Metabolome and transcriptome analyses reveal changes of rapeseed in response to ABA signal during early seedling development.

Seed germination is an important development process in plant growth. The phytohormone abscisic acid (ABA) plays a critical role during seed germination. However, the mechanism of rapeseed in response to ABA is still elusive. In order to understand changes of rapeseed under exogenous ABA treatment, we explored differentially expressed metabolites (DEMs) and the differentially expressed genes (DEGs) between mock- and ABA-treated seedlings. A widely targeted LC-MS/MS based metabolomics were used to identify and quantify metabolic changes in response to ABA during seed germination, and a total of 186 significantly DEMs were identified. There are many compounds which are involved in ABA stimuli, especially some specific ABA transportation-related metabolites such as starches and lipids were screened out. Meanwhile, a total of 4440 significantly DEGs were identified by transcriptomic analyses. There was a significant enrichment of DEGs related to phenylpropanoid and cell wall organization. It suggests that exogenous ABA mainly affects seed germination by regulating cell wall loosening. Finally, the correlation analysis of the key DEMs and DEGs indicates that many DEGs play a direct or indirect regulatory role in DEMs metabolism. The integrative analysis between DEGs and DEMs suggests that the starch and sucrose pathways were the key pathway in ABA responses. The two metabolites from starch and sucrose pathways, levan and cellobiose, both were found significantly down-regulated in ABA-treated seedlings. These comprehensive metabolic and transcript analyses provide useful information for the subsequent post-transcriptional modification and post germination growth of rapeseed in response to ABA signals and stresses.

Functional analysis of a wheat class III peroxidase gene, TaPer12-3A, in seed dormancy and germination.

BACKGROUND: Class III peroxidases (PODs) perform crucial functions in various developmental processes and responses to biotic and abiotic stresses. However, their roles in wheat seed dormancy (SD) and germination remain elusive. RESULTS: Here, we identified a wheat class III POD gene, named TaPer12-3A, based on transcriptome data and expression analysis. TaPer12-3A showed decreasing and increasing expression trends with SD acquisition and release, respectively. It was highly expressed in wheat seeds and localized in the endoplasmic reticulum and cytoplasm. Germination tests were performed using the transgenic Arabidopsis and rice lines as well as wheat mutant mutagenized with ethyl methane sulfonate (EMS) in Jing 411 (J411) background. These results indicated that TaPer12-3A negatively regulated SD and positively mediated germination. Further studies showed that TaPer12-3A maintained H2O2 homeostasis by scavenging excess H2O2 and participated in the biosynthesis and catabolism pathways of gibberellic acid and abscisic acid to regulate SD and germination. CONCLUSION: These findings not only provide new insights for future functional analysis of TaPer12-3A in regulating wheat SD and germination but also provide a target gene for breeding wheat varieties with high pre-harvest sprouting resistance by gene editing technology.

WOX2 functions redundantly with WOX1 and WOX4 to positively regulate seed germination in Arabidopsis.

MAIN CONCLUSION: WOX family gene WOX2 is highly expressed during seed development, which functions redundantly with WOX1 and WOX4 to positively regulate seed germination. WOX (WUSCHEL-related homeobox) is a family of transcription factors in plants. They play essential roles in the regulation of plant growth and development, but their function in seed germination is not well understood. In this report, we show that WOX1, WOX2, and WOX4 are close homologues in Arabidopsis. WOX2 has a redundant function with WOX1 and WOX4, respectively, in seed germination. WOX2 is highly expressed during seed development, from the globular embryonic stage to mature dry seeds, and its expression is decreased after germination. Loss of function single mutant wox2, and double mutants wox1 wox2 and wox2 wox4-1 show decreased germination speed. WOX2 and WOX4 are essential for hypocotyl-radicle zone elongation during germination, potentially by promoting the expression of cell wall-related genes. We also found that WOX2 and WOX4 regulate germination through the gibberellin (GA) pathway. These results suggest that WOX2 and WOX4 integrate the GA pathway and downstream cell wall-related genes during germination.

