Effect of soil additives on biogeochemistry of ultramafic soils-an experimental approach with Brassica napus L.

Ultramafic soils are characterized by low productivity due to the deficiency of macroelements and high content of Ni, Cr, and Co. Incorporation of ultramafic soils for agricultural and food production involves the use of fertilizers. Therefore, this study aims to find the soil additive that decreases the metallic elements uptake by plant using Brassica napus as an example. In this study, we evaluate the effect of manure (0.5 g N/kg of soil), humic acids (1 g of Rosahumus/1 dm3 H2O; 44% C), KNO3 (0.13 g K/kg of soil), lime (12.5 g/kg of soil), (NH4)2SO4 (0.15 g N/kg of soil), and Ca(H2PO4)2) (0.07 g P/kg of soil) on the phytoavailability of metallic elements. The effect of soil additives on metallic elements uptake by Brassica napus was studied in a pot experiment executed in triplicates. Statistical analysis was applied to compare the effects of additives in ultramafic soil on plant chemical composition relative to control unfertilized ultramafic soil (one-way ANOVA and Kruskal-Wallis test). The study shows that in almost all treatments, metallic elements content (Ni, Cr, Co, Al, Fe, Mn) is higher in roots compared to the aboveground parts of Brassica napus except for (NH4)2SO4, in which the mechanism of Mn accumulation is opposite. The main differences between the treatments are observed for the buffer properties of soil and the accumulation of specific metals by studied plants. The soils with the addition of lime and manure have the highest buffer properties in acidic conditions (4.9-fold and 2.1-fold increase relative to control soil, respectively), whereas the soil with (NH4)2SO4 has the lowest effect (0.8-fold decrease relative to control soil). Also, the addition of manure increases the biomass of aboveground parts of B. napus (3.4-fold increase) and decreases the accumulation of Ni (0.6-fold decrease) compared to plants cultivated in the control soil. On the contrary, the addition of (NH4)2SO4 noticeably increases the accumulation of Ni, Co, Mn, and Al in aboveground parts of B. napus (3.2-fold, 18.2-fold, 11.2-fold, and 1.6-fold, respectively) compared to plant grown in control soil, whereas the humic acids increase the accumulation of Cr in roots (1.6-fold increase). Therefore, this study shows that manure is a promising fertilizer in agricultural practices in ultramafic soil, whereas (NH4)2SO4 and humic acids must not be used in ultramafic areas.

Transcriptome analysis suggested that lncRNAs regulate rapeseed seedlings in responding to drought stress by coordinating the phytohormone signal transduction pathways.

The growth, yield, and seed quality of rapeseed are negatively affected by drought stress. Therefore, it is of great value to understand the molecular mechanism behind this phenomenon. In a previous study, long non-coding RNAs (lncRNAs) were found to play a key role in the response of rapeseed seedlings to drought stress. However, many questions remained unanswered. This study was the first to investigate the expression profile of lncRNAs not only under control and drought treatment, but also under the rehydration treatment. A total of 381 differentially expressed lncRNA and 10,253 differentially expressed mRNAs were identified in the comparison between drought stress and control condition. In the transition from drought stress to rehydration, 477 differentially expressed lncRNAs and 12,543 differentially expressed mRNAs were detected. After identifying the differentially expressed (DE) lncRNAs, the comprehensive lncRNAs-engaged network with the co-expressed mRNAs in leaves under control, drought and rehydration was investigated. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis of co-expressed mRNAs identified the most significant pathways related with plant hormones (expecially abscisic acid, auxin, cytokinins, and gibberellins) in the signal transduction. The genes, co-expressed with the most-enriched DE-lncRNAs, were considered as the most effective candidates in the water-loss and water-recovery processes, including protein phosphatase 2 C (PP2C), ABRE-binding factors (ABFs), and SMALL AUXIN UP-REGULATED RNAs (SAURs). In summary, these analyses clearly demonstrated that DE-lncRNAs can act as a regulatory hub in plant-water interaction by controlling phytohormone signaling pathways and provided an alternative way to explore the complex mechanisms of drought tolerance in rapeseed.

