Plant lineages produce distinct alkaloids, indicating the presence of evolutionary mechanisms that prevent the simultaneous metabolism of multiple alkaloid types within the same species. In the Solanaceae family, species produce either tropane alkaloids (TAs) or steroidal glycoalkaloids (SGAs). To investigate the genetic causes underlying this mutually exclusive distribution, we integrated genomic, transcriptomic, and metabolomic data across tens of species representing most Solanaceae clades, focusing on three potential mechanisms: (1) loss of biosynthetic genes, (2) genomic clustering, and (3) regulatory changes. Ancestral trait reconstruction suggests that both pathways were likely present in early Solanaceae lineages but were differentially lost across clades, resulting in transitions between SGA and TA biosynthesis. Notably, SGA and TA genes exhibit contrasting patterns of genomic distribution: SGA genes are organized in large, dynamic biosynthetic clusters, while TA genes are dispersed across the genome. More importantly, SGA genes are ancient and phylogenetically widespread, but are not expressed in clades that produce TAs, implying that a regulatory switch was responsible for the simultaneous silencing of SGA genes in these clades. Finally, we observed the loss and genetic erosion of late-acting SGA and TA genes in clades where they are not expressed. Overall, our study suggests that genomic clustering, regulatory changes, and gene duplication/loss underlie the mutually exclusive distribution of alkaloids in the Solanaceae.

C4 plants have traditionally been classified into NADP-malic enzyme (NADP-ME), NAD-malic enzyme (NAD-ME), and phosphoenolpyruvate carboxykinase (PEPCK) subtypes based on the predominant C4 acid decarboxylating enzyme. To investigate the relative contributions of malate and aspartate to C4 pathway fluxes in each subtype, we performed 13CO2 pulse-chase labelling experiments on four C4 grass species: Zea mays and Setaria viridis (NADP-ME), Panicum miliaceum (NAD-ME), and Megathyrsus maximus (PEPCK). Only a proportion (8-50%) of the total malate pool in the leaves is photosynthetically active, whereas essentially all of the aspartate pool is photosynthetically active. Estimates of metabolic fluxes indicate that approximately two-thirds of the C4 pathway flux is via malate in Z. mays and the remaining third via aspartate, while in S. viridis 50% of the flux is via malate and 50% via aspartate. In P. miliaceum and M. maximus, 91% and 85% of the flux is via aspartate and the remaining 9% and 15% via malate, respectively. The results demonstrate the feasibility of using non-radioactive 13CO2 in pulse-chase labelling experiments to study C4 photosynthesis and to detect C4 pathway fluxes in C3 plants engineered to perform C4 photosynthesis.

Lysine metabolism is a central determinant of plant growth, coupling amino acid biosynthesis to mitochondrial electron transport and the tricarboxylic acid (TCA) cycle to sustain cellular energy homeostasis. However, whether perturbation of lysine biosynthesis reshapes developmental transitions through epigenetic regulatory pathways remains unknown. Here, we show that reduced activity of L,L -diaminopimelate aminotransferase (DAPAT), a key enzyme in lysine biosynthesis, markedly reprograms growth and developmental timing in Arabidopsis thaliana . The lysine biosynthesis-deficient mutant dapat exhibits severe development constraints, displaying a pronounced flowering delay under long-day conditions accompanied by reduced shoot branching, fewer siliques, lower seed number per silique, decreased seed biomass, and reduced leaf production, a comprehensive developmental phenotype that underscores the critical role of lysine metabolism in orchestrating plant growth. Strikingly, neutral-day conditions partially rescue the flowering delay in dapat plants, revealing an unexpected environmental plasticity that positions lysine biosynthesis at the intersection of metabolic regulation, epigenetic remodeling, and developmental timing. Gene expression analysis of key flowering-time regulators that control the photoperiodic and vernalization pathways in Arabidopsis revealed upregulation of CONSTANS ( CO ) and FLOWERING LOCUS C ( FLC ), downregulation of FRIGIDA ( FRI ), and unaltered FLOWERING LOCUS T ( FT ), suggesting that lysine deficiency modulates flowering through a CO / FLC -dependent regulatory framework. Using whole-genome bisulfite sequencing (WGBS) we identified 957 genes with altered methylation patterns across the plant's five chromosomes. These changes occurred in three different DNA sequence contexts: 749 genes showed differences in CpG sites, while 104 genes each showed changes in CHG and CHH contexts. These epigenetic changes coincide with altered expression of genes involved in S -adenosyl-methionine metabolism and chromatin regulation, including upregulation of S -adenosyl-methionine synthase 2 ( MAT2 ), S -adenosyl-methionine synthase 4 ( MAT4 ), and histone methyltransferase SU(VAR)3–9 HOMOLOG 4 ( SUVH4 ), whereas SUVH5 is downregulated and SUVH6 remains unchanged. These findings uncover a previously unrecognized role for lysine biosynthesis via DAPAT enzyme in coordinating plant growth, flowering time, and reproductive output through metabolic-epigenetic regulation.

