Identifying genomic regions associated with C4 photosynthetic activity and leaf anatomy in Alloteropsis semialata.

C4 photosynthesis is a complex trait requiring multiple developmental and metabolic alterations. Despite this complexity, it has independently evolved over 60 times. However, our understanding of the transition to C4 is complicated by the fact that variation in photosynthetic type is usually segregated between species that diverged a long time ago. Here, we perform a genome-wide association study (GWAS) using the grass Alloteropsis semialata, the only known species to have C3, intermediate, and C4 accessions that recently diverged. We aimed to identify genomic regions associated with the strength of the C4 cycle (measured using δ13C), and the development of C4 leaf anatomy. Genomic regions correlated with δ13C include regulators of C4 decarboxylation enzymes (RIPK), nonphotochemical quenching (SOQ1), and the development of Kranz anatomy (SCARECROW-LIKE). Regions associated with the development of C4 leaf anatomy in the intermediate individuals contain additional leaf anatomy regulators, including those responsible for vein patterning (GSL8) and meristem determinacy (GIF1). The parallel recruitment of paralogous leaf anatomy regulators between A. semialata and other C4 lineages implies the co-option of these genes is context-dependent, which likely has implications for the engineering of the C4 trait into C3 species.

Diversification of quantitative morphological traits in wheat.

BACKGROUND AND AIMS: The development and morphology of crop plants have been profoundly altered by evolution under cultivation, initially through unconscious selection, without deliberate foresight, and later by directed breeding. Wild wheats remain an important potential source of variation for modern breeders; however, the sequence and timing of morphological changes during domestication are not fully resolved. METHODS: We grew and measured 142 wheat accessions representing different stages in wheat evolution, including three independent domestication events, and compared their morphological traits to define the morphospace of each group. KEY RESULTS: The results show that wild and domesticated wheats have overlapping morphospaces, but each also occupies a distinct area of morphospace from one another. Polyploid formation in wheat increased leaf biomass and seed weight but had its largest effects on tiller loss. Domestication continued to increase the sizes of wheat leaves and seeds and made wheat grow taller, with more erect architecture. Associated changes to the biomass of domesticated wheats generated more grains and achieved higher yields. Landrace improvement subsequently decreased the numbers of tillers and spikes, to focus resource allocation to the main stem, accompanied by a thicker main stem and larger flag leaves. During the Green Revolution, wheat height was reduced to increase the harvest index and therefore yield. Modern wheats also have more erect leaves and larger flower biomass proportions than landraces. CONCLUSIONS: Quantitative trait history in wheat differs by trait. Some trait values show progressive changes in the same direction (e.g. leaf size, grain weight), whereas others change in a punctuated way at particular stages (e.g. canopy architecture), and other trait values switch directions during wheat evolution (e.g. plant height, flower biomass proportion). Agronomically valued domestication traits arose during different stages of wheat history, such that modern wheats are the product of >10 000 years of morphological evolution.

Impacts of ground-level ozone on sugarcane production.

Sugarcane is a vital commodity crop often grown in (sub)tropical regions which have been experiencing a recent deterioration in air quality. Unlike for other commodity crops, the risk of air pollution, specifically ozone (O3), to this C4 crop has not yet been quantified. Yet, recent work has highlighted both the potential risks of O3 to C4 bioenergy crops, and the emergence of O3 exposure across the tropics as a vital factor determining global food security. Given the large extent, and planned expansion of sugarcane production in places like Brazil to meet global demand for biofuels, there is a pressing need to characterize the risk of O3 to the industry. In this study, we sought to a) derive sugarcane O3 dose-response functions across a range of realistic O3 exposure and b) model the implications of this across a globally important production area. We found a significant impact of O3 on biomass allocation (especially to leaves) and production across a range of sugarcane genotypes, including two commercially relevant varieties (e.g. CTC4, Q240). Using these data, we calculated dose-response functions for sugarcane and combined them with hourly O3 exposure across south-central Brazil derived from the UK Earth System Model (UKESM1) to simulate the current regional impact of O3 on sugarcane production using a dynamic global vegetation model (JULES vn 5.6). We found that between 5.6 % and 18.3 % of total crop productivity is likely lost across the region due to the direct impacts of current O3 exposure. However, impacts depended critically on the substantial differences in O3 susceptibility observed among sugarcane genotypes and how these were implemented in the model. Our work highlights not only the urgent need to fully elucidate the impacts of O3 in this important bioenergetic crop, but the potential implications air quality may have upon tropical food production more generally.

