ABSTRACT To reduce the environmental burden associated with excessive nitrogen fertilizer use in rice cultivation, we developed NR160E, an early heading isogenic line derived from the high‐yielding variety NSIC Rc 160 (NR160). Although early heading is often considered unfavorable for yield and excluded from high‐yield breeding programs, our results challenge this assumption. In field trials conducted under natural long‐day conditions in Tsukuba, Japan (planting density of 18.5 plants m −2 ), heading occurred ~8–10 days earlier in NR160E than in NR160 across all nitrogen fertilization levels (0, 4.8, and 9.6 g N m −2 ). In NR160E, early heading was associated with reduced plant height and a marked increase in harvest index (HI). Specifically, plant height decreased by ~15%, whereas HI increased by 20% compared with NR160, indicating efficient biomass partitioning and suggesting its potential within this genetic and environmental context. Fine‐mapping identified a 33.3‐kbp region on chromosome 3 containing four annotated genes ( Os03G0122600 , Os03g0123100 , Os03g0123200 , and Os03g0123300 ) among which OsMADS50 ( Os03G0122600 ) was upregulated in NR160E under long‐day conditions, accompanied by elevated Ehd1 and RFT1 expression, consistent with the early heading phenotype. Photoperiod experiments revealed that heading occurred significantly earlier in NR160E than in NR160 under long‐day conditions (13.0–14.5 h), whereas no significant difference was observed under short‐day conditions (10.0–11.5 h), indicating that the expression of early heading in NR160E depends strongly on photoperiodic conditions. Field trials across 12 cultivation regimes varying in nitrogen input and planting density consistently showed that NR160E outperformed NR160 in grain yield. At 18.5 plants m −2 , NR160E produced grain yields of 649.0, 797.7, and 922.9 g m −2 under 0‐, 4.8‐, and 9.6‐g N m −2 fertilization, respectively, whereas NR160 yielded 513.0, 666.4, and 692.3 g m −2 under the same conditions. Within each nitrogen–field condition, NR160E produced higher yield than NR160. Seed fertility was consistently higher in NR160E across nitrogen–field conditions (e.g., 0.815 under zero nitrogen input), and NR160E also exhibited a consistently higher agronomical HI, consistent with more efficient assimilate allocation to grain. Growth analysis showed that NR160E maintained dry matter production during grain filling, coinciding with its higher HI and physiological nitrogen‐use efficiency. Although these relationships are observational and the physiological basis remains unclear, the trend was consistent across conditions. Regression analysis also revealed a steeper slope between nitrogen uptake and grain yield in NR160E than in NR160, consistent with higher physiological nitrogen‐use efficiency (as defined in this study). Collectively, these results suggest that earlier heading may be associated with sustained productivity under the long‐day field conditions examined here. However, determining how this trait could inform fertilizer management or nitrogen‐input strategies will require multienvironment and multibackground validation.

The C4 photosynthesis includes intriguing leaf anatomies. The current model supports the placement of C3-C4 intermediates as a middle point in the evolutionary trajectory from C3 to C4 photosynthesis. The known determinants involved in the differentiation of divergent photosynthetic leaves arose from the comparative analysis between both ends, C3 and C4 species. However, much more could be known if evolutionarily close species were analyzed together with intermediate species using advanced-omic approaches. In the present work, by combining leaf anatomical traits and transcriptomic data with machine learning methods, we provided insights on gene regulatory networks involved in complex leaf anatomical characteristics in non-model grasses of subtribe Otachyriinae. For that, self-organizing maps (SOMs) were developed to group genes and phenotypic traits into clusters (neurons) according to their behavior along the leaf developmental gradient. The analysis allowed us to identify a set of genes as potential enablers of key anatomical trait differentiation related to bundle sheath (BS) cell size, vein density, and the interface between mesophyll and BS cells. At the same time, we identified genes that displaced together with the adjustment of the BS cell area suggesting a possible role in the evolution of this distinctive leaf anatomical trait.