Seasonal temperature increases in autumn and winter can significantly disrupt frost tolerance in plants. This inducible trait is acquired during cold acclimation, reduced or lost during de-acclimation, and potentially restored during re-acclimation. The functionality of cellular membranes, closely associated with lipid composition, plays a crucial adaptive role in enabling plants to withstand low-temperature stress. However, studies focusing on effects of de-acclimation and re-acclimation on frost tolerance, particularly in forage grasses, remain limited. Here, we assessed frost tolerance in high (HFT) and low frost tolerant (LFT) genotypes of Festuca arundinacea following cold acclimation, de-acclimation, and re-acclimation, based on the assessment of their cellular membranes integrity as reflected by the parameter describing the temperature causing 50% level of electrolyte leakage from cells. Analyses of membrane integrity and lipid composition during cold acclimation revealed that the HFT genotype maintained greater membrane stability through extensive remodeling of glycerolipids, including increased lipid unsaturation, particularly in phosphatidylethanolamine (PE), preserved a balanced monogalactosyldiacylglycerol/digalactosyldiacylglycerol (MGDG/DGDG) ratio, and enhanced triacylglycerol (TAG) accumulation, compared to the LFT genotype. De-acclimation reversed rearrangements in structural lipids and reduced frost tolerance, whereas re-acclimation induced strong genotype-dependent responses, especially in phosphoglycerolipids profiles. Analysis of mRNA accumulation profiles indicated key enzymes potentially involved in lipid remodeling. Major transcriptional changes were observed in phytyl ester synthase 1 (PES1), digalactosyldiacylglycerol synthase (DGD1), and lipoxygenase (LOX) family transcripts, indicating their importance during cold acclimation, de-acclimation, and re-acclimation. Collectively, these findings demonstrate, for the first time, that genotype-dependent lipid remodeling underlies the capacity of F. arundinacea to maintain and recover frost tolerance. We further suggest that stress-induced TAG accumulation may function as a protective metabolic buffer, sequestering excess free fatty acids, limiting lipid oxidation, and supporting membrane stability under low-temperature stress.

Gibberellins (GAs) play a crucial role in modulating developmental processes throughout the plant life cycle. They are particularly significant during the transition and maintenance of the reproductive meristem, as well as in facilitating the development of floral organs. Additionally, GAs regulate the early stages of fruit development, in coordination with auxin and cytokinin, likely due to their involvement in both division and cell expansion. However, it remains unclear whether fluctuations in endogenous GA levels influence fruit development and metabolism during ripening. To address this, we investigated tomato mutant plants deficient in GAs biosynthesis (gib3, moderately deficient; gib2, intermediate deficiency and gib1, extremely deficient in GAs). Notably, gib2 and gib1 mutants were characterized by a complete interruption of their reproductive development at the floral bud level. Although gib3 plants displayed a slight delay in fruit development, at the end of fruit ripening both wild type (WT) and gib3 fruits were highly similar. Only minor differences were found between WT and gib3 mutant plants in terms of floral development and total fruit yield. Our findings revealed that reduced GA levels in gib3 mutant did not result in morphological modifications in fruits, and relatively few metabolic changes were observed between genotypes during fruit ripening. Typical metabolic changes during ripening, including increments in amino acids and soluble sugars along with decreases in starch, were observed. Collectively, our study demonstrate that GAs play a crucial role in transitioning plants from the vegetative to reproductive stage and in initiating fruit set. Tomato plants with reduced endogenous content of gibberellins show interruption or delay of the reproductive stage.

