants had an altered stress-response phenotype. Interestingly, 832 out of 1007 mutants showed tolerance or sensitivity to more than one abiotic stress treatment, suggesting that genes of unknown function could play an important role in abiotic stress-response signaling, or general acclimation mechanisms. Further analysis of multiple stress-response phenotypes within different populations of mutants revealed interesting links between acclimation to heat, cold and oxidative stresses, as well as between sensitivity to ABA, osmotic, salinity, oxidative and hypoxia stresses. Our findings provide a significant contribution to the biological characterization of genes with unknown function in Arabidopsis and demonstrate that many of these genes play a key role in the response of plants to abiotic stresses.
Full-text · Article · Mar 2013 · Physiologia Plantarum
[Show abstract][Hide abstract] ABSTRACT: Aerobic metabolism of vertebrates is linked to membrane fatty acid (FA) composition. Although the membrane pacemaker hypothesis posits that desaturation of FAs accounts for variation in resting or basal metabolic rate (BMR), little is known about the FA profiles that underpin variation in maximal metabolic rate (MMR). We examined membrane FA composition of liver and skeletal muscle in mice after seven generations of selection for increased MMR. In both liver and skeletal muscle, unsaturation index did not differ between control and high-MMR mice. We also examined membrane FA composition at the individual-level of variation. In liver, 18:0, 20:3 n-6, 20:4 n-6, and 22:6 n-3 FAs were significant predictors of MMR. In gastrocnemius muscle, 18:2 n-6, 20:4 n-6, and 22:6 n-3 FAs were significant predictors of MMR. In addition, muscle 16:1 n-7, 18:1 n-9, and 22:5 n-3 FAs were significant predictors of BMR, whereas no liver FAs were significant predictors of BMR. Our findings indicate that (i) individual variation in MMR and BMR appears to be linked to membrane FA composition in the skeletal muscle and liver, and (ii) FAs that differ between selected and control lines are involved in pathways that can affect MMR or BMR.
Full-text · Article · Feb 2013 · Comparative biochemistry and physiology. Part A, Molecular & integrative physiology