Analysis of the global transcriptome and miRNAome associated with seed dormancy during seed maturation in rice (Oryza sativa L. cv. Nipponbare).

BACKGROUND: Seed dormancy is a biological mechanism that prevents germination until favorable conditions for the subsequent generation of plants are encountered. Therefore, this mechanism must be effectively established during seed maturation. Studies investigating the transcriptome and miRNAome of rice embryos and endosperms at various maturation stages to evaluate seed dormancy are limited. This study aimed to compare the transcriptome and miRNAome of rice seeds during seed maturation. RESULTS: Oryza sativa L. cv. Nipponbare seeds were sampled for embryos and endosperms at three maturation stages: 30, 45, and 60 days after heading (DAH). The pre-harvest sprouting (PHS) assay was conducted to assess the level of dormancy in the seeds at each maturation stage. At 60 DAH, the PHS rate was significantly increased compared to those at 30 and 45 DAH, indicating that the dormancy is broken during the later maturation stage (45 DAH to 60 DAH). However, the largest number of differentially expressed genes (DEGs) and differentially expressed miRNAs (DEmiRs) were identified between 30 and 60 DAH in the embryo and endosperm, implying that the gradual changes in genes and miRNAs from 30 to 60 DAH may play a significant role in breaking seed dormancy. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses confirmed that DEGs related to plant hormones were most abundant in the embryo during 45 DAH to 60 DAH and 30 DAH to 60 DAH transitions. Alternatively, most of the DEGs in the endosperm were related to energy and abiotic stress. MapMan analysis and quantitative real-time polymerase chain reaction identified four newly profiled auxin-related genes (OsSAUR6/12/23/25) and one ethylene-related gene (OsERF087), which may be involved in seed dormancy during maturation. Additionally, miRNA target prediction (psRNATarget) and degradome dataset (TarDB) indicated a potential association between osa-miR531b and ethylene biosynthesis gene (OsACO4), along with osa-miR390-5p and the abscisic acid (ABA) exporter-related gene (OsMATE19) as factors involved in seed dormancy. CONCLUSIONS: Analysis of the transcriptome and miRNAome of rice embryos and endosperms during seed maturation provided new insights into seed dormancy, particularly its relationship with plant hormones such as ABA, auxin, and ethylene.

A transposon insertion in the promoter of OsUBC12 enhances cold tolerance during japonica rice germination.

Low-temperature germination (LTG) is an important agronomic trait for rice (Oryza sativa). Japonica rice generally has greater capacity for germination at low temperatures than the indica subpopulation. However, the genetic basis and molecular mechanisms underlying this complex trait are poorly understood. Here, we report that OsUBC12, encoding an E2 ubiquitin-conjugating enzyme, increases low-temperature germinability in japonica, owing to a transposon insertion in its promoter enhancing its expression. Natural variation analysis reveals that transposon insertion in the OsUBC12 promoter mainly occurs in the japonica lineage. The variation detected in eight representative two-line male sterile lines suggests the existence of this allele introgression by indica-japonica hybridization breeding, and varieties carrying the japonica OsUBC12 locus (transposon insertion) have higher low-temperature germinability than varieties without the locus. Further molecular analysis shows that OsUBC12 negatively regulate ABA signaling. OsUBC12-regulated seed germination and ABA signaling mainly depend on a conserved active site required for ubiquitin-conjugating enzyme activity. Furthermore, OsUBC12 directly associates with rice SUCROSE NON-FERMENTING 1-RELATED PROTEIN KINASE 1.1 (OsSnRK1.1), promoting its degradation. OsSnRK1.1 inhibits LTG by enhancing ABA signaling and acts downstream of OsUBC12. These findings shed light on the underlying mechanisms of UBC12 regulating LTG and provide genetic reference points for improving LTG in indica rice.

