Peisheng Mao’s research while affiliated with China Agricultural University and other places

Background Alfalfa is widely regarded as one of the most important forage crops globally. However, its growth and development are primarily constrained by various abiotic stresses. FIMBRINs are crucial actin-binding proteins involved in regulating cellular dynamics in plants under various stress conditions and during developmental processes. The Fimbrin (FIM) gene family has been reported only in Arabidopsis, while a comprehensive identification of the FIM gene family in alfalfa and the responses of its members to abiotic stresses remain unclear. Results In this study, six MsFIM genes were identified in the alfalfa genome, distributed across three chromosomes. Phylogenetic analysis grouped these genes into four clades, all containing the conserved CH domain. Gene duplication events suggested that large fragment duplications contribute to gene amplification. Furthermore, cis-regulatory element analysis highlighted their pivotal roles in plant development and response to external abiotic stresses. RT-qPCR analyses revealed that the MsFIM genes exhibited differential expression across various tissues, with predominant expression in flowers, stems, and leaves. The MsFIM genes showed elevated expression under abiotic stresses (drought, cold, and salt) as well as hormone treatment (abscisic acid, ABA), suggesting that they served as positive regulators in alfalfa’s resistance to abiotic stresses and its growth and development. Conclusions This study investigates the MsFIM genes in alfalfa, further analyzing their potential roles in plant development and response to abiotic stresses. These findings will provide novel insights into the molecular mechanisms of alfalfa's stress response.

Abstract： This study explores the effects of nitrogen application on superior and inferior grains in smooth bromegrass (Bromus inermis). A randomized block design with two nitrogen treatments (0 and 200 kg·N·ha⁻1) during the 2021-2022 growing seasons. Seed dry weight, fresh weight, and storage protein content were measured at multiple stages after anthesis. PacBio full-length transcriptome sequencing generated a comprehensive transcriptome consisting of 124,425 high-quality transcripts, and metabolomic profiling was performed across developmental stages. The results showed that nitrogen application significantly increased both dry and fresh weights, as well as storage protein content, particularly gliadin and glutelin. Notably, nitrogen treatment upregulated glutamate and asparagine levels in both superior and inferior grains. Transcriptomic analysis revealed enhanced nitrogen transport and protein synthesis pathways. Additionally, two α-gliadin genes, BiGli1 and BiGli2, were identified in inferior grains as nitrogen-responsive. Genetic transformation studies in Arabidopsis confirmed that these genes regulate seed size and vigor, improving seed performance by reducing mean germination time under aging conditions. This study highlights the critical role of α-gliadin in enhancing seed quality, particularly in promoting the development of inferior grains, offering valuable insights for the development of high-yield seed varieties and the optimization of specialized forage seed production. Keywords: α-gliadin, nitrogen application, seed vigor, superior and inferior grains, storage protein, Smooth bromegrass, Multi-omics

Abstract： The common oat (Avena sativa L.) is a globally important hexaploid cereal crop with a large and complex genome, yet its mitochondrial genome has remained unresolved. Here, we report the first complete assembly and comprehensive analysis of the A. sativa mitogenome, which comprises two circular chromosomes totaling 634,262 bp with a GC content of 44.11%. The mitogenome encodes 78 genes, including 37 protein-coding genes, 27 tRNAs, 12 rRNAs, and 2 pseudogenes. Extensive repeat sequences were identified, providing a basis for mitochondrial genome recombination and structural diversity. Codon usage analysis revealed a strong AT bias and distinct codon preferences, while Ka/Ks analysis indicated predominant purifying selection on core OXPHOS genes, with possible signals of positive selection in mttB and nad7. A total of 463 RNA editing sites were predicted, predominantly enhancing protein hydrophobicity and potentially optimizing mitochondrial function. Comparative analyses revealed high collinearity with diploid oat (Avena longiglumis) and frequent chloroplast-to-mitochondrion DNA transfers, highlighting dynamic organellar genome evolution in Poaceae. Phylogenetic reconstruction based on 32 conserved mitochondrial genes robustly resolved relationships within Poaceae and its relatives. Transcriptomic profiling during seed germination demonstrated rapid upregulation of mitochondrial oxidative phosphorylation (OXPHOS) genes during early imbibition, with aged seeds showing even higher expression levels, suggesting compensatory responses to compromised mitochondrial function. This study provides the first complete mitogenome of hexaploid oat, offering valuable insights into mitochondrial genome structure, evolution, and OXPHOS gene expression dynamics, and serves as an important resource for future functional genomics and molecular breeding in oat and related crops. Key words: Avena sativa, mitochondrial genome, RNA editing, OXPHOS, seed germination.

