Photorespiration Research Articles

In silico analysis and functional validation of sinf-yc genes in foxtail millet setaria italica (l.)

Nuclear transcription factor YCs (NF-YCs) are universally involved in plant development and abiotic stress response. As a typical C4 crop, foxtail millet has significant research value for studies of functional genes and mechanisms of plant stress resistance. To further investigate the correlation between the structure and function of the SiNF-YC genes in foxtail millet (Setaria italica), bioinformatic analysis of the SiNF-YCs on the subcellular localization, physicochemical properties, and cis acting elements were performed in this study. The results showed that a total of 12 SiNF-YC subfamily genes that distributed on chromosomes 1-9 were identified from the foxtail millet genome, which encoding 129-469 amino acids. The subcellular localization prediction results showed that the proteins encoded by this gene subfamily are mainly located in the nucleus, with a few being extracellular secreted. The multiple cis-elements related to drought resistance and light response are contained in the gene promoter region. Through haplotype analysis on the phenotypes of drought treated and heading dates that from five different regions, obvious haplotypes were identified except for SiNF-YC8 and SiNF-YC14. At last, drought regulation related gene SiNF-YC5 and heading date regulation related gene SiNF-YC10 were selected to explore superior gene haplotypes, laying a foundation for further exploring the functional genes involved in drought resistance and development within this gene family. Bangladesh j. Bot. 53(3): 535-543, 2024 (September)

Drought and heat stress interactions modify photorespiration and hydrogen peroxide content in Silver fir.

Photorespiration (PR) greatly reduces net carbon assimilation in trees (by c. 25%), but has received recent attention particular for its potential role in stress-signaling through the accumulation of hydrogen peroxide (H2O2), a stress signaling agent. Despite an increasing frequency of drought and heat events affecting forests worldwide, little is known about how concurrent abiotic stressors may interact to affect PR and subsequent H2O2 accumulation in trees. Here, we sought to identify how drought and a compounded one-day heat treatment individually and interactively affect PR (determined under variable O2) in Abies alba Mill. seedlings. Additionally, we quantified foliar H2O2 accumulation and enzymatic scavenging via peroxidase in relation to PR rates. We found drought stress to slightly increase PR (+5.2%) during mild-drought (12 days, Ψmd = -0.85 MPa), but ultimately to decrease PR (-13.6%) during severe-drought (26 days, Ψmd = -1.70 MPa) compared to the control, corresponding to increasing non-stomatal limitations of photosynthesis (i.e., decreased electron transport rate). The response of PR to heat stress was dependent on soil water availability as heat stress increased PR in control seedlings (+37.8%), but not in drought-stressed seedlings. Decreased PR during severe-drought corresponded to ~2x lower foliar H2O2 compared to the control. Despite increased PR under heat stress in control seedlings, foliar H2O2 decreased to near-zero likely due to enhanced scavenging as observed in ~2x greater peroxidase activity. Our results demonstrate that carbon loss to PR during drought stress can be highly dynamic, depending on the severity of soil dehydration. Additionally, increased PR under abiotic stress does not necessarily lead to accumulated H2O2, as tight regulation by scavenging enzymes instead minimize oxidative stress, reducing stress-signaling potential.

Diet in high mediaeval Florence through stable isotope analysis of carbon, nitrogen and sulphur

In this paper, we aim to reconstruct the dietary habits of supposedly lower rank nobles or middle-class High Middle Ages individuals recovered from the cloister arcade of San Pier Scheraggio within the Uffizi Museum complex in Florence, Italy. Notably, the High Middle Ages was a period of cultural and social changes, which is partly reflected in the dietary habits, as suggested by historical sources. Here we apply stable carbon, nitrogen, and sulphur isotope analysis on humans (n = 34) and animals (n = 13) from San Pier Scheraggio to directly investigate food consumption in this peculiar assemblage. The diet of human individuals was based on terrestrial C3 sources without clear contribution of marine fish (δ13C mean and 1SD: −19.5 ± 0.7 ‰; δ15N mean and 1SD: 9.6 ± 0.4 ‰; δ34S values are mean and 1SD: 7.6 ± 1.2 ‰). The comparison with animal and human samples from other Italian Middle Ages contexts shows that the overall diet of the population buried at San Pier Scheraggio is in line with that of other mediaeval communities in Italy, although with a generally higher contribution of terrestrial animal products. Our data seem to suggest that at the site there was no dietary differentiation concerning age at death or biological sex of the individuals. Some differences, however, can be outlined, for example, in the contribution of C4 crops. In addition to this, we identify two individuals as possible non-locals.