Morpho-Physiochemical Indices and Transcriptome Analysis Reveal the Role of Glucosinolate and Erucic Acid in Response to Drought Stress during Seed Germination of Rapeseed.

The global expansion of rapeseed seed quality has been focused on maintaining glucosinolate (GSL) and erucic acid (EA) contents. However, the influence of seed GSL and EA contents on the germination process under drought stress remains poorly understood. Herein, 114 rapeseed accessions were divided into four groups based on GSL and EA contents to investigate their performance during seed imbibition under drought stress. Our results revealed significant variations in seed germination-related traits, particularly with higher GSL and EA, which exhibited higher germination % (G%) and lower mean germination time (MGT) under drought stress conditions. Moreover, osmoregulation, enzymatic system and hormonal regulation were improved in high GSL and high EA (HGHE) versus low GSL and low EA (LGLE) seeds, indicating the essential protective role of GSL and EA during the germination process in response to drought stress. The transcriptional regulation mechanism for coordinating GSL-EA-related pathways in response to drought stress during seed imbibition was found to involve the differential expression of sugar metabolism-, antioxidant-, and hormone-related genes with higher enrichment in HGHE compared to LGLE seeds. GO enrichment analysis showed higher variations in transcription regulator activity and DNA-binding transcription factors, as well as ATP and microtubule motor activity in GSL-EA-related pathways. Furthermore, KEGG analysis identified cellular processes, environmental information processing, and metabolism categories, with varied gene participation between GSL, EA and GSL-EA-related pathways. For further clarification, QY7 (LGLE) seeds were primed with different concentrations of GSL and EA under drought stress conditions. The results showed that 200 µmol/L of GSL and 400 µmol/L of EA significantly improved G%, MGT, and seedling fresh weight, besides regulating stress and fatty acid responsive genes during the seed germination process under drought stress conditions. Conclusively, exogenous application of GSL and EA is considered a promising method for enhancing the drought tolerance of LGLE seeds. Furthermore, the current investigation could provide a theoretical basis of GSL and EA roles and their underlying mechanisms in stress tolerance during the germination process.

Overexpression of the alfalfa (Medicago sativa) gene, MsKMS1, negatively regulates seed germination in transgenic tobacco (Nicotiana tabacum).

Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)-associated proteins are a class of transmembrane proteins involved in intracellular trafficking pathways. However, the functions of many SNARE domain-containing proteins remain unclear. We have previously identified a SNARE-associated gene in alfalfa (Medicago sativa ) KILLING ME SLOWLY1 (MsKMS1 ), which is involved in various abiotic stresses. In this study, we investigated the function of MsKMS1 in the seed germination of transgenic tobacco (Nicotiana tabacum ). Phylogenetic analysis showed that MsKMS1 was homologous to the SNARE-associated or MAPR component-related proteins of other plants. Germination assays revealed that MsKMS1 negatively regulated seed germination under normal, D-mannitol and abscisic acid-induced stress conditions, yet MsKMS1 -overexpression could confer enhanced heat tolerance in transgenic tobacco. The suppressive effect on germination in MsKMS1 -overexpression lines was associated with higher abscisic acid and salicylic acid contents in seeds. This was accompanied by the upregulation of abscisic acid biosynthetic genes (ZEP and NCED ) and the downregulation of gibberellin biosynthetic genes (GA20ox2 and GA20ox3 ). Taken together, these results suggested that MsKMS1 negatively regulated seed germination by increasing abscisic acid and salicylic acid contents through the expression of genes related to abscisic acid and gibberellin biosynthesis. In addition, MsKMS1 could improve heat tolerance during the germination of transgenic tobacco seeds.

The GARP family transcription factor MtHHO3 negatively regulates salt tolerance in Medicago truncatula.