Natural rubber (NR), valued for its elasticity and impact resistance, is essential for numerous industrial and medical applications, with global demand continuously rising. While approximately 2500 plant species from more than 40 families can produce rubber, the majority is sourced from Hevea brasiliensis grown in tropical regions. Alternative rubber-producing plants, such as Parthenium argentatum and Taraxacum kok-saghyz, offer enhanced environmental adaptability and species diversity, making them promising candidates for rubber production. Recent genome sequencing has shed light on rubber biosynthesis pathways, although the mechanisms involved in producing different forms of polyisoprene across species remain unclear. We explore the evolution of rubber biosynthesis and discuss synthetic biological strategies for enhancing NR-production in subtropical plants and a broader range of plant materials (e.g., Manilkara zapota).

Color and flavor are key quality traits in fruits. Using a newly constructed peach pangenome, Chen et al. demonstrated that the PbBL gene, a known regulator of peach fruit color, also contributes to malate accumulation. This finding, along with previous studies, unveils a transcriptional mechanism that co-regulates multiple traits in peaches.

Coffee (Coffea spp.) is one of the most economically important crop species and serves as a rich source of bioactive specialized (secondary) metabolites with various health-promoting properties. Advances in analytical food chemistry and phytochemistry have elucidated an extensive and structurally diverse specialized metabolism in coffee beans, much of which contributes to both organoleptic attributes and adaptive physiological responses in coffee plants. Recent developments in omics-driven methodologies have provided new insights into both coffee metabolism and breeding strategies, particularly those aimed at enhancing both quality traits and environmental resilience. Comparative genomic analyses across Coffea species and cultivars have facilitated the detection of metabolic polymorphisms, enabling inter- and intra-species assessments of biosynthetic pathway variation and the refinement of biosynthetic frameworks for further functional genomics approaches. Such approaches yield critical information regarding the genetic and biochemical determinants underlying specialized metabolite accumulations, which can be directly applied for targeted metabolic engineering and crop improvement. Moreover, cross-species comparative omics and multi-omics integrative analyses, particularly in relation to phylogenetically relevant taxa such as Solanaceae species, exemplified by the model crop tomato (Solanum lycopersicum), provide valuable translational insights into conserved and divergent metabolic architectures.

β-Alanine, an abundant non-proteinogenic amino acid, acts as a precursor for coenzyme A and plays a role in various stress responses. However, a comprehensive understanding of its metabolism in plants remains incomplete. Previous metabolic genome-wide association studies (mGWAS) identified ALANINE:GLYOXYLATE AMINOTRANSFERASE2 (AGT2, AT4G39660) linked to β-alanine levels in Arabidopsis under normal conditions. In this study, we aimed to deepen our insights into β-alanine regulation by conducting mGWAS under two contrasting environmental conditions: control (12 h photoperiod, 21°C, 150 μmol m-2 sec-1) and stress (harvested after 1820 min at 32°C and darkness). We identified two highly significant quantitative trait loci (QTL) for β-alanine, including the AGT2 locus associated in both environments and ALDEHYDE DEHYDROGENASE6B2 (ALDH6B2, AT2G14170) associated only under stress conditions. A coexpression-correlation network revealed that the regulatory pathway involving β-alanine levels, AGT2, and ALDH6B2 connects the branched chained amino acid (BCAA) degradation through the propionate pathway. Metabolic profiles of AGT2 overexpression (OE) and knock-out (KO) lines (agt2) across various organs and developmental stages established the critical role of AGT2 in β-alanine metabolism. This work underscores the importance of β-alanine homeostasis for proper growth and development in Arabidopsis.

