The plant hormone abscisic acid (ABA) is crucial for plant seed germination and abiotic stress tolerance. However, the association between ABA sensitivity and plant abiotic stress tolerance remains largely unknown. In this study, 436 rice accessions were assessed for their sensitivity to ABA during seed germination. The considerable diversity in ABA sensitivity among rice germplasm accessions was primarily reflected by the differentiation between the Xian (indica) and Geng (japonica) subspecies and between the upland-Geng and lowland-Geng ecotypes. The upland-Geng accessions were most sensitive to ABA. Genome-wide association analyses identified four major quantitative trait loci containing 21 candidate genes associated with ABA sensitivity of which a basic helix-loop-helix transcription factor gene, OsbHLH38, was the most important for ABA sensitivity. Comprehensive functional analyses using knockout and overexpression transgenic lines revealed that OsbHLH38 expression was responsive to multiple abiotic stresses. Overexpression of OsbHLH38 increased seedling salt tolerance, while knockout of OsbHLH38 increased sensitivity to salt stress. A salt-responsive transcription factor, OsDREB2A, interacted with OsbHLH38 and was directly regulated by OsbHLH38. Moreover, OsbHLH38 affected rice abiotic stress tolerance by mediating the expression of a large set of transporter genes of phytohormones, transcription factor genes, and many downstream genes with diverse functions, including photosynthesis, redox homeostasis, and abiotic stress responsiveness. These results demonstrated that OsbHLH38 is a key regulator in plant abiotic stress tolerance.

Seed weight is usually associated with seed size and is one of the important agronomic traits that determine yield. Understanding of seed weight control is limited, especially in soybean plants. Here we show that Glycine max JASMONATE-ZIM DOMAIN 3 (GmJAZ3), a gene identified through gene co-expression network analysis, regulates seed-related traits in soybean. Overexpression of GmJAZ3 promotes seed size/weight and other organ sizes in stable transgenic soybean plants likely by increasing cell proliferation. GmJAZ3 interacted with both G. max RESPONSE REGULATOR 18a (GmRR18a) and GmMYC2a to inhibit their transcriptional activation of cytokinin oxidase gene G. max CYTOKININ OXIDASE 3-4 (GmCKX3-4), which usually affects seed traits. Meanwhile, the GmRR18a binds to the promoter of GmMYC2a and activates GmMYC2a gene expression. In GmJAZ3-overexpressing soybean seeds, the protein contents were increased while the fatty acid contents were reduced compared to those in the control seeds, indicating that the GmJAZ3 affects seed size/weight and compositions. Natural variation in JAZ3 promoter region was further analyzed and Hap3 promoter correlates with higher promoter activity, higher gene expression and higher seed weight. The Hap3 promoter may be selected and fixed during soybean domestication. JAZ3 orthologs from other plants/crops may also control seed size and weight. Taken together, our study reveals a novel molecular module GmJAZ3-GmRR18a/GmMYC2a-GmCKXs for seed size and weight control, providing promising targets during soybean molecular breeding for better seed traits.

Exploiting novel endogenous glyphosate-tolerant alleles is highly desirable and has promising potential for weed control in rice breeding. Here, through fusions of different effective cytosine and adenine deaminases with nCas9-NG, we engineered an effective surrogate two-component composite base editing system, STCBE-2, with improved C-to-T and A-to-G base editing efficiency and expanded the editing window. Furthermore, we targeted a rice endogenous OsEPSPS gene for artificial evolution through STCBE-2-mediated near-saturated mutagenesis. After hygromycin and glyphosate selection, we identified a novel OsEPSPS allele with an Asp-213-Asn (D213N) mutation (OsEPSPS-D213N) in the predicted glyphosate-binding domain, which conferred rice plants reliable glyphosate tolerance and had not been reported or applied in rice breeding. Collectively, we developed a novel dual base editor which will be valuable for artificial evolution of important genes in crops. And the novel glyphosate-tolerant rice germplasm generated in this study will benefit weeds management in rice paddy fields.