Genome-Wide Association Study Reveals the Genetic Basis of Crude Fiber Components in Brassica napus L. Shoots at Stem Elongation Stage.

Brassica napus is currently the principal field crop for producing materials for primary, secondary and tertiary industries. B. napus shoots at stem elongation stage are rich in anthocyanins, vitamin C and mineral elements such as selenium, calcium and zinc, and represent a new type of green vegetable. However, the high crude fiber (CF) content of B. napus shoots affects their taste, and few studies have focused on the quality traits of these vegetables. In this study, we investigated five traits related to the CF components, including neutral detergent fiber (NDF), acid detergent fiber (ADF), acid detergent lignin (ADL), hemicellulose (Hem) and cellulose (Cel), of B. napus shoots. Whole-genome resequencing at a depth of ∼20× was utilized to genotype an association panel of 202 diverse accessions, which resulted in the identification of 6,093,649 single nucleotide polymorphisms (SNPs) and 996,252 indels, respectively. A genome-wide association study (GWAS) was performed for the five CF-related traits based on the phenotypic data observed in four environments. A total of 1,285 significant SNPs were detected at the threshold of -log10 (p) = 5.16, and 97 significant association regions were obtained. In addition, seven candidate genes located on chromosomes A2 (one gene), A8 (three genes), A9 (two genes) and C9 (one gene) related to CF traits were identified, and ten lines containing low CF contents were selected as excellent germplasm resources for breeding. Our results contributed new insights into the genetic basis of CF traits and suggested germplasm resources for the quality improvement of B. napus shoots.

Lipid and sugar metabolism play an essential role in pollen development and male sterility: a case analysis in Brassica napus.

AIMS: The genic male sterility (GMS) system is an important strategy for generating heterosis in plants. To better understand the essential role of lipid and sugar metabolism and to identify additional candidates for pollen development and male sterility, transcriptome and metabolome analysis of a GMS line of 1205AB in B. napus was used as a case study. DATA RESOURCES GENERATED: To characterize the GMS system, the transcriptome and metabolome profiles were generated for 24 samples and 48 samples of 1205AB in B. napus, respectively. Transcriptome analysis yielded a total of 156.52 Gb of clean data and revealed the expression levels of 109,541 genes and 8,501 novel genes. In addition, a total of 1,353 metabolites were detected in the metabolomic analysis, including 784 in positive ion mode and 569 in negative ion mode. KEY RESULTS: A total of 15,635 differentially expressed genes (DEGs) and 83 differential metabolites (DMs) were identified from different comparison groups, most of which were involved in lipid and sugar metabolism. The combination of transcriptome and metabolome analysis revealed 49 orthologous GMS genes related to lipid metabolism and 46 orthologous GMS genes related to sugar metabolism, as well as 45 novel genes. UTILITY OF THE RESOURCE: The transcriptome and metabolome profiles and their analysis provide useful reference data for the future discovery of additional GMS genes and the development of more robust male sterility breeding systems for use in the production of plant hybrids.

Genome-wide analysis of TOPLESS/TOPLESS-RELATED co-repressors and functional characterization of BnaA9.TPL regulating the embryogenesis and leaf morphology in rapeseed.

TOPLESS/TOPLESS-RELATED (TPL/TPR) proteins belong to the Groucho (Gro)/Tup1 family co-repressors and act as broad co-repressors that modulate multiple phytohormone signalling pathways and various developmental processes in plant. However, TPL/TPR co-repressors so far are poorly understood in the rapeseed, one of the world-wide important oilseed crops. In this study, we comprehensively characterized eighteen TPL/TPR genes into five groups in the rapeseed genome. Members of TPL/TPR1/TPR4 and TPR2/TPR3 had close evolutionary relationship, respectively. All TPL/TPRs had similar expression patterns and encode conserved protein domain. In addition, we demonstrated that BnaA9.TPL interacted with all known plant repression domain (RD) sequences, which were distributed in non-redundant 24,238 (22.6 %) genes and significantly enriched in transcription factors in the rapeseed genome. These transcription factors were largely co-expressed with the TPL/TPR genes and involved in diverse pathway, including phytohormone signal transduction, protein kinases and circadian rhythm. Furthermore, BnaA9.TPL was revealed to regulate apical embryonic fate by interaction with Bna.IAA12 and suppression of PLETHORA1/2. BnaA9.TPL was also identified to regulate leaf morphology by interaction with Bna.AS1 (Asymmetric leaves 1) and suppression of KNOTTED-like homeobox genes and YABBY5. These data not only suggest the rapeseed TPL/TPRs play broad roles in different processes, but also provide useful information to uncover more TPL/TPR-mediated control of plant development in rapeseed.