High salinity is one of the detrimental environmental factors restricting plant growth and crop production throughout the world. This study demonstrated that the GARP family transcription factor MtHHO3 is involved in response to salt stress and abscisic acid (ABA) signaling in Medicago truncatula. The transcription of MtHHO3 was repressed by salt, osmotic stress, and ABA treatment. The seed germination assay showed that, overexpression of MtHHO3 in Arabidopsis thaliana caused hypersensitivity to salt and osmotic stress, but increased resistance to ABA inhibition. Overexpression of MtHHO3 in M. truncatula resulted in decreased tolerance of salinity, while loss-of-function mutants mthho3-1 and mthho3-2 were more resistant to salt stress compared with wild-type plants. qRT-PCR analyses showed that MtHHO3 downregulated the expression of genes in stress and ABA responsive pathways. We further demonstrated that MtHHO3 repressed the transcription of the pathogenesis-related gene MtPR2 by binding to its promoter. Overall, these results indicate that MtHHO3 negatively regulates salt stress response in plants and deepen our understanding of the role of the GARP subfamily transcription factors in modulating salt stress and ABA signaling.

Insights into drought stress response mechanism of tobacco during seed germination by integrated analysis of transcriptome and metabolome.

Drought stress inhibits seed germination, plant growth and development of tobacco, and seriously affects the yield and quality of tobacco leaves. However, the molecular mechanism underlying tobacco drought stress response remains largely unknown. In this study, integrated analysis of transcriptome and metabolome was performed on the germinated seeds of a cultivated variety K326 and its EMS mutagenic mutant M28 with great drought tolerance. The result showed that drought stress inhibited seed germination of the both varieties, while the germination rate of M28 was faster than that of K326 under drought stress. Besides, the levels of phytohormone ABA, GA19, and zeatin were increased by drought stress in M28. Five vital pathways were identified through integrated transcriptomic and metabolomic analysis, including zeatin biosynthesis, aspartate and glutamate synthesis, phenylamine metabolism, glutathione metabolism, and phenylpropanoid synthesis. Furthermore, 20 key metabolites in the above pathways were selected for further analysis of gene modular-trait relationship, and then four highly correlated modules were found. Then analysis of gene expression network was carried out of Top30 hub gene of these four modules, and 9 key candidate genes were identified, including HSP70s, XTH16s, APX, PHI-1, 14-3-3, SCP, PPO. In conclusion, our study uncovered some key drought-responsive pathways and genes of tobacco during seeds germination, providing new insights into the regulatory mechanisms of tobacco drought stress response.

Exploring water-absorbing capacity: a digital image analysis of seeds from 120 wheat varieties.

Wheat is a staple food crop that provides a significant portion of the world's daily caloric intake, serving as a vital source of carbohydrates and dietary fiber for billions of people. Seed shape studies of wheat typically involve the use of digital image analysis software to quantify various seed shape parameters such as length, width, area, aspect ratio, roundness, and symmetry. This study presents a comprehensive investigation into the water-absorbing capacity of seeds from 120 distinct wheat lines, leveraging digital image analysis techniques facilitated by SmartGrain software. Water absorption is a pivotal process in the early stages of seed germination, directly influencing plant growth and crop yield. SmartGrain, a powerful image analysis tool, was employed to extract precise quantitative data from digital images of wheat seeds, enabling the assessment of various seed traits in relation to their water-absorbing capacity. The analysis revealed significant transformations in seed characteristics as they absorbed water, including changes in size, weight, shape, and more. Through statistical analysis and correlation assessments, we identified robust relationships between these seed traits, both before and after water treatment. Principal Component Analysis (PCA) and Agglomerative Hierarchical Clustering (AHC) were employed to categorize genotypes with similar trait patterns, providing insights valuable for crop breeding and genetic research. Multiple linear regression analysis further elucidated the influence of specific seed traits, such as weight, width, and distance, on water-absorbing capacity. Our study contributes to a deeper understanding of seed development, imbibition, and the crucial role of water absorption in wheat. These insights have practical implications in agriculture, offering opportunities to optimize breeding programs for improved water absorption in wheat genotypes. The integration of SmartGrain software with advanced statistical methods enhances the reliability and significance of our findings, paving the way for more efficient and resilient wheat crop production. Significant changes in wheat seed shape parameters were observed after imbibition, with notable increases in area, perimeter, length, width, and weight. The length-to-width ratio (LWR) and circularity displayed opposite trends, with higher values before imbibition and lower values after imbibition.