Soil salinity is a significant environmental challenge that adversely affects plant yield and quality. Zoysiagrass (Zoysia japonica), a member of the Gramineae family, is highly salt-tolerant, making it an excellent model for studying salt stress response mechanisms. We performed physiological and transcriptomic analyses on two contrasting Zoysiagrass germplasm accessions under high salt conditions. The salt-tolerant germplasm ST68 demonstrated superior growth phenotypes, higher chlorophyll and relative water content, greater photochemical efficiency, and lower relative electrolyte leakage and sodium ion content compared to the salt-sensitive germplasm SS9. Transcriptomic analysis revealed differential expression in pathways involved in photosynthesis, flavonoid biosynthesis, cell wall macromolecule catabolism, phosphate ion homeostasis, and reactive oxygen species response in the tolerant vs the sensitive line under salt stress. Notably, the ZjHEMA gene, which encodes glutamyl-tRNA reductase, a rate-limiting enzyme in chlorophyll biosynthesis, was identified as a key regulator due to its significant upregulation under salt stress in the salt-tolerant germplasm, compared to the sensitive one. Overexpression of the salt-responsive glutamyl-tRNA reductase gene, associated with chlorophyll metabolism in Zoysiagrass, in Arabidopsis led to increased salt tolerance, as evidenced by elevated chlorophyll content, relative water content, and photochemical efficiency compared to wild-type plants. Our findings offer new insights into the mechanisms of salt tolerance in Zoysiagrass, laying a foundation for breeding salt-tolerant germplasm.

Seed priming is an effective way to activate defense mechanisms before germination, enhancing seed vigor and stress resistance. Ascorbic acid (AsA) is an important signaling molecule that plays a crucial role in balancing cellular reactive oxygen species. However, whether AsA priming can enhance seed vigor in oat (Avena sativa) and the underlying mechanisms remain unclear. This study primed aged seeds (controlled deterioration at 45°C for 5 days) with 1.5 mM AsA for 24 h. Primed seeds were then sampled after 36 h of imbibition for seed assays. Significant increases in germination percentage, vigor index, shoot and root length, coupled with a significant reduction in mean germination time, demonstrated that AsA priming effectively restored seed vigor. Ultrastructural observations of mitochondria isolated from embryos presented that AsA priming repaired structural damage in aged seeds, with intact double membranes and clear internal cristae observed. Excessive H2O2 accumulation was discovered in mitochondria of aged seeds, while AsA priming reduced H2O2 levels by increasing the activities of CAT, GR, MDHAR and DHAR. AsA priming also increased antioxidant content, particularly DHA, contributing to reduced oxidative stress. Furthermore, transcriptomic analysis highlighted the upregulation of genes associated with antioxidant defense, including APX, CAT, DHAR and MDHAR, indicating enhanced repair and protection pathways in the mitochondrial AsA‐GSH cycle. This suggests that AsA priming would increase the activity of antioxidant enzymes, the content of antioxidants, and expression of genes related to AsA‐GSH cycle in aged oat seeds, which was conducive to repairing mitochondrial damage and enhancing seed vigor.