Alloteropsis semialata as a study system for C4 evolution in grasses.

BACKGROUND: Numerous groups of plants have adapted to CO2 limitations by independently evolving C4 photosynthesis. This trait relies on concerted changes in anatomy and biochemistry to concentrate CO2 within the leaf and thereby boost productivity in tropical conditions. The ecological and economic importance of C4 photosynthesis has motivated intense research, often relying on comparisons between distantly related C4 and non-C4 plants. The photosynthetic type is fixed in most species, with the notable exception of the grass Alloteropsis semialata. This species includes populations exhibiting the ancestral C3 state in southern Africa, intermediate populations in the Zambezian region and C4 populations spread around the palaeotropics. SCOPE: We compile here the knowledge on the distribution and evolutionary history of the Alloteropsis genus as a whole and discuss how this has furthered our understanding of C4 evolution. We then present a chromosome-level reference genome for a C3 individual and compare the genomic architecture with that of a C4 accession of A. semialata. CONCLUSIONS: Alloteropsis semialata is one of the best systems in which to investigate the evolution of C4 photosynthesis because the genetic and phenotypic variation provides a fertile ground for comparative and population-level studies. Preliminary comparative genomic investigations show that the C3 and C4 genomes are highly syntenic and have undergone a modest amount of gene duplication and translocation since the different photosynthetic groups diverged. The background knowledge and publicly available genomic resources make A. semialata a great model for further comparative analyses of photosynthetic diversification.

Leaf anatomy explains the strength of C4 activity within the grass species Alloteropsis semialata.

C4 photosynthesis results from anatomical and biochemical characteristics that together concentrate CO2 around ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), increasing productivity in warm conditions. This complex trait evolved through the gradual accumulation of components, and particular species possess only some of these, resulting in weak C4 activity. The consequences of adding C4 components have been modelled and investigated through comparative approaches, but the intraspecific dynamics responsible for strengthening the C4 pathway remain largely unexplored. Here, we evaluate the link between anatomical variation and C4 activity, focusing on populations of the photosynthetically diverse grass Alloteropsis semialata that fix various proportions of carbon via the C4 cycle. The carbon isotope ratios in these populations range from values typical of C3 to those typical of C4 plants. This variation is statistically explained by a combination of leaf anatomical traits linked to the preponderance of bundle sheath tissue. We hypothesize that increased investment in bundle sheath boosts the strength of the intercellular C4 pump and shifts the balance of carbon acquisition towards the C4 cycle. Carbon isotope ratios indicating a stronger C4 pathway are associated with warmer, drier environments, suggesting that incremental anatomical alterations can lead to the emergence of C4 physiology during local adaptation within metapopulations.

Defining the scope for altering rice leaf anatomy to improve photosynthesis: a modelling approach.

Leaf structure plays an important role in photosynthesis. However, the causal relationship and the quantitative importance of any single structural parameter to the overall photosynthetic performance of a leaf remains open to debate. In this paper, we report on a mechanistic model, eLeaf, which successfully captures rice leaf photosynthetic performance under varying environmental conditions of light and CO2 . We developed a 3D reaction-diffusion model for leaf photosynthesis parameterised using a range of imaging data and biochemical measurements from plants grown under ambient and elevated CO2 and then interrogated the model to quantify the importance of these elements. The model successfully captured leaf-level photosynthetic performance in rice. Photosynthetic metabolism underpinned the majority of the increased carbon assimilation rate observed under elevated CO2 levels, with a range of structural elements making positive and negative contributions. Mesophyll porosity could be varied without any major outcome on photosynthetic performance, providing a theoretical underpinning for experimental data. eLeaf allows quantitative analysis of the influence of morphological and biochemical properties on leaf photosynthesis. The analysis highlights a degree of leaf structural plasticity with respect to photosynthesis of significance in the context of attempts to improve crop photosynthesis.

