Precision genome editing heralds rapid de novo domestication for new crops.

The de novo domestication has the potential to rapidly capitalize on desirable traits of wild plants. In this issue of Cell, Yu et al. report a route of de novo domestication of an allotetraploid rice, heralding the creation of a novel staple food crop to support global food security.

Meeting the global food demand of the future by engineering crop photosynthesis and yield potential.

Increase in demand for our primary foodstuffs is outstripping increase in yields, an expanding gap that indicates large potential food shortages by mid-century. This comes at a time when yield improvements are slowing or stagnating as the approaches of the Green Revolution reach their biological limits. Photosynthesis, which has been improved little in crops and falls far short of its biological limit, emerges as the key remaining route to increase the genetic yield potential of our major crops. Thus, there is a timely need to accelerate our understanding of the photosynthetic process in crops to allow informed and guided improvements via in-silico-assisted genetic engineering. Potential and emerging approaches to improving crop photosynthetic efficiency are discussed, and the new tools needed to realize these changes are presented.

Perspectives on improving photosynthesis to increase crop yield.

Improving photosynthesis, the fundamental process by which plants convert light energy into chemical energy, is a key area of research with great potential for enhancing sustainable agricultural productivity and addressing global food security challenges. This perspective delves into the latest advancements and approaches aimed at optimizing photosynthetic efficiency. Our discussion encompasses the entire process, beginning with light harvesting and its regulation and progressing through the bottleneck of electron transfer. We then delve into the carbon reactions of photosynthesis, focusing on strategies targeting the enzymes of the Calvin-Benson-Bassham (CBB) cycle. Additionally, we explore methods to increase CO2 concentration near the Rubisco, the enzyme responsible for the first step of CBB cycle, drawing inspiration from various photosynthetic organisms, and conclude this section by examining ways to enhance CO2 delivery into leaves. Moving beyond individual processes, we discuss two approaches to identifying key targets for photosynthesis improvement: systems modeling and the study of natural variation. Finally, we revisit some of the strategies mentioned above to provide a holistic view of the improvements, analyzing their impact on nitrogen use efficiency and on canopy photosynthesis.

Mechanisms controlling metabolite concentrations of the Calvin Benson Cycle.

Maintaining proper metabolite levels in a complex metabolic network is crucial for maintaining a high flux through the network. In this paper, we discuss major regulatory mechanisms over the Calvin Benson Cycle (CBC) with regard to their roles in conferring homeostasis of metabolite levels in CBC. These include: 1) Redox regulation of enzymes in the CBC on one hand ensures that metabolite levels stay above certain lower bounds under low light while on the other hand increases the flux through the CBC under high light. 2) Metabolite regulations, especially allosteric regulations of major regulatory enzymes, ensure the rapid up-regulation of fluxes to ensure sufficient amount of triose phosphate is available for end product synthesis and concurrently avoid phosphate limitation. 3) A balanced activities of enzymes in the CBC help maintain balanced flux through CBC; some innate product feedback mechanisms, in particular the ADP feedback regulation of GAPDH and F6P feedback regulation of FBPase, exist in CBC to achieve such a balanced enzyme activities and hence flux distribution in the CBC for greater photosynthetic efficiency. Transcriptional regulation and natural variations of enzymes controlling CBC metabolite homeostasis should be further explored to maximize the potential of engineering CBC for greater efficiency.

The Gynandropsis gynandra genome provides insights into whole-genome duplications and the evolution of C4 photosynthesis in Cleomaceae.