Smooth bromegrass (Bromus inermis) was adopted as experiment materials for identifying the seed maturity using a combination of multispectral imaging and machine learning. The trials were conducted to investigate the effects of three nitrogen application levels (0, 100 and 200 kg N ha− 1, defined as CK, N1 and N2 respectively) and two spikelet grain positions: superior grain (SG) at the basal position and inferior grain (IG) at the upper position, on smooth bromegrass seeds. The germination characteristics of the seeds revealed that the variations in nitrogen application and grain positions significantly influenced seeds vigor. The seed vigor of increased gradually with their maturity, reaching a high level at 30 and 36 days after anthesis. A stacking ensemble learning approach was employed to identify the seed maturity based on multispectral imaging and autofluorescence imaging. The results demonstrated that the Ensemble model outperformed Support Vector Machine, Bayesian, XGBoost and Random Forest across all evaluated metrics in different scenarios. The model accuracy in CK, N1 and N2 were 89%, 87% and 93%, respectively. Furthermore, the SHapley Additive exPlanations method was selected to interpret the Ensemble model, identifying important features such as 405, 430, 540, 630, 645, 690, 850, 880 and 970 nm. These features exhibited a significant correlation with fresh weight, shoot length and vigor index. These findings showed the high accuracy and generalizability of the Ensemble model for identifying the maturity and quality of smooth bromegrass seeds. Therefore, a new strategy would be offered for evaluating seed maturity and vigor level. Supplementary Information The online version contains supplementary material available at 10.1186/s13007-025-01359-8.

Background The high fat content in oat seeds makes them susceptible to aging during storage, leading to reduced seed vigor, delayed germination, and even seed death. Much evidence suggests that lipid remodeling is closely associated with successful seed germination. However, the dynamic behavior and response mechanisms of lipids during the germination of aged oat seeds remain unclear. In this study, ‘Monida’ (aging-tolerant) and ‘Haywire’ (aging-sensitive), were used to investigate the lipid profiles in the embryo and endosperm and the dynamic transcriptomic differences in the embryo during the germination. Results The results demonstrate that phospholipid alterations during the germination of aged seeds are more significant compared to unaged seeds, indicating that aging affects lipid remodeling during germination, particularly in the ‘Haywire’. Further analysis revealed that the most critical lipid response events occurred at the end of germination stage II (32 h) in embryo, primarily regulated through the PLC-DGK pathway to modulate phospholipid and glycerolipid molecules. Specifically, transcripts of PLC, DGK, and DGAT were upregulated, promoting the generation of diacylglycerol (DG) from various phospholipids, which further increased the monogalactosyldiacylglycerol/digalactosyldiacylglycerol (MGDG/DGDG) ratio, thereby influencing membrane repair. Additionally, at 6 h of germination in aged seeds, PC(3:0/0:0) levels significantly decreased. Compared to ‘Monida,’ the aging-sensitive ‘Haywire’ seeds exhibited substantial production of PE(19:0/0:0) and PC(15:0/0:0) at 32 h of germination, which may be key factors contributing to the seed's sensitivity to aging and the significant reduction in germination percentage after aging. Therefore, PC(3:0/0:0), PE(19:0/0:0), and PC(15:0/0:0) could serve as important lipid metabolic markers in future studies on the mechanisms of oat seed vigor. Conclusions The findings of this study provide insights into the specificity of lipid remodeling and its response mechanisms during the germination of aged oat seeds, providing a theoretical foundation for the safe preservation of oat germplasm and the development of aging-tolerant varieties.