Physical contact between genes distant on chromosomes is a potentially important way for genes to coordinate their expressions. To investigate the potential importance of distant contacts, we performed high-throughput chromatin conformation capture (Hi-C) experiments on leaf nuclei isolated from Brassica rapa and Brassica oleracea. We then combined our results with published Hi-C data from Arabidopsis thaliana. We found that distant genes come into physical contact and do so preferentially between the proximal promoter of one gene and the downstream region of another gene. Genes with higher numbers of conserved noncoding sequences (CNSs) nearby were more likely to have contact with distant genes. With more CNSs came higher numbers of transcription factor binding sites and more histone modifications associated with the activity. In addition, for the genes we studied, distant contacting genes with CNSs were more likely to be transcriptionally coordinated. These observations suggest that CNSs may enrich active histone modifications and recruit transcription factors, correlating with distant contacts to ensure coordinated expression. This study advances our knowledge of gene contacts and provides insights into the relationship between CNSs and distant gene contacts in plants.

Maize (Zea mays) cultivation is strongly affected by both abiotic and biotic stress, leading to reduced growth and productivity. It has recently become clear that regulators of plant stress responses, including the phytohormones abscisic acid (ABA), ethylene (ET), and jasmonic acid (JA), together with reactive oxygen species (ROS), shape plant growth and development. Beyond their well established functions in stress responses, these molecules play crucial roles in balancing growth and defense, which must be finely tuned to achieve high yields in crops while maintaining some level of defense. In this review, we provide an in-depth analysis of recent research on the developmental functions of stress regulators, focusing specifically on maize. By unraveling the contributions of these regulators to maize development, we present new avenues for enhancing maize cultivation and growth while highlighting the potential risks associated with manipulating stress regulators to enhance grain yields in the face of environmental challenges.

Inorganic phosphate (Pi) availability is an important factor which affects the growth and yield of crops, thus an appropriate and effective response to Pi fluctuation is critical. However, how crops orchestrate Pi signaling and growth under Pi starvation conditions to optimize the growth defense tradeoff remains unclear. Here we show that a Pi starvation-induced transcription factor NIGT1 (NITRATE-INDUCIBLE GARP-TYPE TRANSCRIPTIONAL REPRESSOR 1) controls plant growth and prevents a hyper-response to Pi starvation by directly repressing the expression of growth-related and Pi-signaling genes to achieve a balance between growth and response under a varying Pi environment. NIGT1 directly binds to the promoters of Pi starvation signaling marker genes, like IPS1, miR827, and SPX2, under Pi-deficient conditions to mitigate the Pi-starvation responsive (PSR). It also directly represses the expression of vacuolar Pi efflux transporter genes VPE1/2 to regulate plant Pi homeostasis. We further demonstrate that NIGT1 constrains shoot growth by repressing the expression of growth-related regulatory genes, including brassinolide signal transduction master regulator BZR1, cell division regulator CYCB1;1, and DNA replication regulator PSF3. Our findings reveal the function of NIGT1 in orchestrating plant growth and Pi starvation signaling, and also provide evidence that NIGT1 acts as a safeguard to avoid hyper-response during Pi starvation stress in rice.

Plant cells possess a two-layered immune system consisting of pattern-triggered immunity (PTI) and effector-triggered immunity (ETI), mediated by cell surface pattern-recognition receptors and intracellular nucleotide-binding leucine-rich repeat receptors (NLRs), respectively. The CONSTITUTIVE EXPRESSION OF PR GENES 5 (CPR5) nuclear pore complex protein negatively regulates ETI, including ETI-associated hypersensitive response. Here, we show that CPR5 is essential for the activation of various PTI responses in Arabidopsis, such as resistance to the non-adapted bacterium Pseudomonas syringae pv. tomato DC3000 hrcC^-. In a forward-genetic screen for suppressors of cpr5, we identified the mediator protein MED4. Mutation of MED4 in cpr5 greatly restored the defective PTI of cpr5. Our findings reveal that CPR5 plays opposite roles in regulating PTI and ETI, and genetically regulates PTI via MED4.