ABSTRACTFerula violacea Korovin, an endemic Tajikistani plant with purported medicinal properties, remains understudied. This study employs untargeted metabolomics to characterize the metabolite profiles of ethanol extracts and juices from F. violacea roots and seeds. In total, 540 distinct metabolites are putatively identified, 419 of which are previously unreported in the Ferula genus, representing a substantial expansion of its known chemical diversity. The most abundant metabolites are terpenoids, amino acid derivatives, and alkaloids. A particularly abundant group of daucane sesquiterpenoids, sharing a common (6‐methyl‐azulen‐4‐yl)cyclohexanecarboxylate substructure, is identified, including known metabolites such as ferutidin and ferutinin. Comparative analysis reveals organ‐specific metabolic specialization: roots are enriched in terpenoids, whereas seeds exhibit higher concentrations of alkaloids and amino acids. Additionally, processing methods influence metabolite composition, with ethanol extracts being rich in terpenoids and amino acids, and juices displaying a greater diversity of phenylpropanoid‐derived compounds. These findings expand the phytochemical richness of F. violacea and suggest its potential as a valuable source of bioactive compounds for pharmacological exploration.

ABSTRACT The ubiquitin‐dependent Arg/N‐degron pathway relates the stability of a substrate protein to the nature of its N‐terminal amino acid residue or its biochemical modifications, with some N‐terminal residues being recognized by specific E3 ubiquitin ligases, resulting in the ubiquitylation and degradation of the substrate protein. Work in the model plant Arabidopsis thaliana has shown that the Arg/N‐degron pathway is a key regulator of plant responses to hypoxia, which can be either physiological or a stress in the context of waterlogging or submergence. The role of the Arg/N‐degron pathway in hypoxia response is mediated via the oxygen‐dependent degradation of group VII ETHYLENE RESPONSE FACTOR (ERFVII) transcription factors, which act as the master regulators of the hypoxia response program in plants. Analysis of Arabidopsis mutants for different enzymatic components of the Arg/N‐degron pathway has also revealed its roles in the regulation of responses to other abiotic stresses (e.g., salt stress), as well as to pathogens. Although much has been learned from studies in Arabidopsis about the functions of the Arg/N‐degron pathway, very little is known about this pathway in crops, including in Brassica crops such as oilseed rape, cabbage, or turnip. To determine functional similarities and divergence of the Arg/N‐degron pathway between Arabidopsis and Brassica crops, we isolated and characterized the first Arg/N‐degron pathway mutants in Brassica rapa (turnip, pak choi), a diploid Brassica crop closely related to oilseed rape. We focused on two enzymatic components, namely, the arginine‐transferases ( ATE s) and the E3 ubiquitin ligase PROTEOLYSIS6 ( PRT6 ). Our results show both similarities and divergence of function for these Arg/N‐degron pathway components in B. rapa compared to Arabidopsis. Specifically, ATE mutants in B. rapa arrest their development at the seedling stage, which contrasts with the mild phenotypic defects of the equivalent Arabidopsis mutants. Double mutant lines for two of the three PRT6 genes in B. rapa indicated a constitutive activation of hypoxia response genes at the transcriptional level, as shown in the single prt6 mutant in Arabidopsis. However, contrary to Arabidopsis, the B. rapa double mutants were more sensitive to waterlogging and hypoxia and did not show differential response to salt stress or to biotic stress compared to the wild type. The functional divergence identified likely reflects variability in each species in the substrate repertoire and/or in the regulation of pathways or targets downstream of Arg/N‐degron pathway substrates. Such differences could be driven by direct selective pressures at N‐termini (e.g., gain or loss of a destabilizing N‐terminal residue) or by species‐specific proteases that may generate destabilizing neo‐N‐termini after cleavage. These similarities and differences highlight the difficulties in translating research findings from Arabidopsis to crops, even within the same plant family (Brassicaceae), and highlight the need to study pathways in crops.