Utilizing a high-throughput visualization screening technology to develop a genetically encoded biosensor for monitoring 5-aminolevulinic acid production in engineered Escherichia coli

5-Aminolevulinic acid (5-ALA) is a non-protein amino acid widely used in agriculture, animal husbandry and medicine. Currently, microbial cell factories are a promising production pathway, but the lack of high-throughput fermentation strain screening tools often hinders the exploration of engineering strategies to increase cell factory yields. Here, mutant AC103-3H was screened from libraries of saturating mutants after response-specific engineering of the transcription factor AsnC of L-asparagine (Asn). Based on mutant AC103-3H, a whole-cell biosensor EAC103-3H with a specific response to 5-ALA was constructed, which has a linear dynamic detection range of 1–12 mM and a detection limit of 0.094 mM, and can be used for in situ screening of potential high-producing 5-ALA strains. With its support, overexpression of the C5 pathway genes using promoter engineering assistance resulted in a 4.78-fold enhancement of 5-ALA production in the engineered E. coli. This study provides an efficient strain screening tool for exploring approaches to improve the 5-ALA productivity of engineered strains.

PredPSP: a novel computational tool to discover pathway-specific photosynthetic proteins in plants.

Photosynthetic proteins play a crucial role in agricultural productivity by harnessing light energy for plant growth. Understanding these proteins, especially within C3 and C4 pathways, holds promise for improving crops in challenging environments. Despite existing models, a comprehensive computational framework specifically targeting plant photosynthetic proteins is lacking. The underutilization of plant datasets in computational algorithms accentuates the gap this study aims to fill by introducing a novel sequence-based computational method for identifying these proteins. The scope of this study encompassed diverse plant species, ensuring comprehensive representation across C3 and C4 pathways. Utilizing six deep learning models and seven shallow learning algorithms, paired with six sequence-derived feature sets followed by feature selection strategy, this study developed a comprehensive model for prediction of plant-specific photosynthetic proteins. Following 5-fold cross-validation analysis, LightGBM with 65 and 90 LGBM-VIM selected features respectively emerged as the best models for C3 (auROC: 91.78%, auPRC: 92.55%) and C4 (auROC: 99.05%, auPRC: 99.18%) plants. Validation using an independent dataset confirmed the robustness of the proposed model for both C3 (auROC: 87.23%, auPRC: 88.40%) and C4 (auROC: 92.83%, auPRC: 92.29%) categories. Comparison with existing methods demonstrated the superiority of the proposed model in predicting plant-specific photosynthetic proteins. This study further established a free online prediction server PredPSP ( https://iasri-sg.icar.gov.in/predpsp/ ) to facilitate ongoing efforts for identifying photosynthetic proteins in C3 and C4 plants. Being first of its kind, this study offers valuable insights into predicting plant-specific photosynthetic proteins which holds significant implications for plant biology.