Drought seriously impacts wheat production (Triticum aestivum L.), while the exploitation and utilization of genes for drought tolerance are insufficient. Leaf wilting is a direct reflection of drought tolerance in plants. Clade A PP2Cs are abscisic acid (ABA) co-receptors playing vital roles in the ABA signaling pathway, regulating drought response. However, the roles of other clade PP2Cs in drought tolerance, especially in wheat, remain largely unknown. Here, we identified a gain-of-function drought-induced wilting 1 (DIW1) gene from the wheat Aikang 58 mutant library by map-based cloning, which encodes a clade I protein phosphatase 2C (TaPP2C158) with enhanced protein phosphatase activity. Phenotypic analysis of overexpression and CRISPR/Cas9 mutant lines demonstrated that DIW1/TaPP2C158 is a negative regulator responsible for drought resistance. We found that TaPP2C158 directly interacts with TaSnRK1.1 and de-phosphorylates it, thus inactivating the TaSnRK1.1–TaAREB3 pathway. TaPP2C158 protein phosphatase activity is negatively correlated with ABA signaling. Association analysis suggested that C-terminal variation of TaPP2C158 changing protein phosphatase activity is highly correlated with the canopy temperature, and seedling survival rate under drought stress. Our data suggest that the favorable allele with lower phosphatase activity of TaPP2C158 has been positively selected in Chinese breeding history. This work benefits us in understanding the molecular mechanism of wheat drought tolerance, and provides elite genetic resources and molecular markers for improving wheat drought tolerance.

Reactive oxygen species (ROS) are produced as undesirable by-products of metabolism in various cellular compartments, especially in response to unfavorable environmental conditions, throughout the life cycle of plants. Stress-induced ROS production disrupts normal cellular function and leads to oxidative damage. To cope with excessive ROS, plants are equipped with a sophisticated antioxidative defense system consisting of enzymatic and non-enzymatic components that scavenge ROS or inhibit their harmful effects on biomolecules. Nonetheless, when maintained at relatively low levels, ROS act as signaling molecules that regulate plant growth, development, and adaptation to adverse conditions. Here, we provide an overview of current approaches for detecting ROS. We also discuss recent advances in understanding ROS signaling, ROS metabolism, and the roles of ROS in plant growth and responses to various abiotic stresses.

Although root nodules are essential for biological nitrogen fixation in legumes, the cell types and molecular regulatory mechanisms contributing to nodule development and nitrogen fixation in determinate nodule legumes, such as soybean (Glycine max), remain incompletely understood. Here, we generated a single-nucleus resolution transcriptomic atlas of soybean roots and nodules at 14 days post inoculation (dpi) and annotated 17 major cell types, including six that are specific to nodules. We identified the specific cell types responsible for each step in the ureides synthesis pathway, which enables spatial compartmentalization of biochemical reactions during soybean nitrogen fixation. By utilizing RNA velocity analysis, we reconstructed the differentiation dynamics of soybean nodules, which differs from those of indeterminate nodules in Medicago truncatula. Moreover, we identified several putative regulators of soybean nodulation and two of these genes, GmbHLH93 and GmSCL1, were as-yet uncharacterized in soybean. Overexpression of each gene in soybean hairy root systems validated their respective roles in nodulation. Notably, enrichment for cytokinin-related genes in soybean nodules led to identification of the cytokinin receptor, GmCRE1, as a prominent component of the nodulation pathway. GmCRE1 knockout in soybean resulted in a striking nodule phenotype with decreased nitrogen fixation zone and depletion of leghemoglobins, accompanied by downregulation of nodule-specific gene expression, as well as almost complete abrogation of biological nitrogen fixation. In summary, this study provides a comprehensive perspective of the cellular landscape during soybean nodulation, shedding light on the underlying metabolic and developmental mechanisms of soybean nodule formation.