Goji berries, an important medicinal and edible plant, possess substantial medicinal and economic potential. While research has extensively investigated the polar metabolites of goji berries, their lipid metabolism has received comparatively less attention. Here, we performed comprehensive lipid profiling of 78 goji berry samples using UPLC-MS. We identified a total of 180 lipid compounds, classified into five classes: glycerolipids, glycerophospholipids, glycoglycerolipids, sphingolipids, and pigments. The data reveal the natural variation and accumulation patterns of lipid compounds in cultivated and wild goji berries, and assess lipid distribution characteristics across different harvesting periods and maturation stages. Heatmap clustering and PCA analysis indicated that all samples displayed a notable spatial aggregation. Furthermore, the OPLS-DA analysis demonstrated key markers for distinguishing goji berries from different harvesting periods. These findings offer valuable insights into the lipid metabolism in Lycium and establish a basis for assessing the quality of this genus.

The Anaphase-Promoting Complex/Cyclosome (APC/C) is an E3 ubiquitin ligase that plays a crucial role in ubiquitin-dependent proteolysis of key cell cycle regulators, which is completed by the 26S proteasome. Previously, SAMBA, a plant-specific regulator of the APC/C, was identified in Arabidopsis as a critical factor controlling organ size through the regulation of cell proliferation. Here, by assessing its role in the crop tomato (Solanum lycopersicum), we confirm that SAMBA is a conserved APC/C regulator in plants and shows additional roles, including the modulation of fruit shape and changes in sugar metabolism. Two slsamba genome-edited lines were produced and characterized, and showed delayed growth, reduced plant size, and altered fruit morphology, which were linked to changes in cell division and expansion. Notably, untargeted metabolomics revealed altered flavonoid profiles, along with elevated Brix values in the fruits, indicating a sweeter taste. Accordingly, transcriptomics uncovered a change in temporal gene expression gradients during early fruit development, correlating with the alterations in sugar metabolism and revealing changes in cell wall biosynthesis genes. This study provides the first evidence of SAMBA's role in regulating fruit development, metabolic content, and ultimately, quality. These important findings offer potential applications for improving the nutritional quality and overall performance of tomatoes.

The growth-differentiation trade-off limits productivity in secondary metabolite-rich crops. Here, using Stevia rebaudiana as a model plant, we find that the N source strongly regulates the trade-off between biomass and steviol glycoside (SG) accumulation, with a nitrate [NO3-]-to-ammonium [NH4+] ratio of 75:25 yielding optimal SG production. Metabolomics analyses attributed the benefits of a high proportion of NO3- supply to redirected carbon flux into phenolic and terpenoid pathways. While overall rhizomicrobial diversity remained unaffected by N forms, the composition of the bacterial, rather than fungal, community changed significantly. NO3- supply favored bacterial taxa from the Burkholderiales, Hyphalales, and Cytophagales orders while diminishing those belonging to Vicinamibacterales. Notably, NH4+-associated bacteria were linked to amino acid synthesis, while NO3--enriched taxa, including Sphingomonadaceae and Thermoanaerobaculaceae, correlated with secondary metabolite accumulation. Our study highlighted the tight link between N-form-regulated trade-offs and rhizobacterial community assembly, providing insights into plant-microbe interactions and strategies for optimizing crop yield and bioactive compound accumulation.

In recent years, several studies investigating multifactorial stresses have emerged. This shift has been driven by the recognition that one of the primary reasons for the inconsistency between laboratory-based results and field observations of plant responses is that, in natural environments, plants are routinely exposed to a combination of biotic and/or abiotic stresses, which they encounter either simultaneously or sequentially. Within this review, we address current advances in multifactorial studies focusing on metabol(om)ic changes in model as well as cereal crop species. The common consensus is that currently, studies on phenotypic and transcriptomics analysis are prevailing, while metabolic studies are scarce. Despite the need for further studies to validate the findings in this review, two clear biological messages emerge. First, and perhaps unsurprisingly, proline stands out as a universal stress metabolite, closely followed by branched-chain amino acids. Interestingly, while multifactorial stress responses are often considered non-additive and unpredictable, our findings reveal that many metabolic changes are both. Expanding the scope of studies to include more species and a wider range of stresses at the metabolic level will be essential for uncovering additional metabolic reprogramming in response to multifactorial stress. This will provide invaluable insights for developing breeding strategies aimed at future-proofing crops.