Metabolic control of seed germination in legumes.

Seed development, dormancy, and germination are connected with changes in metabolite levels. Not surprisingly, a complex regulatory network modulates biosynthesis and accumulation of storage products. Seed development has been studied profusely in Arabidopsis thaliana and has provided valuable insights into the genetic control of embryo development. However, not every inference applies to crop legumes, as these have been domesticated and selected for high seed yield and specific metabolic profiles and fluxes. Given its enormous economic relevance, considerable work has contributed to shed light on the mechanisms that control legume seed growth and germination. Here, we summarize recent progress in the understanding of regulatory networks that coordinate seed metabolism and development in legumes.

Redundant role of OsCNGC4 and OsCNGC5 encoding cyclic nucleotide-gated channels in rice pollen germination and tube growth.

In staple crops, such as rice (Oryza sativa L.), pollen plays a crucial role in seed production. However, the molecular mechanisms underlying rice pollen germination and tube growth remain underexplored. Notably, we recently uncovered the redundant expression and mutual interaction of two rice genes encoding cyclic nucleotide-gated channels (CNGCs), OsCNGC4 and OsCNGC5, in mature pollen. Building on these findings, the current study focused on clarifying the functional roles of these two genes in pollen germination and tube growth. To overcome functional redundancy, we produced gene-edited rice plants with mutations in both genes using the CRISPR-Cas9 system. The resulting homozygous OsCNGC4 and OsCNGC5 gene-edited mutants (oscngc4/5) exhibited significantly lower pollen germination rates than the wild type (WT), along with severely reduced fertility. Transcriptome analysis of the double oscngc4/5 mutant revealed downregulation of genes related to receptor kinases, transporters, and cell wall metabolism. To identify the direct regulators of OsCNGC4, which form a heterodimer with OsCNGC5, we screened a yeast two-hybrid library containing rice cDNAs from mature anthers. Subsequently, we identified two calmodulin isoforms (CaM1-1 and CaM1-2), NETWORKED 2 A (NET2A), and proline-rich extension-like receptor kinase 13 (PERK13) proteins as interactors of OsCNGC4, suggesting its roles in regulating Ca2+ channel activity and F-actin organization. Overall, our results suggest that OsCNGC4 and OsCNGC5 may play critical roles in pollen germination and elongation by regulating the Ca2+ gradient in growing pollen tubes.

RGA1 regulates grain size, rice quality and seed germination in the small and round grain mutant srg5.

BACKGROUND: Generating elite rice varieties with high yield and superior quality is the main goal of rice breeding programs. Key agronomic traits, including grain size and seed germination characteristics, affect the final yield and quality of rice. The RGA1 gene, which encodes the α-subunit of rice G-protein, plays an important role in regulating rice architecture, seed size and abiotic stress responses. However, whether RGA1 is involved in the regulation of rice quality and seed germination traits is still unclear. RESULTS: In this study, a rice mutant small and round grain 5 (srg5), was identified in an EMS-induced rice mutant library. Systematic analysis of its major agronomic traits revealed that the srg5 mutant exhibited a semi-dwarf plant height with small and round grain and reduced panicle length. Analysis of the physicochemical properties of rice showed that the difference in rice eating and cooking quality (ECQ) between the srg5 mutant and its wild-type control was small, but the appearance quality was significantly improved. Interestingly, a significant suppression of rice seed germination and shoot growth was observed in the srg5 mutant, which was mainly related to the regulation of ABA metabolism. RGA1 was identified as the candidate gene for the srg5 mutant by BSA analysis. A SNP at the splice site of the first intron disrupted the normal splicing of the RGA1 transcript precursor, resulting in a premature stop codon. Additional linkage analysis confirmed that the target gene causing the srg5 mutant phenotype was RGA1. Finally, the introduction of the RGA1 mutant allele into two indica rice varieties also resulted in small and round rice grains with less chalkiness. CONCLUSIONS: These results indicate that RGA1 is not only involved in the control of rice architecture and grain size, but also in the regulation of rice quality and seed germination. This study sheds new light on the biological functions of RGA1, thereby providing valuable information for future systematic analysis of the G-protein pathway and its potential application in rice breeding programs.