Radish (Raphanus sativus L.) is an important root vegetable in the cruciferous family. The yield and quality of radish is seriously affected by the premature bolting and flowering. Although the microRNAs (miRNAs) in regulating flower development have been established in radish, the identification and characterization of long noncoding RNAs (lncRNAs) have yet to be explored. In this study, miRNAs and lncRNAs in vegetative and flower stage were conducted by RNA-seq and small RNA sequencing, respectively. A total of 5315 differentially expressed genes (DGEs), 263 DElncRNAs, and 38 DEmiRNAs were detected in two stages. GO analysis found that many flower DGEs associated with reproductive process, response to hormone, and pollination were enriched. In total, 202 DElncRNAs and 257 DElncRNAs were found to have potential cis- and trans-regulatory effects on 572 DEmRNAs and 3902 DGEs, respectively. A total of 93 and 82 DEGs were predicted as putative targets of 31 DEmiRNAs and 29 DEmiRNAs, respectively. Five mRNA-lncRNA-miRNA regulatory pairs involved in flowering time regulation were proposed, including miRNA156a-5p, miRNA399b, miRNA novel-23, miRNA164c-5p, and miRNA165a-5p. The qRT-PCR results showed that four mRNAs, three lncRNAs, and three miRNAs were consistent with the results of RNA-seq and small RNA sequencing. Transient overexpression of miR156a-5p significantly inhibited the expression levels of RsSPL10, RsSPL15, lncRNAs RsLinc1162, and RsLinc214. The results showed that miR156 co-expressed with RsSPL10 and RsSPL15 significantly inhibited the luciferase activity of RsSPL10 and RsSPL15 genes, indicating miR156 can directly target RsSPL10 and RsSPL15 and inhibit their expression. These findings provide a theoretical foundation for further elucidating the molecular regulation mechanism of mRNAs, lncRNAs, and miRNAs in bolting and flowering in radish.

ABSTRACTThe pulp color of jackfruit reflects variations in its nutritional composition and influences market preference. We investigated the mechanism underlying pulp coloration through an integrated transcriptomic and metabolomic analysis of three jackfruit cultivars—light yellow “THA”, yellow “GTM”, and orange “YNH”. Twenty‐five differentially accumulated flavonoids and 32 differentially accumulated carotenoids were identified. Naringenin chalcone, eriodictyol, taxifolin, zeaxanthin, and lutein were identified as the key flavonoids and carotenoids associated with the light‐yellow tone of THA pulp. Phlorizin and lutein were associated with the yellow tone of GTM pulp, whereas apigenin, luteolin, zeaxanthin, and violaxanthin dipalmitate were the key pigments regulating the orange tone of YNH pulp. Differentially expressed genes involved in flavonoid and carotenoid biosynthesis included PAL, C4H, 4CL, F3H, FLS, ANS, ANR, PSY, PDS, ZDS, LCYE, LUT5, ZEP, and VDE. This study provides a foundation for elucidating the molecular mechanisms underlying jackfruit pulp coloration at transcriptional and metabolic levels.