Purine-related metabolites are central to primary metabolic pathways in plants and serve as precursors for purine alkaloid biosynthesis in caffeinated species such as tea plants (Camellia sinensis). In this study, metabolite profiling of two tissues (young and mature leaves) was performed across 183 genetically diverse tea accessions, identifying and quantifying 10 purine alkaloid-related metabolites. Metabolite genome-wide association studies revealed 17 significant loci associated with these metabolites, including both known loci such as caffeine synthase and 16 novel loci (P < 1.05 × 10-5). Through functional annotation and in vitro enzymatic assay, we characterized 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase (CsMTAN) as the causal gene underlying natural variation in adenosine and adenine content. CsMTAN can catalyze the degradation of both 5'-methylthioadenosine and S-adenosylhomocysteine to release adenine. The T → A nucleotide substitution at SNP55151898, which leads to a phenylalanine → tyrosine substitution at residue 179 (F179Y), resulted in a significant alteration of enzyme activity in vitro, as evidenced by an approximately 50% reduction in adenine abundance (P < 0.05). Transient overexpression of CsMTAN-A and CsMTAN-T in Nicotiana benthamiana both significantly increased adenine content and dramatically decreased adenosine content, providing direct evidence for the functional involvement of CsMTAN in plant purine metabolism. CsMTAN-T overexpression resulted in significantly lower adenosine level than CsMTAN-A (P < 0.05). Phylogenetic analysis across 115 species and protein structural modeling revealed a distinct evolutionary divergence between plant MTAN evolution and species phylogeny, strongly suggesting the occurrence of horizontal gene transfer events in the evolutionary history of plant MTANs. This study thus furthered our understanding of the genetics and molecular mechanisms regulating purine metabolism and purine alkaloid biosynthesis in tea plants and provided novel targets for molecular breeding and synthetic biology applications.

Plant metabolism is remarkably flexible. Rapid changes in the rate and mode of primary metabolism are essential to meet the demands of plants under changeable conditions. While it is evident that photosynthetic metabolism must be regulated to match changes in illumination, the principles that govern the regulation of respiratory metabolism have remained less obvious, even though plant respiratory rates can vary profoundly. An extreme transition in respiratory metabolism occurs when a thermogenic plant tissue enters the phase of heat generation. Here, we review our current understanding of what is required to re-model plant metabolism toward thermogenesis and highlight recent advances. We propose plant thermogenesis as a model to uncover novel mechanisms that control respiration rate. Those mechanisms may aid engineering carbon use efficiency and improve stress resilience in plants and beyond.

In nature, environmental conditions strongly fluctuate, frequently subjecting plants to periods of immediate photo-oxidative stress. The small molecule ascorbate allows plants to cope with such stress conditions. Ascorbate scavenges reactive oxygen species and enables the rapid and full induction of photoprotective non-photochemical quenching (NPQ). NPQ is dependent on zeaxanthin, which requires ascorbate as the electron donor during its synthesis by the violaxanthin de-epoxidase. The VTC2 gene encodes one of two isoforms of GDP-l-galactose phosphorylase, the rate-controlling enzyme of ascorbate biosynthesis. In the current study, by including a newly identified vtc2 allele, we found that loss of VTC2 depleted ascorbate mainly from the mature leaves and thereby limited NPQ specifically in this tissue. Growth in fluctuating light and controlled climate suppressed the slow NPQ induction phenotype of vtc2 mature leaves to some degree. This was concurrent with a constitutively higher accumulation of zeaxanthin under this condition. When plants were shifted to natural conditions, with strongly fluctuating light and temperature, the ascorbate-deficient mature leaves of vtc2 bleached. Together, our results reveal developmental and environmental effects on VTC2-dependent ascorbate accumulation and function. The VTC2 enzyme controls ascorbate accumulation primarily in the mature leaves of Arabidopsis thaliana . The resulting developmental effects on photoprotection and stress resilience are environmentally modulated.