Investigating the cis-regulatory basis of C3 and C4 photosynthesis in grasses at single-cell resolution

While considerable knowledge exists about the enzymes pivotal for C4 photosynthesis, much less is known about the cis-regulation important for specifying their expression in distinct cell types. Here, we use single-cell-indexed ATAC-seq to identify cell-type-specific accessible chromatin regions (ACRs) associated with C4 enzymes for five different grass species. This study spans four C4 species, covering three distinct photosynthetic subtypes: Zea mays and Sorghum bicolor (NADP-dependent malic enzyme), Panicum miliaceum (NAD-dependent malic enzyme), Urochloa fusca (phosphoenolpyruvate carboxykinase), along with the C3 outgroup Oryza sativa. We studied the cis-regulatory landscape of enzymes essential across all C4 species and those unique to C4 subtypes, measuring cell-type-specific biases for C4 enzymes using chromatin accessibility data. Integrating these data with phylogenetics revealed diverse co-option of gene family members between species, showcasing the various paths of C4 evolution. Besides promoter proximal ACRs, we found that, on average, C4 genes have two to three distal cell-type-specific ACRs, highlighting the complexity and divergent nature of C4 evolution. Examining the evolutionary history of these cell-type-specific ACRs revealed a spectrum of conserved and novel ACRs, even among closely related species, indicating ongoing evolution of cis-regulation at these C4 loci. This study illuminates the dynamic and complex nature of cis-regulatory elements evolution in C4 photosynthesis, particularly highlighting the intricate cis-regulatory evolution of key loci. Our findings offer a valuable resource for future investigations, potentially aiding in the optimization of C3 crop performance under changing climatic conditions.

Response of Bacterial Community Structure and Function in Rhizosphere Soil on the Photosynthesis of Selected Plant Types C3 and C4 under Bis(2,4,6-tribromophenoxy) Ethane Exposure

This study investigated the response of a bacterial community’s structure and function in the rhizosphere soil of C3 and C4 plants under bis(2,4,6-tribromophenoxy) ethane (BTBPE) exposure. The bacterial community composition was determined using 16S rRNA sequencing, while FAPROTAX and PICRUSt 2 were employed for functional predictions. Results showed significant differences between C3 and C4 plants in terms of bacterial community structure. C3 plants exhibited higher abundances of Proteobacteria, Bacteroidetes at the phylum level and Sphingomicrobium at the genus level, compared to C4 plants. Conversely, C4 plants had higher abundances of Actinobacteria and Patescibacteria at the phylum level and Nocardioides at the genus level. LEfSe and function prediction analyses revealed that the rhizosphere soil bacteria in C3 plants exhibited significantly higher enrichment in nitrogen fixation functions (p < 0.05), whereas C4 plants showed a significantly higher relative abundance of bacteria and functions related to organic pollutant degradation (p < 0.05). These findings suggest that the rhizosphere soil bacteria of C3 plants exhibit a stronger response to BTBPE exposure in nitrogen metabolism-related processes, while C4 plants possess superior biodegradation ability compared to C3 plants.

Mesophyll airspace unsaturation drives C4 plant success under vapor pressure deficit stress

A fundamental assumption in plant science posits that leaf air spaces remain vapor saturated, leading to the predominant view that stomata alone control leaf water loss. This concept has been pivotal in photosynthesis and water-use efficiency research. However, recent evidence has refuted this longstanding assumption by providing evidence of unsaturation in the leaf air space of C3 plants under relatively mild vapor pressure deficit (VPD) stress. This phenomenon represents a nonstomatal mechanism restricting water loss from the mesophyll. The potential ubiquity and physiological implications of this phenomenon, its driving mechanisms in different plant species and habitats, and its interaction with other ecological adaptations have. In this context, C4 plants spark particular interest for their importance as crops, bundle sheath cells' unique anatomical characteristics and specialized functions, and notably higher water-use efficiency relative to C3 plants. Here, we confirm reduced relative humidities in the substomatal cavity of the C4 plants maize, sorghum, and proso millet down to 80% under mild VPD stress. We demonstrate the critical role of nonstomatal control in these plants, indicating that the role of the CO2 concentration mechanism in CO2 management at a high VPD may have been overestimated. Our findings offer a mechanistic reconciliation between discrepancies in CO2 and VPD responses reported in C4 species. They also reveal that nonstomatal control is integral to maintaining an advantageous microclimate of relatively higher CO2 concentrations in the mesophyll air space of C4 plants for carbon fixation, proving vital when these plants face VPD stress.