Gynandropsis gynandra (Cleomaceae) is a cosmopolitan leafy vegetable and medicinal plant, which has also been used as a model to study C4 photosynthesis due to its evolutionary proximity to C3 Arabidopsis (Arabidopsis thaliana). Here, we present the genome sequence of G. gynandra, anchored onto 17 main pseudomolecules with a total length of 740 Mb, an N50 of 42 Mb and 30,933 well-supported gene models. The G. gynandra genome and previously released genomes of C3 relatives in the Cleomaceae and Brassicaceae make an excellent model for studying the role of genome evolution in the transition from C3 to C4 photosynthesis. Our analyses revealed that G. gynandra and its C3 relative Tarenaya hassleriana shared a whole-genome duplication event (Gg-α), then an addition of a third genome (Th-α, +1×) took place in T. hassleriana but not in G. gynandra. Analysis of syntenic copy number of C4 photosynthesis-related gene families indicates that G. gynandra generally retained more duplicated copies of these genes than C3T. hassleriana, and also that the G. gynandra C4 genes might have been under positive selection pressure. Both whole-genome and single-gene duplication were found to contribute to the expansion of the aforementioned gene families in G. gynandra. Collectively, this study enhances our understanding of the polyploidy history, gene duplication and retention, as well as their impact on the evolution of C4 photosynthesis in Cleomaceae.

Increased α-ketoglutarate links the C3-C4 intermediate state to C4 photosynthesis in the genus Flaveria.

As a complex trait, C4 photosynthesis has multiple independent origins in evolution. Phylogenetic evidence and theoretical analysis suggest that C2 photosynthesis, which is driven by glycine decarboxylation in the bundle sheath cell, may function as a bridge from C3 to C4 photosynthesis. However, the exact molecular mechanism underlying the transition between C2 photosynthesis to C4 photosynthesis remains elusive. Here, we provide evidence suggesting a role of higher α-ketoglutarate (AKG) concentration during this transition. Metabolomic data of 12 Flaveria species, including multiple photosynthetic types, show that AKG concentration initially increased in the C3-C4 intermediate with a further increase in C4 species. Petiole feeding of AKG increases the concentrations of C4-related metabolites in C3-C4 and C4 species but not the activity of C4-related enzymes. Sequence analysis shows that glutamate synthase (Fd-GOGAT), which catalyzes the generation of glutamate using AKG, was under strong positive selection during the evolution of C4 photosynthesis. Simulations with a constraint-based model for C3-C4 intermediate further show that decreasing the activity of Fd-GOGAT facilitated the transition from a C2-dominant to a C4-dominant CO2 concentrating mechanism. All these results provide insight into the mechanistic switch from C3-C4 intermediate to C4 photosynthesis.

GLK transcription factors accompany ELONGATED HYPOCOTYL5 to orchestrate light-induced seedling development in Arabidopsis.

Light-induced de-etiolation is an important aspect of seedling photomorphogenesis. GOLDEN2 LIKE (GLK) transcriptional regulators are involved in chloroplast development, but to what extent they participate in photomorphogenesis is not clear. Here, we show that ELONGATED HYPOCOTYL5 (HY5) binds to GLK promoters to activate their expression, and also interacts with GLK proteins in Arabidopsis (Arabidopsis thaliana). The chlorophyll content in the de-etiolating Arabidopsis seedlings of the hy5 glk2 double mutants was lower than that in the hy5 single mutant. GLKs inhibited hypocotyl elongation, and the phenotype could superimpose on the hy5 phenotype. Correspondingly, GLK2 regulated the expression of photosynthesis and cell elongation genes partially independent of HY5. Before exposure to light, DE-ETIOLATED 1 (DET1) affected accumulation of GLK proteins. The enhanced etioplast development and photosystem gene expression observed in the det1 mutant were attenuated in the det1 glk2 double mutant. Our study reveals that GLKs act downstream of HY5, or additive to HY5, and are likely quantitatively adjusted by DET1, to orchestrate multiple developmental traits during the light-induced skotomorphogenesis-to-photomorphogenesis transition in Arabidopsis.

Regulatory NADH dehydrogenase-like complex optimizes C4 photosynthetic carbon flow and cellular redox in maize.