Physiology and transcriptome of Eucommia ulmoides seeds at different germination stages.

E. ulmoides (Eucommia ulmoides) has significant industrial and medicinal value and high market demand. E. ulmoides grows seedlings through sowing. According to previous studies, plant hormones have been shown to regulate seed germination. To understand the relationship between hormones and E. ulmoides seed germination, we focused on examining the changes in various indicators during the germination stage of E. ulmoides seeds. We measured the levels of physiological and hormone indicators in E. ulmoides seeds at different germination stages and found that the levels of abscisic acid (ABA), gibberellin (GA), and indole acetic acid (IAA) significantly varied as the seeds germinated. Furthermore, we confirmed that ABA, GA, and IAA are essential hormones in the germination of E. ulmoides seeds using Gene Ontology and Kyoto Encyclopedia of Genes and Genomics enrichment analyses of the transcriptome. The discovery of hormone-related synthesis pathways in the control group of Eucommia seeds at different germination stages further confirmed this conclusion. This study provides a basis for further research into the regulatory mechanisms of E. ulmoides seeds at different germination stages and the relationship between other seed germination and plant hormones.

Interplay between coding and non-coding regulation drives the Arabidopsis seed-to-seedling transition.

Translation of seed stored mRNAs is essential to trigger germination. However, when RNAPII re-engages RNA synthesis during the seed-to-seedling transition has remained in question. Combining csRNA-seq, ATAC-seq and smFISH in Arabidopsis thaliana we demonstrate that active transcription initiation is detectable during the entire germination process. Features of non-coding regulation such as dynamic changes in chromatin accessible regions, antisense transcription, as well as bidirectional non-coding promoters are widespread throughout the Arabidopsis genome. We show that sensitivity to exogenous ABSCISIC ACID (ABA) during germination depends on proximal promoter accessibility at ABA-responsive genes. Moreover, we provide genetic validation of the existence of divergent transcription in plants. Our results reveal that active enhancer elements are transcribed producing non-coding enhancer RNAs (eRNAs) as widely documented in metazoans. In sum, this study defining the extent and role of coding and non-coding transcription during key stages of germination expands our understanding of transcriptional mechanisms underlying plant developmental transitions.

Seed longevity and genome damage.

Seeds are the mode of propagation for most plant species and form the basis of both agriculture and ecosystems. Desiccation tolerant seeds, representative of most crop species, can survive maturation drying to become metabolically quiescent. The desiccated state prolongs embryo viability and provides protection from adverse environmental conditions, including seasonal periods of drought and freezing often encountered in temperate regions. However, the capacity of the seed to germinate declines over time and culminates in the loss of seed viability. The relationship between environmental conditions (temperature and humidity) and the rate of seed deterioration (ageing) is well defined, but less is known about the biochemical and genetic factors that determine seed longevity. This review will highlight recent advances in our knowledge that provide insight into the cellular stresses and protective mechanisms that promote seed survival, with a focus on the roles of DNA repair and response mechanisms. Collectively, these pathways function to maintain the germination potential of seeds. Understanding the molecular basis of seed longevity provides important new genetic targets for the production of crops with enhanced resilience to changing climates and knowledge important for the preservation of plant germplasm in seedbanks.

ABA functions in low phosphate-induced anthocyanin accumulation through the transcription factor ABI5 in Arabidopsis.