Characterization, Phylogenetic Analyses, and Pathogenicity of Eutiarosporella dactylidis on Sorghum in China.

Sorghum, the fifth-largest cereal crop globally and a C4 crop, mainly grows in arid and semi-arid areas. In 2021-2023, a new foliar disease of sorghum occurred in China. The diseased leaves showed water-soaked symptoms in the leaf tip and margins, resulting in half- and full-leaf desiccation and necrosis, thus affecting plant photosynthesis. A total of 24 Eutiarosporella strains were isolated from symptomatic leaves. Based on morphological characteristics and multi-locus phylogenetic analysis involving ITS, LSU, and EF1-α sequences, and the pathogenicity test, the pathogen of sorghum causing leaf blight in China was identified as Eutiarosporella dactylidis. The virulence of all E. dactylidis strains was evaluated using the spray-mycelium method. Different strains showed significantly different pathogenicities toward a susceptible cultivar, Longza 10, with disease indexes ranging from 23.76 to 60.37. This study first reported leaf blight of sorghum caused by E. dactylidis and named it "Eutiarosporella leaf blight", which provides a theoretical basis for farmers in disease management.

Regulatory Network of the Late-Recruited Primary Decarboxylase C4NADP-ME in Sugarcane.

In agronomically important C4 grasses, efficient CO2 delivery to Rubisco is facilitated by NADP-malic enzyme (C4NADP-ME), which decarboxylates malate in bundle sheath cells. However, understanding the molecular regulation of the C4NADP-ME gene in sugarcane (Saccharum spp.) is hindered by its complex genetic background. Enzymatic activity assays demonstrated that decarboxylation in sugarcane Saccharum spontaneum predominantly relies on the NADP-ME pathway, similar to sorghum (Sorghum bicolor) and maize (Zea mays). Comparative genomics analysis revealed the recruitment of eight core C4 shuttle genes, including C4NADP-ME (SsC4NADP-ME2), in the C4 pathway of sugarcane. Contrasting to sorghum and maize, the expression of SsC4NADP-ME2 in sugarcane is regulated by different transcription factors (TFs). We propose a gene regulatory network for SsC4NADP-ME2, involving candidate TFs identified through gene co-expression analysis and yeast one-hybrid experiment. Among these, ABA INSENSITIVE5 (ABI5) was validated as the predominant regulator of SsC4NADP-ME2 expression, binding to a G-box within its promoter region. Interestingly, the core element ACGT within the regulatory G-box was conserved in sugarcane, sorghum, maize, and rice (Oryza sativa), suggesting an ancient regulatory code utilized in C4 photosynthesis. This study offers insights into SsC4NADP-ME2 regulation, crucial for optimizing sugarcane as a bioenergy crop.

Sources and transport of organic matter in the Ganges-Brahmaputra-Meghna River system of Bengal Basin, Bangladesh

This study aimed to understand the sources and transport mechanism of organic matter (OM) in the Ganges-Brahmaputra-Meghna (GBM) river system in Bangladesh. We conducted analyses of total organic carbon (TOC), total nitrogen (TN), their stable isotopes (δ13C and δ15N), and sediment grain size. The results reveal a heterogeneous mixture of OM derived from terrestrial plants, aquatic environments, and anthropogenic sources. The Brahmaputra River exhibited higher concentrations of TOC and TN, with δ13C and δ15N values indicating that the OM is primarily sourced from C3 plants. Conversely, the Ganges River demonstrated lower TOC levels and higher isotopic values, reflecting significant anthropogenic inputs. The Lower Meghna showed a mixture of terrestrial and marine sources. Variations in the TOC/TN ratios across the river system underscore the complex interplay between natural and anthropogenic factors. Additionally, sediment grain size plays a crucial role, with finer sediments in the Brahmaputra River associated with increased OM concentrations, while coarser sediments in the Ganges River correlate with lower TOC and TN levels.