Savanna fire regimes depend on grass trait diversity.

Grasses fuel most fires on Earth and strongly influence local fire behaviour through traits that determine how flammable they are. Therefore, grass communities that differ in their species and trait compositions give rise to significant spatial variation in savanna fire regimes across the world, which cannot be otherwise explained. Likewise, fire regimes are continuously modified by alterations to savanna grass community traits, through species introductions and climatic changes. However, current representation of grassy fuels in global fire models misses important variation and therefore limits predictive power. The inclusion of grass trait diversity in models, using remotely sensed trait proxies, for example, will greatly improve our ability to understand and project savanna fires and their roles in the Earth system.

Upregulation of C4 characteristics does not consistently improve photosynthetic performance in intraspecific hybrids of a grass.

C4 photosynthesis is thought to have evolved via intermediate stages, with changes towards the C4 phenotype gradually enhancing photosynthetic performance. This hypothesis is widely supported by modelling studies, but experimental tests are missing. Mixing of C4 components to generate artificial intermediates can be achieved via crossing, and the grass Alloteropsis semialata represents an outstanding study system since it includes C4 and non-C4 populations. Here, we analyse F1 hybrids between C3 and C4 , and C3 +C4 and C4 genotypes to determine whether the acquisition of C4 characteristics increases photosynthetic performance. The hybrids have leaf anatomical characters and C4 gene expression profiles that are largely intermediate between those of their parents. Carbon isotope ratios are similarly intermediate, which suggests that a partial C4 cycle coexists with C3 carbon fixation in the hybrids. This partial C4 phenotype is associated with C4 -like photosynthetic efficiency in C3 +C4 × C4 , but not in C3 × C4 hybrids, which are overall less efficient than both parents. Our results support the hypothesis that the photosynthetic gains from the upregulation of C4 characteristics depend on coordinated changes in anatomy and biochemistry. The order of acquisition of C4 components is thus constrained, with C3 +C4 species providing an essential step for C4 evolution.

Drought exposure leads to rapid acquisition and inheritance of herbicide resistance in the weed Alopecurus myosuroides.

Globally, herbicide resistance in weeds poses a threat to food security. Resistance evolves rapidly through the co-option of a suite of physiological mechanisms that evolved to allow plants to survive environmental stress. Consequently, we hypothesize that stress tolerance and herbicide resistance are functionally linked. We address two questions: (i) does exposure to stress in a parental generation promote the evolution of resistance in the offspring? (ii) Is such evolution mediated through non-genetic mechanisms? We exposed individuals of a grass weed to drought, and tested whether this resulted in herbicide resistance in the first generation. In terms of both survival and dry mass, we find enhanced resistance to herbicide in the offspring of parents that had been exposed to drought. Our results suggest that exposure of weeds to drought can confer herbicide resistance in subsequent generations, and that the mechanism conferring heritability of herbicide resistance is non-genetic.

Disparities among crop species in the evolution of growth rates: the role of distinct origins and domestication histories.

Growth rates vary widely among plants with different strategies. For crops, evolution under predictable and high-resource environments might favour rapid resource acquisition and growth, but whether this strategy has consistently evolved during domestication and improvement remains unclear. Here we report a comprehensive study of the evolution of growth rates based on comparisons among wild, landrace, and improved accessions of 19 herbaceous crops grown under common conditions. We also examined the underlying growth components and the influence of crop origin and history on growth evolution. Domestication and improvement did not affect growth consistently, that is growth rates increased or decreased or remained unchanged in different crops. Crops selected for fruits increased the physiological component of growth (net assimilation rate), whereas leaf and seed crops showed larger domestication effects on morphology (leaf mass ratio and specific leaf area). Moreover, climate and phylogeny contributed to explaining the effects of domestication and changes in growth. Crop-specific responses to domestication and improvement suggest that selection for high yield has not consistently changed growth rates. The trade-offs between morpho-physiological traits and the distinct origins and histories of crops accounted for the variability in growth changes. These findings have far-reaching implications for our understanding of crop performance and adaptation.

