Accurately representing the relationships between nitrogen supply and photosynthesis is crucial for reliably predicting carbon-nitrogen cycle coupling in Earth System Models (ESMs). Most ESMs assume positive correlations amongst soil nitrogen supply, leaf nitrogen content, and photosynthetic capacity. However, leaf photosynthetic nitrogen demand may influence the leaf nitrogen response to soil nitrogen supply; thus, responses to nitrogen supply are expected to be the largest in environments where demand is the greatest. Using a nutrient addition experiment replicated across 26 sites spanning four continents, we demonstrated that climate variables were stronger predictors of leaf nitrogen content than soil nutrient supply. Leaf nitrogen increased more strongly with soil nitrogen supply in regions with the highest theoretical leaf nitrogen demand, increasing more in colder and drier environments than warmer and wetter environments. Thus, leaf nitrogen responses to nitrogen supply are primarily influenced by climatic gradients in photosynthetic nitrogen demand, an insight that could improve ESM predictions.

Natural selection favors growth by selecting a combination of plant traits that maximize photosynthetic CO2 assimilation at the lowest combined carbon costs of resource acquisition and use. We quantified how soil nutrient availability, plant nutrient acquisition strategies, and aridity modulate the variability in plant costs of nutrient acquisition relative to water acquisition (β). We used an eco-evolutionary optimality framework and a global carbon isotope dataset to quantify β. Under low soil nitrogen-to-carbon (N : C) ratios, a mining strategy (symbioses with ectomycorrhizal and ericoid mycorrhizal fungi) reduced β by mining organic nitrogen, compared with a scavenging strategy (symbioses with arbuscular mycorrhizal fungi). Conversely, under high N : C ratios, scavenging strategies reduced β by effectively scavenging soluble nitrogen, compared with mining strategies. N2-fixing plants did not exhibit reduced β under low N : C ratios compared with non-N2-fixing plants. Moisture increased β only in plants using a scavenging strategy, reflecting direct impacts of aridity on the carbon costs of maintaining transpiration in these plants. Nitrogen and phosphorus colimitation further modulated β. Our findings provide a framework for simulating the variability of plant economics due to plant nutrient acquisition strategies in earth system models.

Plants respond to increasing atmospheric CO2 concentrations by reducing leaf nitrogen content and photosynthetic capacity-patterns that correspond with increased net photosynthesis and growth. Despite the longstanding notion that nitrogen availability regulates these responses, eco-evolutionary optimality theory posits that leaf-level responses to elevated CO2 are driven by leaf nitrogen demand for building and maintaining photosynthetic enzymes and are independent of nitrogen availability. In this study, we examined leaf and whole-plant responses of Glycine max L. (Merr) subjected to full-factorial combinations of two CO2, two inoculation, and nine nitrogen fertilization treatments. Nitrogen fertilization and inoculation did not alter leaf photosynthetic responses to elevated CO2. Instead, elevated CO2 decreased the maximum rate of ribulose-1,5-bisophosphate oxygenase/carboxylase (Rubisco) carboxylation more strongly than it decreased the maximum rate of electron transport for ribulose-1,5-bisphosphate (RuBP) regeneration, increasing net photosynthesis by allowing rate-limiting steps to approach optimal coordination. Increasing fertilization enhanced positive whole-plant responses to elevated CO2 due to increased below-ground carbon allocation and nitrogen uptake. Inoculation with nitrogen-fixing bacteria did not influence plant responses to elevated CO2. These results reconcile the role of nitrogen availability in plant responses to elevated CO2, showing that leaf photosynthetic responses are regulated by leaf nitrogen demand while whole-plant responses are constrained by nitrogen availability.