Does the response of Rubisco and photosynthesis to elevated [CO2] change with unfavourable environmental conditions?

Climate change due to anthropogenic CO2 emissions affects plant performance globally. To improve crop resilience, we need to understand the effects of elevated CO2 concentration (e[CO2]) on CO2 assimilation and Rubisco biochemistry. However, the interactive effects of e[CO2] and abiotic stress are especially unclear. This study analyses the CO2 effect on photosynthetic capacity under different water availability and temperature conditions in 42 different crop species, varying in functional group, photosynthetic pathway and phenological stage. We analysed close to 3000 data points extracted from 120 published manuscripts. For C3 species, e[CO2] increases net photosynthesis and intercellular [CO2], while reducing stomatal conductance and transpiration. Vmaxc, Rubisco in vitro extractable maximal activity and content also decrease with e[CO2] in C3 species, while C4 crops are less responsive to e[CO2]. The interaction with drought and/or heat stress does not significantly alter these photosynthetic responses, indicating that the photosynthetic capacity of stressed plants responds to e[CO2]. Moreover, e[CO2] has strong effect on the photosynthetic capacity of grasses mainly in the final stages of development. This study provides insight into the intricate interactions within the plant photosynthetic apparatus under the influence of climate change, enhancing the understanding of mechanisms governing plant responses to environmental parameters.

One place, many worlds – nutrition strategies of human groups inhabiting the site 3 in Miechów (southern Poland) since the Neolithic until the Iron Age

Abstract Studies on the ratios of stable isotopes of carbon and nitrogen constitute a research tool commonly applied in bioarchaeology to reconstruct human diets, including the origins of proteins consumed, the relationship between plant and animal foods, weaning processes, and parameters of the environment from which the food was obtained. Within the comprehensive analysis of the excavation results carried out at site 3 in Miechów (approximately 30 km north of Kraków), isotope analyses of δ13C and δ15N were performed on the human groups that inhabited the site from the Neolithic to the Iron Age. In total, 41 skeletons were investigated. The analyses aimed to hypothesise the nutritional strategies of communities of successive periods and to compare the knowledge obtained with current data and interpretations on food management in prehistoric communities of Central Europe. The results indicate a general predominance of a terrestrial diet, with plant-based foods prevailing, although not conspicuously. However, there are indications of slightly varying nutritional strategies among successive human groups. These differences are related to the contribution of animal protein and the relationship between C3 and C4 plants. For instance, in the Middle Neolithic (4th millennium BC), a greater consumption of meat and/or cereals from manured fields can be postulated compared to the Early Neolithic (late 6th and 5th millennia BC). On the other hand, the proportion of meat in the diet of Early Bronze Age inhabitants (2nd millennium BC) was possibly the lowest in the entire history of site occupation. This proportion returned to near-Neolithic levels in the developed Bronze Age and Iron Age (1st millennium BC). Regarding plant food, the results indicate greater variability in the plants consumed in the Bronze and Iron Ages. Levels of δ13C imply some contribution from C4 plants, that is, millet, during these periods. This aligns with recently formulated theories suggesting the post-Neolithic beginnings of millet cultivation and consumption in Central Europe.

Identifying the botanical origin of alcohol using 2H SNIF NMR: A case study of “polish vodka” PGI

In this study, we utilized 2H SNIF NMR and chemometric techniques to differentiate the botanical origin of raw materials used in vodka production in Poland, specifically focusing on plants with C3 metabolism such as grain, potato, and sugar beet. Additionally, for the first time, mixtures of alcohols from different C3 plants were analysed to detect adulteration. Our goal was to determine if it is possible to detect the addition of a different raw material in vodka made from a mixture of alcohols derived from C3 plants. Significant isotopic differences were confirmed using analysis of variance and Tukey's tests. Linear relationships in grain-potato, grain-sugar beet, and beet-potato mixtures enabled composition determination. The detectability threshold for adulterants ranged from 10 % to 50 %, depending on the type of raw material. Our findings suggest that 2H SNIF-NMR is an effective tool for authenticating vodka and detecting adulteration in products marketed as “Polish vodka.”