Crop origins explain variation in global agricultural relevance.

Human food production is dominated globally by a small number of crops. Why certain crops have attained high agricultural relevance while others have remained minor might partially stem from their different origins. Here, we analyse a dataset of 866 crops to show that seed crops and species originating from seasonally dry environments tend to have the greatest agricultural relevance, while phylogenetic affinities play a minor role. These patterns are nuanced by root and leaf crops and herbaceous fruit crops having older origins in the aseasonal tropics. Interestingly, after accounting for these effects, we find that older crops are more likely to be globally important and are cultivated over larger geographical areas than crops of recent origin. Historical processes have therefore left a pervasive global legacy on the food we eat today.

Fertile Crescent crop progenitors gained a competitive advantage from large seedlings.

Cereal domestication during the transition to agriculture resulted in widespread food production, but why only certain species were domesticated remains unknown. We tested whether seedlings of crop progenitors share functional traits that could give them a competitive advantage within anthropogenic environments, including higher germination, greater seedling survival, faster growth rates, and greater competitive ability.Fifteen wild grass species from the Fertile Crescent were grown individually under controlled conditions to evaluate differences in growth between cereal crop progenitors and other wild species that were never domesticated. Differences in germination, seedling survival, and competitive ability were measured by growing a subset of these species as monocultures and mixtures.Crop progenitors had greater germination success, germinated more quickly and had greater aboveground biomass when grown in competition with other species. There was no evidence of a difference in seedling survival, but seed size was positively correlated with a number of traits, including net assimilation rates, greater germination success, and faster germination under competition. In mixtures, the positive effect of seed mass on germination success and speed of germination was even more beneficial for crop progenitors than for other wild species, suggesting greater fitness. Thus, selection for larger seeded individuals under competition may have been stronger in the crop progenitors.The strong competitive ability of Fertile Crescent cereal crop progenitors, linked to their larger seedling size, represents an important ecological difference between these species and other wild grasses in the region. It is consistent with the hypothesis that competition within plant communities surrounding human settlements, or under early cultivation, benefited progenitor species, favoring their success as crops.

Traits explain sorting of C4 grasses along a global precipitation gradient.

Species distributions are closely associated with moisture availability, but the underlying mechanisms remain unresolved. Drought relations are especially important for plants such as C4 grasses that dominate seasonally dry ecosystems. Here, we test the hypothesis that C4 grass species sampled across global precipitation gradients show variation in survival under drought that can be explained by their traits.Our experiment subjected 18 C4 grass species to a lethal drought under controlled environmental conditions. The number of days until death was measured, along with root traits, senescence, and aspects of hydraulic function.We identified two strategies: Drought-avoiding species that stayed green as the water potential declined and drought-tolerating species that senesced more quickly but could extend survival via drought-tolerant meristems.Plants that stay-green for longer occupied drier habitats and had the longest survival under drought, facilitated by narrow root diameter and isohydric stomatal behavior. Plants that senesced quickly had thicker roots, an anisohydric strategy, and occupied wetter habitats.Global distributions of C4 grasses can be predicted by variation in rates of senescence, meristem survival, root traits, and stomatal strategy, showing the value of these traits for understanding plant distributions in relation to climate.

Developmental and biophysical determinants of grass leaf size worldwide.

One of the most notable ecological trends-described more than 2,300  years ago by Theophrastus-is the association of small leaves with dry and cold climates, which has recently been recognized for eudicotyledonous plants at a global scale1-3. For eudicotyledons, this pattern has been attributed to the fact that small leaves have a thinner boundary layer that helps to avoid extreme leaf temperatures4 and their leaf development results in vein traits that improve water transport under cold or dry climates5,6. However, the global distribution of leaf size and its adaptive basis have not been tested in the grasses, which represent a diverse lineage that is distinct in leaf morphology and that contributes 33% of terrestrial primary productivity (including the bulk of crop production)7. Here we demonstrate that grasses have shorter and narrower leaves under colder and drier climates worldwide. We show that small grass leaves have thermal advantages and vein development that contrast with those of eudicotyledons, but that also explain the abundance of small leaves in cold and dry climates. The worldwide distribution of leaf size in grasses exemplifies how biophysical and developmental processes result in convergence across major lineages in adaptation to climate globally, and highlights the importance of leaf size and venation architecture for grass performance in past, present and future ecosystems.