Many plant species form symbiotic associations with nitrogen-fixing bacteria. Through this symbiosis, plants allocate photosynthate belowground to the bacteria in exchange for nitrogen fixed from the atmosphere. This symbiosis forms an important link between carbon and nitrogen cycles in many ecosystems. However, the economics of this relationship under soil nitrogen availability gradients is not well understood, as plant investment toward symbiotic nitrogen fixation tends to decrease with increasing soil nitrogen availability. Here, we used a manipulation experiment to examine how costs of nitrogen acquisition vary under a factorial combination of soil nitrogen availability and inoculation with Bradyrhizobium japonicum in Glycine max L. (Merr.). We found that inoculation decreased belowground biomass carbon costs to acquire nitrogen and increased total leaf area and total biomass, but these patterns were only observed under low fertilization and were the result of increased plant nitrogen uptake and no change in belowground carbon allocation. These results suggest that symbioses with nitrogen-fixing bacteria reduce carbon costs of nitrogen acquisition by increasing plant nitrogen uptake, but only when soil nitrogen is low, allowing individuals to increase nitrogen allocation to structures that support aboveground growth. This pattern may help explain the prevalence of plants capable of forming these associations in less fertile soils and provides useful insight into understanding the role of nutrient acquisition strategy on plant nitrogen uptake across nitrogen availability gradients.

Interactions between carbon (C) and nitrogen (N) cycles in terrestrial ecosystems are simulated in advanced vegetation models, yet methodologies vary widely, leading to divergent simulations of past land C balance trends. This underscores the need to reassess our understanding of ecosystem processes, given recent theoretical advancements and empirical data. We review current knowledge, emphasising evidence from experiments and trait data compilations for vegetation responses to CO2 and N input, alongside theoretical and ecological principles for modelling. N fertilisation increases leaf N content but inconsistently enhances leaf-level photosynthetic capacity. Whole-plant responses include increased leaf area and biomass, with reduced root allocation and increased aboveground biomass. Elevated atmospheric CO2 also boosts leaf area and biomass but intensifies belowground allocation, depleting soil N and likely reducing N losses. Global leaf traits data confirm these findings, indicating that soil N availability influences leaf N content more than photosynthetic capacity. A demonstration model based on the functional balance hypothesis accurately predicts responses to N and CO2 fertilisation on tissue allocation, growth and biomass, offering a path to reduce uncertainty in global C cycle projections.

Repository contains data and analyses reported in Perkowski et al. (in prep), which reports findings from a growth chamber experiment that grew soybean (Glycine max L. (Merr)) seedlings under full factorial combinations of two CO2 treatments, two inoculation treatments, and nine soil nitrogen fertilization treatments. New in v1.1: remove unnecessary columns in data/NxCO2xI_data.csv

Data and scripts for Perkowski et al. (in prep) describing interactive effects of nitrogen fertilization and inoculation on carbon costs to acquire nitrogen in soybean (Glycine max).

The connection between soil nitrogen availability, leaf nitrogen, and photosynthetic capacity is not perfectly understood. Because these three components tend to be positively related over large spatial scales, some posit that soil nitrogen positively drives leaf nitrogen, which positively drives photosynthetic capacity. Alternatively, others posit that photosynthetic capacity is primarily driven by above-ground conditions. Here, we examined the physiological responses of a non-nitrogen-fixing plant (Gossypium hirsutum) and a nitrogen-fixing plant (Glycine max) in a fully factorial combination of light by soil nitrogen availability to help reconcile these competing hypotheses. Soil nitrogen stimulated leaf nitrogen in both species, but the relative proportion of leaf nitrogen used for photosynthetic processes was reduced under elevated soil nitrogen in all light availability treatments due to greater increases in leaf nitrogen content than chlorophyll and leaf biochemical process rates. Leaf nitrogen content and biochemical process rates in G. hirsutum were more responsive to changes in soil nitrogen than those in G. max, probably due to strong G. max investments in root nodulation under low soil nitrogen. Nonetheless, whole-plant growth was significantly enhanced by increased soil nitrogen in both species. Light availability consistently increased relative leaf nitrogen allocation to leaf photosynthesis and whole-plant growth, a pattern that was similar between species. These results suggest that the leaf nitrogen-photosynthesis relationship varies under different soil nitrogen levels and that these species preferentially allocated more nitrogen to plant growth and non-photosynthetic leaf processes, rather than photosynthesis, as soil nitrogen increased.