KEY MESSAGE: ABI5 functions in ABA-mediated anthocyanin accumulation in plant response to low phosphate. Low phosphate (LP)-induced anthocyanin biosynthesis and accumulation play an important role in plant adaptive response to phosphate starvation conditions. However, whether and how the stress phytohormone abscisic acid (ABA) participates in LP-induced anthocyanin accumulation remain elusive. Here, we report that ABA is required for LP-induced anthocyanin accumulation in Arabidopsis thaliana. Disrupting ABA DEFICIENT2 (ABA2), a key ABA-biosynthetic gene, or BETA-GLUCOSIDASE1 (BG1), a major gene implicated in converting conjugated ABA to active ABA, significantly impairs LP-induced anthocyanin accumulation, as LP-induced expression of the anthocyanin-biosynthetic genes Chalcone Synthase (CHS) is dampened in the aba2 and bg1 mutant. In addition, LP-induced anthocyanin accumulation is defective in the mutants of ABA signaling pathway, including ABA receptors, ABA Insensitive2, and the transcription factors ABA Insensitive5 (ABI5), suggesting a role of ABI5 in ABA-mediated upregulation of anthocyanin-biosynthetic genes in plant response to LP. Indeed, LP-induced expression of CHS is repressed in the abi5-7 mutant but further promoted in the ABI5-overexpressing plants compared to the wild-type. Moreover, ABI5 can bind to and transcriptionally activate CHS, and the defectiveness of LP-induced anthocyanin accumulation in abi5-7 can be restored by overexpressing CHS. Collectively, our findings illustrates that ABI5 functions in ABA-mediated LP-induced anthocyanin accumulation in Arabidopsis.

Integrated transcriptomic and metabolomic analyses elucidate the mechanism of flavonoid biosynthesis in the regulation of mulberry seed germination under salt stress.

Seed propagation is the main method of mulberry expansion in China, an important economic forest species. However, seed germination is the most sensitive stage to various abiotic stresses, especially salinity stress. To reveal the molecular regulatory mechanism of mulberry seed germination under salt stress, flavonoid metabolomics and transcriptomics analyses were performed on mulberry seeds germinated under 50 and 100 mmol/L NaCl stress. Analysis of the flavonoid metabolome revealed that a total of 145 differential flavonoid metabolites (DFMs) were classified into 9 groups, 40 flavonols, 32 flavones, 16 chalcones and 14 flavanones. Among them, 61.4% (89) of the DFMs accumulated continuously with increasing salt concentration, reaching the highest level at a 100 mmol/L salt concentration; these DFMs included quercetin-3-O-glucoside (isoquercitrin), kaempferol (3,5,7,4'-tetrahydroxyflavone), quercetin-7-O-glucoside, taxifolin (dihydroquercetin) and apigenin (4',5,7-trihydroxyflavone), indicating that these flavonoids may be key metabolites involved in the response to salt stress. Transcriptional analysis identified a total of 3055 differentially expressed genes (DEGs), most of which were enriched in flavonoid biosynthesis (ko00941), phenylpropanoid biosynthesis (ko00940) and biosynthesis of secondary metabolites (ko01110). Combined analysis of flavonoid metabolomic and transcriptomic data indicated that phenylalanine ammonia-lyase (PAL), 4-coumarate-CoA ligase (4CL), chalcone synthase (CHS), flavonol synthase (FLS), bifunctional dihydroflavonol 4-reductase/flavanone 4-reductase (DFR) and anthocyanidin reductase (ANR) were the key genes involved in flavonoid accumulation during mulberry seed germination under 50 and 100 mmol/L NaCl stress. In addition, three transcription factors, MYB, bHLH and NAC, were involved in the regulation of flavonoid accumulation under salt stress. The results of quantitative real-time PCR (qRT‒PCR) validation showed that the expression levels of 11 DEGs, including 7 genes involved in flavonoid biosynthesis, under different salt concentrations were consistent with the transcriptomic data, and parallel reaction monitoring (PRM) results showed that the expression levels of 6 key enzymes (proteins) involved in flavonoid synthesis were consistent with the accumulation of flavonoids. This study provides a new perspective for investigating the regulatory role of flavonoid biosynthesis in the regulation of mulberry seed germination under salt stress at different concentrations.