Identification and expression profiling of heteroglycan glucosidase 1 enzyme of Cenchrus americanus

Maltose metabolism is a critical process during plant growth, which provides energy during development and reproduction. To investigate maltose metabolism in C4 plants, we identified and analyzed the expression of the heteroglycan glucosidase 1 (HGL1) enzyme from Cenchrus americanus (L.) Morrone. The sequenced cDNA of CaHGL1 (3469 bp) encoded for a deduced protein of 1047 aa. Transmembrane topology revealed that CaHGL1 is a membrane-bound protein that comprises a signal peptide and a transmembrane helix. The promoter of CaHGL1 contains cis-elements related to the responses to light, abiotic stress and phytohormones. Real-time PCR revealed high expression in inflorescence and roots during the vegetative stage. Moreover, phytohormone treatments caused an activation of CaHGL1 expression in the seedling root and shoot by ABA, cytokinin, and BL, and an inhibition by JA in the seedling root and shoot. However, treatment with GA and IAA caused an activation in the CaHGL1 expression only in the shoot. Stress treatment induced the expression of CaHGL1 under drought, cold, and salt stress. The results of the current study give insight into the activity of the HGL1 enzyme in maltose metabolism under abiotic stress, which can aid in understanding the different metabolic pathways in Cenchrus americanus under stress.

Response of maize (Zea mays L.) on yield, physiology and stomatal behaviour under two different elevated CO<sub>2</sub> concentrations. Do these anatomical changes affect the physiology of the C4 crop plant under high CO<sub>2</sub> conditions?

Response of maize (Zea mays L.) on yield, physiology and stomatal behaviour under two different elevated CO<sub>2</sub> concentrations. Do these anatomical changes affect the physiology of the C4 crop plant under high CO<sub>2</sub> conditions?

Taxonomic and stable isotope analyses of mammal remains from the Lateglacial site of Grotta Polesini (central Italy): Paleoenviromental implications

ABSTRACTGrotta Polesini is one of the most famous paleontological and archaeological sites of central Italy, which testifies to its human occupation during the Lateglacial. The site comprises a cave system where systematic excavation campaigns have been carried out since the 1950s. In 1974, 656 mammal remains were collected but never studied. This fossil collection is here described for the first time through taxonomic and stable isotope analyses of the enamel of selected mammal teeth. The aim is to reconstruct the paleoenvironmental and climatic conditions of the site and to offer new information on terrestrial ecosystems during the Lateglacial in central Italy. The faunal assemblage studied herein, in addition to other species reported in previous works, suggests cold climate conditions. We also describe a right radius of an adult individual of Homo sapiens, increasing the human fossil record of the site. Carbon isotope data point to a scenario dominated by C3 plants in open and dry habitats, such as grasslands and steppes, in accordance with the pollen data from central Italy. The oxygen isotope data suggest the use of water resources with a local origin, i.e. local precipitation and surface waters with a provenance from the nearby Apennine chain. The ecology of the taxa influenced the oxygen isotope values, especially in the case of semi‐obligate to non‐obligate drinker species.

Native arbuscular mycorrhizal fungi drive ecophysiology through phenotypic integration and functional plasticity under the Sonoran desert conditions.