Plants respond to elevated atmospheric CO2 concentrations by reducing leaf nitrogen content and photosynthetic capacity – patterns that correspond with increased net photosynthesis rates, total leaf area, and total biomass. Nitrogen supply has been hypothesized to be the primary factor controlling these responses, as nitrogen availability limits net primary productivity globally. Recent work using evo-evolutionary optimality theory suggests that leaf photosynthetic responses to elevated CO2 are independent of nitrogen supply and are instead driven by leaf nitrogen demand to build and maintain photosynthetic enzymes, which optimizes resource allocation to photosynthetic capacity and maximizes allocation to growth. Here, Glycine max L. (Merr) seedlings were grown under two CO2 concentrations, with and without inoculation with Bradyrhizobium japonicum , and across nine soil nitrogen fertilization treatments in a full-factorial growth chamber experiment to reconcile the role of nitrogen supply and demand on leaf and whole-plant responses to elevated CO2. After seven weeks, elevated CO2 increased net photosynthesis rates despite reduced leaf nitrogen content and maximum rates of Ribulose-1,5-bisphosphate (RuBP) carboxylase/oxygenase (Rubisco) carboxylation and electron transport for RuBP regeneration. Effects of elevated CO2 on net photosynthesis and indices of photosynthetic capacity were independent of nitrogen fertilization and inoculation. However, increasing nitrogen fertilization enhanced positive effects of elevated CO2 on total leaf area and total biomass due to increased nitrogen uptake and reduced carbon costs to acquire nitrogen. Whole-plant responses to elevated CO2 were not modified by inoculation across the nitrogen fertilization gradient, as plant investment toward symbiotic nitrogen fixation was similar between CO2 treatments. These results indicate that leaf nitrogen demand to build and maintain photosynthetic enzymes drives leaf photosynthetic responses to elevated CO2, while nitrogen supply regulates whole-plant responses. Our findings build on previous work suggesting that terrestrial biosphere models may improve simulations of photosynthetic processes under future novel environments by adopting optimality principles.### Competing Interest StatementThe authors have declared no competing interest.

Representations of interactions between the C and N cycles in terrestrial ecosystems are now implemented in a majority of state-of-the-art Dynamic Global Vegetation Models (C-N models). Standard models for simulating the response of individual processes to changes in N availability have not yet emerged and widely used models have not been tested against the full diversity of empirical data. Large remaining model structural uncertainty has important implications for projections and hindcasts of the land C uptake.Here, we summarise the current state of global land C balance simulations by comparing C-N models to C-only models; summarise data from field surveys and experiments to elucidate the role of soil N in controlling photosynthesis and its acclimation, stoichiometry, allocation, and growth; and demonstrate how optimality principles can guide the representation of acclimation and allocation for simulating ecosystem responses to experimental treatments of CO2 and soil N – consistent with observations. Promising model results are achieved by assuming that the atmospheric environment, including CO2, is the principal driver for photosynthetic capacities and leaf N following optimality theory of photosynthetic acclimation (Prentice et al., 2014). In turn, the functional balance hypothesis (Bloom et al., 1985) yields accurate predictions for how soil N availability and CO2 influence allocation and growth in different tissues.Our results show how confronting new theoretical approaches to simulating ecosystem C-N interactions against the collective constraints from diverse types of observations can guide model development and potentially reduce the large uncertainty in global carbon cycle projections.ReferencesBloom, Arnold J, F Stuart Chapin, and Harold A Mooney. “Resource Limitation in Plants--An Economic Analogy” Annual Review of Ecology and Systematics, 16, no. 1 (1985): 363–92. https://doi.org/10.1146/annurev.es.16.110185.002051.Prentice, I. Colin, Ning Dong, Sean M. Gleason, Vincent Maire, and Ian J. Wright. “Balancing the Costs of Carbon Gain and Water Transport: Testing a New Theoretical Framework for Plant Functional Ecology.” Ecology Letters 17, 1 (2014): 82–91. https://doi.org/10.1111/ele.12211.