C4 plants typically operate a CO2 concentration mechanism from mesophyll (M) cells into bundle sheath (BS) cells. NADH dehydrogenase-like (NDH) complex is enriched in the BS cells of many NADP-malic enzyme (ME) type C4 plants and is more abundant in C4 than in C3 plants, but to what extent it is involved in the CO2 concentration mechanism remains to be experimentally investigated. We created maize and rice mutants deficient in NDH function and then used a combination of transcriptomic, proteomic, and metabolomic approaches for comparative analysis. Considerable decreases in growth, photosynthetic activities, and levels of key photosynthetic proteins were observed in maize but not rice mutants. However, transcript abundance for many cyclic electron transport (CET) and Calvin-Benson cycle components, as well as BS-specific C4 enzymes, was increased in maize mutants. Metabolite analysis of the maize ndh mutants revealed an increased NADPH : NADP ratio, as well as malate, ribulose 1,5-bisphosphate (RuBP), fructose 1,6-bisphosphate (FBP), and photorespiration intermediates. We suggest that by optimizing NADPH and malate levels and adjusting NADP-ME activity, NDH functions to balance metabolic and redox states in the BS cells of maize (in addition to ATP supply), coordinating photosynthetic transcript abundance and protein content, thus directly regulating the carbon flow in the two-celled C4 system of maize.

The Evolution of C4 Photosynthesis in Flaveria (Asteraceae): Insights from the Flaveria linearis Complex.

Flaveria is a leading model for C4 plant evolution due to the presence of a dozen C3-C4 intermediate species, many of which are associated with a phylogenetic complex centered around Flaveria linearis. To investigate C4 evolution in Flaveria, we updated the Flaveria phylogeny and evaluated gas exchange, starch δ13C, and activity of C4 cycle enzymes in 19 Flaveria species and 28 populations within the F. linearis complex. A principal component analysis identified six functional clusters: (1) C3, (2) sub-C2, (3) full C2, (4) enriched C2, (5) sub-C4, and (6) fully C4 species. The sub-C2 species lacked a functional C4 cycle, while a gradient was present in the C2 clusters from little to modest C4 cycle activity as indicated by δ13C and enzyme activities. Three Yucatan populations of F. linearis had photosynthetic CO2 compensation points equivalent to C4 plants but showed little evidence for an enhanced C4 cycle, indicating they have an optimized C2 pathway that recaptures all photorespired CO2 in the bundle sheath (BS) tissue. All C2 species had enhanced aspartate aminotransferase activity relative to C3 species and most had enhanced alanine aminotransferase activity. These aminotransferases form aspartate and alanine from glutamate and in doing so could help return photorespiratory nitrogen (N) from BS to mesophyll cells, preventing glutamate feedback onto photorespiratory N assimilation. Their use requires upregulation of parts of the C4 metabolic cycle to generate carbon skeletons to sustain N return to the mesophyll, and thus could facilitate the evolution of the full C4 photosynthetic pathway.

Rethinking the potential productivity of crassulacean acid metabolism by integrating metabolic dynamics with shoot architecture, using the example of Agave tequilana.

Terrestrial CAM plants typically occur in hot semiarid regions, yet can show high crop productivity under favorable conditions. To achieve a more mechanistic understanding of CAM plant productivity, a biochemical model of diel metabolism was developed and integrated with 3-D shoot morphology to predict the energetics of light interception and photosynthetic carbon assimilation. Using Agave tequilana as an example, this biochemical model faithfully simulated the four diel phases of CO2 and metabolite dynamics during the CAM rhythm. After capturing the 3-D form over an 8-yr production cycle, a ray-tracing method allowed the prediction of the light microclimate across all photosynthetic surfaces. Integration with the biochemical model thereby enabled the simulation of plant and stand carbon uptake over daily and annual courses. The theoretical maximum energy conversion efficiency of Agave spp. is calculated at 0.045-0.049, up to 7% higher than for C3 photosynthesis. Actual light interception, and biochemical and anatomical limitations, reduced this to 0.0069, or 15.6 Mg ha-1 yr-1 dry mass annualized over an 8-yr cropping cycle, consistent with observation. This is comparable to the productivity of many C3 crops, demonstrating the potential of CAM plants in climates where little else may be grown while indicating strategies that could raise their productivity.

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.