Knowledge is scarce to what extent environmental drivers and native symbiotic fungi in soil induce abrupt (short-term), systemic (multiple traits), or specific (a subset of traits) shifts in C3 plants' ecophysiological/mycorrhizal responses. We cultivated an emblematic native C3 species (Capsicum annuum var. glabriusculum, "Chiltepín") to look at how the extreme heat of the Sonoran desert, sunlight regimes (low = 2, intermediate = 15, high = 46 mol m2 d-1) and density of native arbuscular mycorrhizal fungi in soil (low AMF = 1% v/v, high AMF = 100% v/v), drive shifts on mycorrhizal responses through multiple functional traits (106 traits). The warming thresholds were relentlessly harsh even under intensive shade (e.g. superheat maximum thresholds reached ranged between 47-63°C), and several pivotal traits were synergistically driven by AMF (e.g. photosynthetic capacity, biomass gain/allometry, and mycorrhizal colonization traits); whereas concurrently, sunlight regimes promoted most (76%) alterations in functional acclimation traits in the short-term and opposite directions (e.g. survival, phenology, photosynthetic, carbon/nitrogen economy). Multidimensional reduction analysis suggests that the AMF promotes a synergistic impact on plants' phenotypic integration and functional plasticity in response to sunlight regimes; however, complex relationships among traits suggest that phenotypic variation determines the robustness degree of ecophysiological/mycorrhizal phenotypes between/within environments. Photosynthetic canopy surface expansion, Rubisco activity, photosynthetic nitrogen allocation, carbon gain, and differential colonization traits could be central to plants' overall ecophysiological/mycorrhizal fitness strengthening. In conclusion, we found evidence that a strong combined effect among environmental factors in which AMF are key effectors could drive important trade-offs on plants' ecophysiological/mycorrhizal fitness in the short term.

Instantaneous growth: a compact measure of efficient carbon and nitrogen allocation in leaves and roots of C3 and C4 plants.

Elucidating plant functions and identifying crop productivity bottlenecks requires the accurate quantification of their performance. This task has been attained through photosynthetic models. However, their traditional focus on the leaf's capacity to uptake CO2 is becoming increasingly restrictive. Advanced bioengineering of C3 plants has made it possible to increase rates of CO2 assimilation by packing photosynthetic structures more densely within leaves. The operation of mechanisms that concentrate CO2 inside leaves can boost rates of assimilation while requiring a lower investment in carboxylating enzymes. Therefore, whether in the context of spontaneous plants or modern manipulation, considering trade-offs in resource utilization efficiency emerges as a critical necessity. I've developed a concise and versatile analytical model that simulates concurrent leaf and root growth by balancing instantaneous fluxes of carbon and nitrogen. Carbon is made available by leaf photosynthesis, encompassing all types of biochemistries, while nitrogen is either taken up by roots or remobilized after senescence. The allocation of leaf nitrogen between light or carbon reactions was determined using a fitting algorithm: growth maximisation was the only reliable fitting goal. Both the leaf nitrogen pool and the root-to-leaf ratio responded realistically to various environmental drivers (CO2 concentration, light intensity, soil nitrogen), replicating trends typically observed in plants. Furthermore, modifying the strength of CO2 concentrating mechanisms proved sufficient to alter the root-to-leaf ratio between C3 and C4 types. This direct and mechanistic one-to-one link convincingly demonstrates, for the first time, the functional dependence of a morphological trait on a single biochemical property.

Stomatal response to VPD in C4 plants with different biochemical sub-pathways.

C4 NAD-malic enzyme (NAD-ME) species occurs in drier regions and exhibit different drought responses compared to C4 NADP-malic enzyme (NADP-ME) species. However, a physiological mechanism explaining the geographical discrepancies remains uncertain. This study examined gas exchange patterns that might explain different distributions observed between two subtypes of C4 photosynthesis. We measured the response of leaf gas exchange to vapour pressure deficit (VPD) and CO2 in plants from six distinct C4 clades having closely related NAD-ME and NADP-ME species using a Li-Cor 6400 gas exchange system. We found that NAD-ME species exhibited greater relative reductions in stomatal conductance with increases in VPD than NADP-ME species but observed no consistent subtype differences in C4 cycle activity as indicated by the initial slope of the A response to intercellular CO2 concentration. Based on these results, we hypothesise the greater response of gs to increasing VPD may enable NAD-ME plants to outperform NADP-ME plants in hot, dry environments where VPD is normally high.