The plant hormone ethylene is essential for climacteric fruit ripening, although it is unclear how other phytohormones and their interactions with ethylene might affect fruit ripening. Here, we explored how brassinosteroids (BRs) regulate fruit ripening in tomato (Solanum lycopersicum) and how they interact with ethylene. Exogenous BR treatment and increased endogenous BR contents in tomato plants overexpressing the BR biosynthetic gene SlCYP90B3 promoted ethylene production and fruit ripening. Genetic analysis indicated that the BR signaling regulators Brassinazole-resistant1 (SlBZR1) and BRI1-EMS-suppressor1 (SlBES1) act redundantly in fruit softening. Knocking out SlBZR1 inhibited ripening through transcriptome reprogramming at the onset of ripening. Combined transcriptome deep sequencing and chromatin immunoprecipitation followed by sequencing identified 73 SlBZR1-repressed targets and 203 SlBZR1-induced targets involving major ripening-related genes, suggesting that SlBZR1 positively regulates tomato fruit ripening. SlBZR1 directly targeted several ethylene and carotenoid biosynthetic genes to contribute to the ethylene burst and carotenoid accumulation to ensure normal ripening and quality formation. Furthermore, knock-out of Brassinosteroid-insensitive2 (SlBIN2), a negative regulator of BR signaling upstream of SlBZR1, promoted fruit ripening and carotenoid accumulation. Taken together, our results highlight the role of SlBZR1 as a master regulator of tomato fruit ripening with potential for tomato quality improvement and carotenoid biofortification.

Common buckwheat (Fagopyrum esculentum) and Tartary buckwheat (Fagopyrum tataricum), the two most widely cultivated buckwheat species, differ greatly in flavonoid content and reproductive mode. Here, we report the first high-quality and chromosome-level genome assembly of common buckwheat with 1.2 Gb. Comparative genomic analysis revealed that common buckwheat underwent a burst of long terminal repeat retrotransposons insertion accompanied by numerous large chromosome rearrangements after divergence from Tartary buckwheat. Moreover, multiple gene families involved in stress tolerance and flavonoid biosynthesis such as multidrug and toxic compound extrusion (MATE) and chalcone synthase (CHS) underwent significant expansion in buckwheat, especially in common buckwheat. Integrated multi-omics analysis identified high expression of catechin biosynthesis-related genes in flower and seed in common buckwheat and high expression of rutin biosynthesis-related genes in seed in Tartary buckwheat as being important for the differences in flavonoid type and content between these buckwheat species. We also identified a candidate key rutin- degrading enzyme gene (Ft8.2377) that was highly expressed in Tartary buckwheat seed. In addition, we identified a haplotype-resolved candidate locus containing many genes reportedly associated with the development of flower and pollen, which was potentially related to self-incompatibility in common buckwheat. Our study provides important resources facilitating future functional genomics-related research of flavonoid biosynthesis and self- incompatibility in buckwheat.

Nitrogen (N) is an essential macronutrient for plants and profoundly affects crop yields and qualities. Ammonium (NH₄⁺) and nitrate (NO₃?) are major inorganic N forms absorbed by plants from the surrounding environments. Intriguingly, NH₄⁺ is usually toxic to plants when it serves as the sole or dominant N source. It is thus important for plants to coordinate the utilization of NH₄⁺ and the alleviation of NH₄⁺ toxicity. To fully decipher the molecular mechanisms underlying how plants minimize NH₄⁺ toxicity may broadly benefit agricultural practice. In the current minireview, we attempt to discuss recent discoveries in the strategies for mitigating NH₄⁺ toxicity in plants, which may provide potential solutions for improving the nitrogen use efficiency (NUE) and stress adaptions in crops.

The phragmoplast, a structure crucial for the completion of cytokinesis in plant cells, is composed of antiparallel microtubules (MTs) and actin filaments (AFs). However, how the parallel structure of phragmoplast MTs and AFs is maintained, especially during centrifugal phragmoplast expansion, remains elusive. Here, we analyzed a new Arabidopsis thaliana MT and AF crosslinking protein (AtMAC). When AtMAC was deleted, the phragmoplast showed disintegrity during centrifugal expansion, and the resulting phragmoplast fragmentation led to incomplete cell plates. Overexpression of AtMAC increased the resistance of phragmoplasts to depolymerization and caused the formation of additional phragmoplasts during cytokinesis. Biochemical experiments showed that AtMAC crosslinked MTs and AFs in vitro, and the truncated AtMAC protein, N-CC1, was the key domain controlling the ability of AtMAC. Further analysis showed that N-CC1(51–154) is the key domain for binding MTs, and N-CC1(51–125) for binding AFs. In conclusion, AtMAC is the novel MT and AF crosslinking protein found to be involved in regulation of phragmoplast organization during centrifugal phragmoplast expansion, which is required for complete cytokinesis.