Due to low availability of CO 2 in aquatic environment, microalgae have evolved a CO 2 concentrating mechanism (CCM). It has long been thought that operation of CCM would suppress photorespiration by increasing the CO 2 concentration at the Rubisco active site, but experimental evidence is scarce. To better explore the function of photorespiration in algae, we first characterized a Chlamydomonas reinhardtii mutant defected in low-CO 2 inducible 20 (LCI20) and show that LCI20 is a chloroplast-envelope glutamate/malate transporter playing a role in photorespiration. By monitoring growth and glycolate excretion in mutants deficient in either CCM or photorespiration, we conclude that: ( i. ) CCM induction does not depend on photorespiration, ( ii. ) glycolate excretion together with glycolate dehydrogenase down-regulation prevents the toxic accumulation of non-metabolized photorespiratory metabolites, and ( iii .) photorespiration is active at low CO 2 when the CCM is operational. This work provides a foundation for a better understanding of the carbon cycle in the ocean where significant glycolate concentrations have been found.

High-altitude environments expose plants to increased levels of UV-B radiation, necessitating the evolution of protective mechanisms to mitigate stress. Buckwheat is a flavonoid-rich pseudocereal naturally adapted to high-altitude environments with elevated UV-B exposure. Although flavonoid biosynthesis is thought to contribute to this adaptation, the molecular and metabolic basis underlying flavonoid-mediated UV-B tolerance remains largely uncharacterized. In this study, we comprehensively assessed the relationship between flavonoid content and UV-B resistance across several cultivated and wild buckwheat species, including Fagopyrum esculentum, F. tataricum, F. cymosum, F. gracilipes and F. urophyllum. Our findings demonstrate that the synthesis of rutin strongly correlates with enhanced UV-B tolerance in buckwheat species, and the synthesis of rutin, along with isoquercitrin, positively influences the growth of diverse crops under UV-B stress. Functional validation of key enzymes revealed that the G125D variation in FtFLS4 and variations within the PGSG-box of FtRT1 significantly impact rutin-related metabolite synthesis in buckwheat. Notably, the Tartary buckwheat genes FtFLS4, FtUF3GT1 and FtRT1 exhibited both catalytic activity and UV-B inducible promoter responses, collectively underpinning F. tataricum's superior UV-B tolerance. Furthermore, we characterised the distinct UV-B response characteristics of FgFLS4 and FgFLS7 in the tetraploid wild buckwheat F. gracilipes, suggesting diversified adaptive strategies. Our findings provide novel insights into the functional basis of UV-B adaptation in Tartary buckwheat and offer potential targets for breeding or engineering UV-B-resilient crops.

The foundation of all biological processes is the network of diverse and dynamic protein interactions with other molecules in cells known as the interactome. Understanding the interactome is crucial for elucidating molecular mechanisms but has been a longstanding challenge. Recent developments in mass spectrometry (MS)-based techniques, including affinity purification, proximity labeling, cross-linking, and co-fractionation mass spectrometry (MS), have significantly enhanced our abilities to study the interactome. They do so by identifying and quantifying protein interactions, yielding profound insights into protein organizations and functions. This review summarizes recent advances in MS-based interactomics, focusing on the development of techniques that capture protein-protein, protein-metabolite, and protein-nucleic acid interactions. Additionally, we discuss how integrated MS-based approaches have been applied to diverse biological samples, focusing on significant discoveries that have leveraged our understanding of cellular functions. Finally, we highlight state-of-the-art bioinformatic approaches for predictions of interactome and complex modeling, as well as strategies for combining experimental interactome data with computation methods, thereby enhancing the ability of MS-based techniques to identify protein interactomes. Indeed, advances in MS technologies and their integrations with computational biology provide new directions and avenues for interactome research, leveraging new insights into mechanisms that govern the molecular architecture of living cells and, thereby, our comprehension of biological processes.