<p>Plants acclimate to increasing CO<sub>2</sub> by reducing leaf nutrient allocation and photosynthetic capacity at the leaf level, a response that often occurs alongside growth stimulation at the whole plant level. Nutrient limitation has been hypothesized to be the primary mechanism driving leaf and whole plant acclimation responses to CO<sub>2</sub>, as nutrient availability commonly limits primary productivity and may decrease with increasing CO<sub>2</sub> over time. However, recent work leveraging photosynthetic least-cost theory indicates that these acclimation responses may instead be the result of optimal resource investment toward photosynthetic capacity, which maximizes nutrient allocation to whole plant growth. Acclimation responses to CO<sub>2</sub> may also vary in species with different nutrient acquisition strategies, but few studies have examined these responses across a soil nitrogen availability gradient and in species with different nutrient acquisition strategies. To test whether nutrient limitation or optimal leaf resource investment controls leaf and whole plant acclimation responses to CO<sub>2</sub> and how nutrient acquisition strategy modifies these responses, we grew <em>Glycine max</em> L. (Merr) seedlings under two atmospheric CO<sub>2</sub> levels, with and without <em>Bradyrhizobium japonicum</em> inoculation, and across nine soil nitrogen fertilization treatments in a full factorial growth chamber experiment. After seven weeks, <em>G. max</em> demonstrated a strong downregulation in leaf nitrogen content, <em>V</em><sub>cmax25</sub>, and <em>J</em><sub>max25</sub> under elevated CO<sub>2</sub>, patterns that were not causally linked to changes in soil nitrogen fertilization or inoculation treatment. A relatively stronger downregulation in leaf nitrogen content than <em>V</em><sub>cmax25</sub> increased the proportion of leaf nitrogen content allocated to photosynthesis, while a relatively stronger downregulation in <em>V</em><sub>cmax25</sub> than <em>J</em><sub>max25</sub> stimulated <em>J</em><sub>max25</sub>:<em>V</em><sub>cmax25</sub> under elevated CO<sub>2</sub>. These leaf acclimation responses to elevated CO<sub>2</sub> corresponded with strong stimulations in total leaf area and total biomass, a pattern that was generally stronger with increasing fertilization and in inoculated pots. Whole plant acclimation responses to CO<sub>2</sub> were driven by reductions in the cost of acquiring nitrogen with increasing fertilization and inoculation. Overall, these results provide strong support for patterns expected from photosynthetic least-cost theory, showing that optimal resource investment is the primary mechanism governing <em>G. max</em> acclimation responses to elevated CO<sub>2</sub>.</p>

Plant nitrogen acquisition requires carbon to be allocated belowground to build roots and sustain microbial associations. This carbon cost to acquire nitrogen varies by nitrogen acquisition strategy; however, the degree to which these costs vary due to nitrogen availability or demand has not been well tested under controlled conditions. We grew a species capable of forming associations with nitrogen-fixing bacteria (Glycine max) and a species not capable of forming such associations (Gossypium hirsutum) under four soil nitrogen levels to manipulate nitrogen availability and four light levels to manipulate nitrogen demand in a full-factorial greenhouse experiment. We quantified carbon costs to acquire nitrogen as the ratio of total root carbon to whole-plant nitrogen within each treatment combination. In both species, light availability increased carbon costs due to a larger increase in root carbon than whole-plant nitrogen, while nitrogen fertilization generally decreased carbon costs due to a larger increase in whole-plant nitrogen than root carbon. Nodulation data indicated that G. max shifted relative carbon allocation from nitrogen fixation to direct uptake with increased nitrogen fertilization. These findings suggest that carbon costs to acquire nitrogen are modified by changes in light and nitrogen availability in species with and without associations with nitrogen-fixing bacteria.