On the rate of phytoplankton respiration in the light.

The rate of algal and cyanobacterial respiration in the light is an important ecophysiological term that remains to be completely characterized and quantified. To address this issue, we exploited process-specific decarboxylation rates from flux balance analysis and isotopically nonstationary metabolic flux analysis. Our study, based on published data, suggested that decarboxylation is about 22% of net CO2 assimilation when the tricarboxylic acid cycle is completely open (characterized by the commitment of alpha ketoglutarate to amino acid synthesis and very low rates of succinate formation). This estimate was supported by calculating the decarboxylation rates required to synthesize the major components of biomass (proteins, lipids, and carbohydrates) at their typical abundance. Of the 22 CO2 molecules produced by decarboxylation (normalized to net assimilation = 100), approximately 13 were from pyruvate and 3 were from isocitrate. The remaining six units of decarboxylation were in the amino acid synthesis pathways outside the tricarboxylic acid cycle. A small additional flux came from photorespiration, decarboxylations of six phosphogluconate in the oxidative pentose phosphate pathway, and decarboxylations in the syntheses of lower-abundance compounds, including pigments and ribonucleic acids. This general approach accounted for the high decarboxylation rates in algae and cyanobacteria compared to terrestrial plants. It prompts a simple speculation for the origin of the Kok effect and helps constrain the photoautotrophic respiration rate, in the light, in the euphotic zone of the ocean and lakes.

The evolution of stomatal traits along the trajectory toward C4 photosynthesis.

C4 photosynthesis optimizes plant carbon and water relations, allowing high photosynthetic rates with low stomatal conductance. Stomata have long been considered a part of the C4 syndrome. However, it remains unclear how stomatal traits evolved along the path from C3 to C4. Here, we examined stomata in the Flaveria genus, a model used for C4 evolutionary study. Comparative, transgenic, and semi-in vitro experiments were performed to study the molecular basis that underlies the changes of stomatal traits in C4 evolution. The evolution from C3 to C4 species is accompanied by a gradual rather than an abrupt change in stomatal traits. The initial change appears near the Type I intermediate stage. Co-evolution of the photosynthetic pathway and stomatal traits is supported. On the road to C4, stomata tend to be fewer in number but larger in size and stomatal density dominates changes in anatomical maximum stomatal conductance (gsmax). Reduction of FSTOMAGEN expression underlies decreased gsmax in Flaveria and likely occurs in other C4 lineages. Decreased gsmax contributes to the increase in intrinsic water-use efficiency in C4 evolution. This work highlights the stomatal traits in the current C4 evolutionary model. Our study provides insights into the pattern, mechanism, and role of stomatal evolution along the road toward C4.

Two major metabolic factors for an efficient NADP-malic enzyme type C4 photosynthesis.

Compared to the large number of studies focused on the factors controlling C3 photosynthesis efficiency, there are relatively fewer studies of the factors controlling photosynthetic efficiency in C4 leaves. Here, we used a dynamic systems model of C4 photosynthesis based on maize (Zea mays) to identify features associated with high photosynthetic efficiency in NADP-malic enzyme (NADP-ME) type C4 photosynthesis. We found that two additional factors related to coordination between C4 shuttle metabolism and C3 metabolism are required for efficient C4 photosynthesis: (1) accumulating a high concentration of phosphoenolpyruvate through maintaining a large PGA concentration in the mesophyll cell chloroplast and (2) maintaining a suitable oxidized status in bundle sheath cell chloroplasts. These identified mechanisms are in line with the current cellular location of enzymes/proteins involved in the starch synthesis, the Calvin-Benson cycle and photosystem II of NADP-ME type C4 photosynthesis. These findings suggested potential strategies for improving C4 photosynthesis and engineering C4 rice.

The energy cost of repairing photoinactivated photosystem II: an experimental determination in cotton leaf discs.