Physiological and molecular changes of onion (Allium cepa L.) seeds under different aging conditions.

BACKGROUND: Onion seeds have limited storage capacity compared to other vegetable seeds. It is crucial to identify the mechanisms that induce tolerance to storage conditions and reduce seed deterioration. To address this goal, an experiment was conducted to evaluate changes in germination, biochemical, physiological, and molecular characteristics of onion seed landraces (Horand, Kazerun landraces and Zargan cultivar) at different aging levels (control, three-days and six-days accelerated aging, and natural aging for one year). RESULTS: The findings suggest that there was an increase in glucose, fructose, total sugar, and electrolyte leakage in the Horand (HOR), Kazerun (KAZ) landraces, and Zarghan (ZAR) cultivar, with Kazerun exhibiting the greatest increase. The percentage and rate of germination of Kazerun decreased by 54% and 33%, respectively, in six-day accelerated aging compared to the control, while it decreased by 12% and 14%, respectively, in Horand. Protein content decreased with increasing levels of aging, with a decrease of 26% in Kazerun landrace at six days of aging, while it was 16% in Horand landrace. The antioxidant activities of catalase, superoxide dismutase, and glutathione peroxidase decreased more intensively in Kazerun. The expression of AMY1, BMY1, CTR1, and NPR1 genes were lower in Kazerun landraces than in Horand and Zargan at different aging levels. CONCLUSIONS: The AMY1, BMY1, CTR1, and NPR1 genes play a pivotal role in onion seed germination, and their downregulation under stressful conditions has been shown to decrease germination rates. In addition, the activity of CAT, SOD, and GPx enzymes decreased by seed aging, and the amount of glucose, fructose, total sugar and electrolyte leakage increased, which ultimately led to seed deterioration. Based on the results of this experiment, it is recommended to conduct further studies into the molecular aspects involved in onion seed deterioration. More research on the genes related to this process is suggested, as well as investigating the impact of different priming treatments on the genes expression involved in the onion seed aging process.

Comparison of the genetic basis of salt tolerance at germination, seedling, and reproductive stages in an introgression line population of rice.

BACKGROUND: Salinity is a major limitation for rice farming due to climate change. Since salt stress adversely impact rice plants at germination, seedling, and reproductive stages resulting in poor crop establishment and reduced grain yield, enhancing salt tolerance at these vulnerable growth stages will enhance rice productivity in salinity prone areas. METHODS AND RESULTS: An introgression line (ILs) population from a cross between a high yielding cultivar 'Cheniere' and a salt tolerant donor 'TCCP' was evaluated to map quantitative trait loci (QTLs) for traits associated with salt tolerance at germination, seedling, and reproductive stages. Using a genotyping-by-sequencing based high density SNP linkage map, a total of 7, 16, and 30 QTLs were identified for five germination traits, seven seedling traits, and ten reproductive traits, respectively. There was overlapping of QTLs for some traits at different stages indicating the pleiotropic effects of these QTLs or clustering of linked genes. Candidate genes identified for salt tolerance were OsSDIR1 and SERF for the seedling stage, WRKY55 and OsUBC for the reproductive stage, and MYB family transcription factors for all three stages. Gene ontology analysis revealed significant GO terms related to nucleotide binding, protein binding, protein kinase activity, antiporter activity, active transmembrane transporter activity, calcium-binding protein, and F- box protein interaction domain containing protein. CONCLUSIONS: The colocalized QTLs for traits at different growth stages would be helpful to improve multiple traits simultaneously using marker-assisted selection. The salt tolerant ILs have the potential to be released as varieties or as pre-breeding lines for developing salt tolerant rice varieties.