Soil salinization poses an increasingly severe threat to sustainable agriculture globally, 13 while conventional seed priming optimization remains time-and resource-intensive, 14 limiting its practical implementation. This study developed an innovative smart seed 15 priming framework that seamlessly combined multispectral imaging technology with 16 explainable machine learning algorithms to optimize alfalfa seed priming under 17 saline-alkali stress (75 mM Na₂SO₄). Through systematic evaluation of six distinct 18 priming agents (gamma-aminobutyric acid (GABA), ascorbic acid (AsA), melatonin 19 (MT), salicylic acid (SA), spermidine (Spd), and sodium sulfate (Na₂SO₄)), our 20 sophisticated stacking ensemble model demonstrated exceptional performance in both 21 priming parameter classification (accuracy: 0.812-0.920, ROC-AUC: 0.941-0.994) 22 and effect prediction (accuracy: 0.820-0.950, ROC-AUC: 0.928-0.960). 23 Comprehensive SHapley Additive exPlanations (SHAP) analysis identified critical 24 spectral features across multiple analytical scales: the parameter model highlighted 25 wavelengths at 570 nm and 970 nm as key indicators of moisture content and 26 flavonoid compositional changes; for priming effects assessment, wavelengths at 850 27 nm (associated with storage materials) and 490 nm exhibited significant negative 28 synergistic effects specifically on MT priming "first" category; detailed individual 29 seed analysis revealed complex nonlinear relationships between priming agent 30 concentration, treatment duration, and corresponding spectral variations. The 31 developed smart seed priming framework enabled rapid and efficient non-destructive 32 optimization of seed priming, significantly accelerating priming technology 33 development processes and advancing sustainable agricultural practices by providing 34 personalized priming solutions under various stress conditions, particularly in regions 35 affected by soil salinization. 36

Abstract： Smooth bromegrass (Bromus inermis) was adopted as experiment materials for identifying the seed maturity using a combination of multispectral imaging and machine learning. The trials were conducted to investigate the effects of three nitrogen application levels (0, 100 and 200 kg N ha-1, defined as CK, N1 and N2 respectively) and two spikelet grain positions (spikelet basal position for SG and spikelet upper position for IG) on smooth bromegrass seeds. The germination characteristics of the seeds revealed that the variations in nitrogen application and grain positions significantly influenced seeds vigor. The seed vigor of increased gradually with their maturity, reaching a high level at 30 and 36 days after anthesis. A stacking ensemble learning approach was employed to identify the seed maturity based on multispectral imaging and autofluorescence imaging. The results demonstrated that the Ensemble model outperformed Support Vector Machine, Bayesian, XGBoost and Random Forest across all evaluated metrics in different scenarios. The model accuracy in CK, N1 and N2 were 89%, 87% and 93%, respectively. Furthermore, the SHapley Additive exPlanations method was selected to interpret the Ensemble model, identifying important features such as 405, 430, 540, 630, 645, 690, 850, 880 and 970 nm. These features exhibited a significant correlation with fresh weight, shoot length and vigor index. These findings showed the high accuracy and generalizability of the Ensemble model for identifying the maturity and quality of smooth bromegrass seeds. Therefore, a new strategy would be offered for evaluating seed maturity and vigor level. Key words: smooth bromegrass; nitrogen; seed maturity; multispectral imaging; stacking ensemble

Seed aging poses a significant challenge to agronomic production and germplasm conservation. Reactive oxygen species (ROS) are highly involved in the aging process. However, dynamic response of germination characteristics and antioxidant system to seed aging are not yet very clear. This study explored the potential physiological mechanisms responsible for the reduced and rapid loss of seed vigor in alfalfa, and identified key genes regulating seed vigor. The germination percentage exhibited a decreased trend with the prolongation of aging duration. From 16 to 32 days of aging, the antioxidant enzyme activities of SOD, POD, CAT, DHAR and MDHAR declined significantly, which lead to the disruption of ROS balance and a significant increase in ROS levels, exacerbating seed aging. Based on transcriptome, 29 differentially expressed genes (DEGs) including SOD1, APX-2 and GST-7 within the ROS scavenging system showed a significantly down-regulated expression trend at aging of 16 and 24 days, indicating the abnormal function of antioxidant metabolism. Furthermore, some related genes including ATPF1B, ATPeF0C-3, NDUFS1, NDUFS3 and ND2 in the mitochondrial ETC exhibited a downturn following seed aging, which would result in the losing of seed vigor. This study has uncovered a significant array of potential target genes within the seed antioxidant system and mitochondrial ETC. These discoveries offer a wider lens for delving into the molecular regulatory mechanisms of seed aging. Further research is crucial to comprehensively elucidate the precise pathways through which these pivotal genes regulate seed vigor.