Auxin is an important phytohormone in plants, and auxin signaling pathways in rice play key roles in regulating its growth, development, and productivity. To investigate how rice grain yield traits are regulated by auxin signaling pathways and to facilitate their application in rice improvement, we validated the functional relationships among regulatory genes such as OsIAA10, OsSK41, and OsARF21 that are involved in one of the auxin (OsIAA10) signaling pathways. We assessed the phenotypic effects of these genes on several grain yield traits across two environments using knockout and/or overexpression transgenic lines. Based on the results, we constructed a model that showed how grain yield traits were regulated by OsIAA10 and OsTIR1, OsAFB2, and OsSK41 and OsmiR393 in the OsSK41-OsIAA10-OsARF module and by OsARF21 in the transcriptional regulation of downstream auxin response genes in the OsSK41-OsIAA10-OsARF module. The population genomic analyses revealed rich genetic diversity and the presence of major functional alleles at most of these loci in rice populations. The strong differentiation of many major alleles between Xian/indica and Geng/japonica subspecies and/or among modern varieties and landraces suggested that they contributed to improved productivity during evolution and breeding. We identified several important aspects associated with the genetic and molecular bases of rice grain and yield traits that were regulated by auxin signaling pathways. We also suggested rice auxin response factor (OsARF) activators as candidate target genes for improving specific target traits by overexpression and/or editing subspecies-specific alleles and by searching and pyramiding the ‘best’ gene allelic combinations at multiple regulatory genes in auxin signaling pathways in rice breeding programs.

Abiotic stress is one of the most important factors reducing soybean yield. It is essential to identify regulatory factors contributing to stress responses. A previous study found that the tandem CCCH zinc-finger protein GmZF351 is an oil level regulator. In this study, we discovered that the GmZF351 gene is induced by stress and that the overexpression of GmZF351 confers stress tolerance to transgenic soybean. GmZF351 directly regulates the expression of GmCIPK9 and GmSnRK, leading to stomata closing, by binding to their promoter regions, which carry two CT(G/C)(T/A)AA elements. Stress induction of GmZF351 is mediated through reduction in the H3K27me3 level at the GmZF351 locus. Two JMJ30-demethylase-like genes, GmJMJ30-1 and GmJMJ30-2, are involved in this demethylation process. Overexpression of GmJMJ30-1/2 in transgenic hairy roots enhances GmZF351 expression mediated by histone demethylation and confers stress tolerance to soybean. Yield-related agronomic traits were evaluated in stable GmZF351-transgenic plants under mild drought stress conditions. Our study reveals a new mode of GmJMJ30-GmZF351 action in stress tolerance, in addition to that of GmZF351 in oil accumulation. Manipulation of the components in this pathway is expected to improve soybean traits and adaptation under unfavorable environments.

Establishing how lineages with similar traits are phylogenetically related remains critical for understanding the origin of biodiversity on Earth. Floral traits in plants are widely used to explore phylogenetic relationships and to delineate taxonomic groups. The subtribe Swertiinae (Gentianaceae) comprises more than 350 species with high floral diversity ranging from rotate to tubular corollas and possessing diverse nectaries. Here we performed phylogenetic analysis of 60 species from all 15 genera of the subtribe Swertiinae sensu Ho and Liu, representing the range of floral diversity, using data from the nuclear and plastid genomes. Extensive topological conflicts were present between the nuclear and plastome trees. Three of the 15 genera represented by multiple species are polyphyletic in both trees. Key floral traits including corolla type, absence or presence of lobe scales, nectary type, nectary position, and stigma type are randomly distributed in the nuclear and plastome trees without phylogenetic correlation. We also revealed the likely ancient hybrid origin of one large clade comprising 10 genera with diverse floral traits. These results highlight the complex evolutionary history of this subtribe. The phylogenies constructed here provide a basic framework for further exploring the ecological and genetic mechanisms underlying both species diversification and floral diversity.