ABSTRACTDrought has emerged as one of the most severe and widespread environmental stresses affecting plants. Crops exposed to varying levels of drought, ranging from moderate to severe, often experience notable declines in yield or reduced harvest quality. Investigating the molecular mechanisms and cellular factors involved in plant defense against drought is crucial—not only for advancing our understanding of these processes but also for ensuring sustainable food production and supporting humanity's survival. Our previous work identified the small intrinsically disordered protein DSS1 (deleted in split‐hand/split‐foot) as a key factor in the stress defense mechanisms of Arabidopsis thaliana. The absence of DSS1(V) led to increased sensitivity of plants to oxidative stress induced by hydrogen peroxide or methyl viologen. As drought can induce oxidative stress in plant cells, we investigated if DSS1(V) protein can mitigate stress caused by mild to moderate drought. Alongside the wild‐type (WT) strain, the analysis included knockout plants lacking the DSS1(V) gene and plants overexpressing this gene. Various stress‐related parameters, including lipid peroxidation, total phenol content, chlorophyll levels, and protein oxidation, were measured. Results indicated that the DSS1(V) knockout line displayed significantly higher sensitivity to drought compared to WT plants. However, elevated levels of DSS1(V) transcripts in the overexpressing lines did not confer a protective effect, as these lines did not exhibit reduced drought sensitivity. These findings provide compelling evidence highlighting the critical involvement of the DSS1(V) protein in the mechanisms underlying plant responses to environmental stress, particularly water deficiency. This protein appears to enable plants to cope with the challenges posed by drought conditions, emphasizing its importance in maintaining cellular homeostasis and mitigating the adverse effects of water scarcity.

Soybean (Glycine max) seed coat color variation is determined by the accumulation of flavonoid-derived pigments, although the molecular mechanisms underlying this trait remain poorly understood. This study integrated RNA sequencing (RNA-Seq) and high-performance liquid chromatography (HPLC)-based metabolite measurements to investigate black and yellow seed coat soybean lines derived from the same genetic background. Metabolite analysis revealed significantly higher total phenolic content (TPC), total flavonoid content (TFC), total anthocyanin content (TAC), total proanthocyanidin content (TPAC), and antioxidant activity (DPPH, ABTS) in black seed coats, whereas yellow seed coats exhibited elevated total isoflavone content (TIC). RNA-seq at 110 days after sowing (DAS) identified differential expression of flavonoid pathway genes associated with these metabolic differences. Genes upregulated in black seed coats included flavanone 3-hydroxylase (F3H), anthocyanidin synthase (ANS), UDP-glycosyltransferases (UGT78D2, UGT79B6), and glutathione S-transferase (GSTF11), encoding enzymes reported to function in anthocyanin biosynthesis, glycosylation, and vacuolar transport, respectively. Conversely, leucoanthocyanidin reductase (LAR) genes showed higher expression in yellow seed coats despite lower proanthocyanidin (PA) levels, whereas LAC5 exhibited black seed-specific expression consistent with potential PA polymerization activity. R2R3-MYB transcription factor genes along with small heat shock protein genes (sHSPs) were also upregulated in black seed coats, suggesting candidate regulatory roles in pigmentation and stress responses. Cytochrome P450 genes showed preferential expression in yellow seed coats, consistent with isoflavonoid pathway activation. Together, these findings elucidate the genetic and metabolic regulation of seed coat color in soybean and identify candidate genes relevant for functional breeding and genomics research.

ABSTRACTChaperones are essential for facilitating the import of nuclear‐encoded precursor proteins into chloroplasts. In the intermembrane space (IMS) of the chloroplasts, this process is mediated by the transport‐associated domains (POTRA) of the translocon at the outer envelope membrane (Toc75) and translocon at the inner envelope membrane (Tic22) proteins. The present work aims to understand the interaction between the Toc75 POTRA domain and Tic22 in the IMS and determine their relationship in facilitating protein import. Expression of the POTRA1 domain deleted TOC75 (TOC75ΔP1) in the tic22‐III mutant background resulted in a more severe phenotype than the individual mutants, indicating that the two proteins functionally interact in the IMS. Using an insulin aggregation assay, we have demonstrated that Tic22‐III also possesses chaperone‐like activity. In vitro import experiments suggest that TOC75ΔP1/tic22‐III plants are compromised in importing stromal and thylakoid membrane proteins. Therefore, we propose that the Toc75 POTRA domains and Tic22‐III both provide chaperone activity necessary to prevent the misfolding of incoming pre‐proteins, acting as chaperones and facilitating the protein import process through the IMS of the chloroplast.