Photosystem II (PSII), which splits water molecules at minimal excess photochemical potential, is inevitably photoinactivated during photosynthesis, resulting in compromised photosynthetic efficiency unless it is repaired. The energy cost of PSII repair is currently uncertain, despite attempts to calculate it. We experimentally determined the energy cost of repairing each photoinactivated PSII in cotton (Gossypium hirsutum) leaves, which are capable of repairing PSII in darkness. As an upper limit, 24 000 adenosine triphosphate (ATP) molecules (including any guanosine triphosphate synthesized at the expense of ATP) were required to repair one entire PSII complex. Further, over a 7-h illumination period at 526-1953 µmol photons m-2 s-1 , the ATP requirement for PSII repair was on average up to 4.6% of the ATP required for the gross carbon assimilation. Each of these two measures of ATP requirement for PSII repair is two- to three-fold greater than the respective reported calculated value. Possible additional energy sinks in the PSII repair cycle are discussed.

Potential metabolic mechanisms for inhibited chloroplast nitrogen assimilation under high CO2.

Improving photosynthesis is considered a major and feasible option to dramatically increase crop yield potential. Increased atmospheric CO2 concentration often stimulates both photosynthesis and crop yield, but decreases protein content in the main C3 cereal crops. This decreased protein content in crops constrains the benefits of elevated CO2 on crop yield and affects their nutritional value for humans. To support studies of photosynthetic nitrogen assimilation and its complex interaction with photosynthetic carbon metabolism for crop improvement, we developed a dynamic systems model of plant primary metabolism, which includes the Calvin-Benson cycle, the photorespiration pathway, starch synthesis, glycolysis-gluconeogenesis, the tricarboxylic acid cycle, and chloroplastic nitrogen assimilation. This model successfully captures responses of net photosynthetic CO2 uptake rate (A), respiration rate, and nitrogen assimilation rate to different irradiance and CO2 levels. We then used this model to predict inhibition of nitrogen assimilation under elevated CO2. The potential mechanisms underlying inhibited nitrogen assimilation under elevated CO2 were further explored with this model. Simulations suggest that enhancing the supply of α-ketoglutarate is a potential strategy to maintain high rates of nitrogen assimilation under elevated CO2. This model can be used as a heuristic tool to support research on interactions between photosynthesis, respiration, and nitrogen assimilation. It also provides a basic framework to support the design and engineering of C3 plant primary metabolism for enhanced photosynthetic efficiency and nitrogen assimilation in the coming high-CO2 world.

Green giant-a tiny chloroplast genome with mighty power to produce high-value proteins: history and phylogeny.

Free-living cyanobacteria were entrapped by eukaryotic cells ~2 billion years ago, ultimately giving rise to chloroplasts. After a century of debate, the presence of chloroplast DNA was demonstrated in the 1960s. The first chloroplast genomes were sequenced in the 1980s, followed by ~100 vegetable, fruit, cereal, beverage, oil and starch/sugar crop chloroplast genomes in the past three decades. Foreign genes were expressed in isolated chloroplasts or intact plant cells in the late 1980s and stably integrated into chloroplast genomes, with typically maternal inheritance shown in the 1990s. Since then, chloroplast genomes conferred the highest reported levels of tolerance or resistance to biotic or abiotic stress. Although launching products with agronomic traits in important crops using this concept has been elusive, commercial products developed include enzymes used in everyday life from processing fruit juice, to enhancing water absorption of cotton fibre or removal of stains as laundry detergents and in dye removal in the textile industry. Plastid genome sequences have revealed the framework of green plant phylogeny as well as the intricate history of plastid genome transfer events to other eukaryotes. Discordant historical signals among plastid genes suggest possible variable constraints across the plastome and further understanding and mitigation of these constraints may yield new opportunities for bioengineering. In this review, we trace the evolutionary history of chloroplasts, status of autonomy and recent advances in products developed for everyday use or those advanced to the clinic, including treatment of COVID-19 patients and SARS-CoV-2 vaccine.

Canopy occupation volume as an indicator of canopy photosynthetic capacity.