Low dispersal and ploidy differences in a grass maintain photosynthetic diversity despite gene flow and habitat overlap.

Geographical isolation facilitates the emergence of distinct phenotypes within a single species, but reproductive barriers or selection are needed to maintain the polymorphism after secondary contact. Here, we explore the processes that maintain intraspecific variation of C4 photosynthesis, a complex trait that results from the combined action of multiple genes. The grass Alloteropsis semialata includes C4 and non-C4 populations, which have coexisted as a polyploid series for more than 1 million years in the miombo woodlands of Africa. Using population genomics, we show that there is genome-wide divergence for the photosynthetic types, but the current geographical distribution does not reflect a simple habitat displacement scenario as the genetic clusters overlap, being occasionally mixed within a given habitat. Despite evidence of recurrent introgression between non-C4 and C4 groups, in both diploids and polyploids, the distinct genetic lineages retain their identity, potentially because of selection against hybrids. Coupled with strong isolation by distance within each genetic group, this selection created a geographical mosaic of photosynthetic types. Diploid C4 and non-C4 types never grew together, and the C4 type from mixed populations constantly belonged to the hexaploid lineage. By limiting reproductive interactions between photosynthetic types, the ploidy difference probably allows their co-occurrence, reinforcing the functional diversity within this species. Together, these factors enabled the persistence of divergent physiological traits of ecological importance within a single species despite gene flow and habitat overlap.

Estimating uncertainty: A Bayesian approach to modelling photosynthesis in C3 leaves.

The Farquhar-von Caemmerer-Berry (FvCB) model is extensively used to model photosynthesis from gas exchange measurements. Since its publication, many methods have been developed to measure, or more accurately estimate, parameters of this model. Here, we have created a tool that uses Bayesian statistics to fit photosynthetic parameters using concurrent gas exchange and chlorophyll fluorescence measurements whilst evaluating the reliability of the parameter estimation. We have tested this tool on synthetic data and experimental data from rice leaves. Our results indicate that reliable parameter estimation can be achieved whilst only keeping one parameter, Km , that is, Michaelis constant for CO2 by Rubisco, prefixed. Additionally, we show that including detailed low CO2 measurements at low light levels increases reliability and suggests this as a new standard measurement protocol. By providing an estimated distribution of parameter values, the tool can be used to evaluate the quality of data from gas exchange and chlorophyll fluorescence measurement protocols. Compared to earlier model fitting methods, the use of a Bayesian statistics-based tool minimizes human interaction during fitting, reducing the subjectivity which is essential to most existing tools. A user friendly, interactive Bayesian tool script is provided.

Resprouting grasses are associated with less frequent fire than seeders.

Plant populations persist under recurrent fire via resprouting from surviving tissues (resprouters) or seedling recruitment (seeders). Woody species are inherently slow maturing, meaning that seeders are confined to infrequent fire regimes. However, for grasses, which mature faster, the relationships between persistence strategy and fire regime remain unknown. Globally, we analysed associations between fire regimes experienced by hundreds of grass species and their persistence strategy, within a phylogenetic context. We also tested whether persistence strategies are associated with morphological and physiological traits. Resprouters were associated with less frequent fire than seeders. Whilst modal fire frequencies were similar (fire return interval of 4-6 yr), seeders were restricted to regions with more frequent fire than resprouters, suggesting that greater competition with long-lived resprouters restricts seeder recruitment and survival when fire is rare. Resprouting was associated with lower leaf N, higher C:N ratios and the presence of belowground buds, but was unrelated to photosynthetic pathway. Differences between the life histories of grasses and woody species led to a contrasting prevalence of seeders and resprouters in relation to fire frequency. Rapid sexual maturation in grasses means that seeder distributions, relative to fire regime, are determined by competitive ability and recruitment, rather than time to reproductive maturity.