Leaf angle and leaf area index together influence canopy light interception and canopy photosynthesis. However, so far, there is no effective method to identify the optimal combination of these two parameters for canopy photosynthesis. In this study, first a robust high-throughput method for accurate segmentation of maize organs based on 3D point clouds data was developed, then the segmented plant organs were used to generate new 3D point clouds for the canopy of altered architectures. With this, we simulated the synergistic effect of leaf area and leaf angle on canopy photosynthesis. The results show that, compared to the traditional parameters describing the canopy photosynthesis including leaf area index, facet angle and canopy coverage, a new parameter - the canopy occupation volume (COV) - can better explain the variations of canopy photosynthetic capacity. Specifically, COV can explain > 79% variations of canopy photosynthesis generated by changing leaf angle and > 84% variations of canopy photosynthesis generated by changing leaf area. As COV can be calculated in a high-throughput manner based on the canopy point clouds, it can be used to evaluate canopy architecture in breeding and agronomic research.

Honoring Bacon Ke at 100: a legend among the many luminaries and a highly collaborative scientist in photosynthesis research.

Bacon Ke, who did pioneering research on the primary photochemistry of photosynthesis, was born in China on July 26, 1920, and currently, he is living in a senior home in San Francisco, California, and is a centenarian. To us, this is a very happy and unique occasion to honor him. After providing a brief account of his life, and a glimpse of his research in photosynthesis, we present here "messages" for Bacon Ke@ 100 from: Robert Alfano (USA), Charles Arntzen (USA), Sandor Demeter (Hungary), Richard A. Dilley (USA), John Golbeck (USA), Isamu Ikegami (Japan), Ting-Yun Kuang (China), Richard Malkin (USA), Hualing Mi (China), Teruo Ogawa (Japan), Yasusi Yamamoto (Japan), and Xin-Guang Zhu (China).

Natural variation in the fast phase of chlorophyll a fluorescence induction curve (OJIP) in a global rice minicore panel.

Photosynthesis can be probed through Chlorophyll a fluorescence induction (FI), which provides detailed insight into the electron transfer process in Photosystem II, and beyond. Here, we have systematically studied the natural variation of the fast phase of the FI, i.e. the OJIP phase, in rice. The OJIP phase of the Chl a fluorescence induction curve is referred to as "fast transient" lasting for less than a second; it is obtained after a dark-adapted sample is exposed to saturating light. In the OJIP curve, "O" stands for "origin" (minimal fluorescence), "P" for "peak" (maximum fluorescence), and J and I for inflection points between the O and P levels. Further, Fo is the fluorescence intensity at the "O" level, whereas Fm is the intensity at the P level, and Fv (= Fm - Fo) is the variable fluorescence. We surveyed a set of quantitative parameters derived from the FI curves of 199 rice accessions, grown under both field condition (FC) and growth room condition (GC). Our results show a significant variation between Japonica (JAP) and Indica (IND) subgroups, under both the growth conditions, in almost all the parameters derived from the OJIP curves. The ratio of the variable to the maximum (Fv/Fm) and of the variable to the minimum (Fv/Fo) fluorescence, the performance index (PIabs), as well as the amplitude of the I-P phase (AI-P) show higher values in JAP compared to that in the IND subpopulation. In contrast, the amplitude of the O-J phase (AO-J) and the normalized area above the OJIP curve (Sm) show an opposite trend. The performed genetic analysis shows that plants grown under GC appear much more affected by environmental factors than those grown in the field. We further conducted a genome-wide association study (GWAS) using 11 parameters derived from plants grown in the field. In total, 596 non-unique significant loci based on these parameters were identified by GWAS. Several photosynthesis-related proteins were identified to be associated with different OJIP parameters. We found that traits with high correlation are usually associated with similar genomic regions. Specifically, the thermal phase of FI, which includes the amplitudes of the J-I and I-P subphases (AJ-I and AI-P) of the OJIP curve, is, in turn, associated with certain common genomic regions. Our study is the first one dealing with the natural variations in rice, with the aim to characterize potential candidate genes controlling the magnitude and half-time of each of the phases in the OJIP FI curve.