Contrasted histories of organelle and nuclear genomes underlying physiological diversification in a grass species.

C4 photosynthesis evolved multiple times independently in angiosperms, but most origins are relatively old so that the early events linked to photosynthetic diversification are blurred. The grass Alloteropsis semialata is an exception, as this species encompasses C4 and non-C4 populations. Using phylogenomics and population genomics, we infer the history of dispersal and secondary gene flow before, during and after photosynthetic divergence in A. semialata. We further analyse the genome composition of individuals with varied ploidy levels to establish the origins of polyploids in this species. Detailed organelle phylogenies indicate limited seed dispersal within the mountainous region of origin and the emergence of a C4 lineage after dispersal to warmer areas of lower elevation. Nuclear genome analyses highlight repeated secondary gene flow. In particular, the nuclear genome associated with the C4 phenotype was swept into a distantly related maternal lineage probably via unidirectional pollen flow. Multiple intraspecific allopolyploidy events mediated additional secondary genetic exchanges between photosynthetic types. Overall, our results show that limited dispersal and isolation allowed lineage divergence, with photosynthetic innovation happening after migration to new environments, and pollen-mediated gene flow led to the rapid spread of the derived C4 physiology away from its region of origin.

High silicon concentrations in grasses are linked to environmental conditions and not associated with C4 photosynthesis.

The uptake and deposition of silicon (Si) as silica phytoliths is common among land plants and is associated with a variety of functions. Among these, herbivore defense has received significant attention, particularly with regard to grasses and grasslands. Grasses are well known for their high silica content, a trait which has important implications ranging from defense to global Si cycling. Here, we test the classic hypothesis that C4 grasses evolved stronger mechanical defenses than C3 grasses through increased phytolith deposition, in response to extensive ungulate herbivory ("C4 -grazer hypothesis"). Despite mixed support, this hypothesis has received broad attention, even outside the realm of plant biology. Because C3 and C4 grasses typically dominate in different climates, with the latter more abundant in hot, dry regions, we also investigated the effects of water availability and temperature on Si deposition. We compiled a large dataset of grasses grown under controlled environmental conditions. Using phylogenetically informed generalized linear mixed models and character evolution models, we evaluated whether photosynthetic pathway or growth condition influenced Si concentration. We found that C4 grasses did not show consistently elevated Si concentrations compared with C3 grasses. High temperature treatments were associated with increased concentration, especially in taxa adapted to warm regions. Although the effect was less pronounced, reduced water treatment also promoted silica deposition, with slightly stronger response in dry habitat species. The evidence presented here rejects the "C4 -grazer hypothesis." Instead, we propose that the tendency for C4 grasses to outcompete C3 species under hot, dry conditions explains previous observations supporting this hypothesis. These findings also suggest a mechanism via which anthropogenic climate change may influence silica deposition in grasses and, by extension, alter the important ecological and geochemical processes it affects.

The morphogenesis of fast growth in plants.

Growth rate represents a fundamental axis of life history variation. Faster growth associated with C4 photosynthesis and annual life history has evolved multiple times, and the resulting diversity in growth is typically explained via resource acquisition and allocation. However, the underlying changes in morphogenesis remain unknown. We conducted a phylogenetic comparative experiment with 74 grass species, conceptualising morphogenesis as the branching and growth of repeating modules. We aimed to establish whether faster growth in C4 and annual grasses, compared with C3 and perennial grasses, came from the faster growth of individual modules or higher rates of module initiation. Morphogenesis produces fast growth in different ways in grasses using C4 and C3 photosynthesis, and in annual compared with perennial species. C4 grasses grow faster than C3 species through a greater enlargement of shoot modules and quicker secondary branching of roots. However, leaf initiation is slower and there is no change in shoot branching. Conversely, faster growth in annuals than perennials is achieved through greater branching and enlargement of shoots, and possibly faster root branching. The morphogenesis of fast growth depends on ecological context, with C4 grasses tending to promote resource capture under competition, and annuals enhancing branching to increase reproductive potential.