Human Sensory, Taste Receptor, and Quantitation Studies on Kaempferol Glycosides Derived from Rapeseed/Canola Protein Isolates.

Beyond the key bitter compound kaempferol 3-O-(2‴-O-sinapoyl-ß-d-sophoroside) previously described in the literature (1), eight further bitter and astringent-tasting kaempferol glucosides (2-9) have been identified in rapeseed protein isolates (Brassica napus L.). The bitterness and astringency of these taste-active substances have been described with taste threshold concentrations ranging from 3.3 to 531.7 and 0.3 to 66.4 µmol/L, respectively, as determined by human sensory experiments. In this study, the impact of 1 and kaempferol 3-O-ß-d-glucopyranoside (8) on TAS2R-linked proton secretion by HGT-1 cells was analyzed by quantification of the intracellular proton index. mRNA levels of bitter receptors TAS2R3, 4, 5, 13, 30, 31, 39, 40, 43, 45, 46, 50 and TAS2R8 were increased after treatment with compounds 1 and 8. Using quantitative UHPLC-MS/MSMRM measurements, the concentrations of 1-9 were determined in rapeseed/canola seeds and their corresponding protein isolates. Depending on the sample material, compounds 1, 3, and 5-9 exceeded dose over threshold (DoT) factors above one for both bitterness and astringency in selected protein isolates. In addition, an increase in the key bitter compound 1 during industrial protein production (apart from enrichment) was observed, allowing the identification of the potential precursor of 1 to be kaempferol 3-O-(2‴-O-sinapoyl-ß-d-sophoroside)-7-O-ß-d-glucopyranoside (3). These results may contribute to the production of less bitter and astringent rapeseed protein isolates through the optimization of breeding and postharvest downstream processing.

Enhancing germination and growth of canola (Brassica napus L.) through hydropriming and NaCl priming.

The excessive accumulation of sodium chloride (NaCl) in soil can result in soil salinity, which poses a significant challenge to plant growth and crop production due to impaired water and nutrient uptake. On the other hand, hydropriming (WP) and low level of NaCl priming can improve the germination of seeds, chlorophyll contents, oil and seed yield in plants. That's why this study investigates the impact of hydro and different levels of NaCl (0.5, 1.0, 1.5 and 2.0%) priming, as pre-treatment techniques on canola seeds germination, growth and yield of two varieties Punjab and Faisal Canola. Results showed that, WP performed significant best for increase in germination (~ 20 and ~ 22%) and shoot length (~ 6 and ~ 10%) over non-priming (NP) in Punjab Canola and Faisal Canola respectively. A significant increase in plant height (~ 6 and ~ 7%), root length (~ 1 and ~ 7%), shoot fresh weight (~ 5 and ~ 7%), root fresh weight (~ 6 and ~ 7%) in Punjab Canola and Faisal Canola respectively. It was also observed that plants under WP and 0.5%NaCl priming were also better in production of seed yield per plant, oil contents, silique per plant, seeds per silique, and branches per plant chlorophyll contents and leaf relative water contents over NP. In conclusion, WP and 0.5%NaCl has potential to improve the germination, growth, yield and oil attributes of canola compared to non-priming, 1.0%NaCl priming, 1.5%NaCl priming and 2.0%NaCl priming.

Cold Acclimation and Deacclimation of Winter Oilseed Rape, with Special Attention Being Paid to the Role of Brassinosteroids.

Winter plants acclimate to frost mainly during the autumn months, through the process of cold acclimation. Global climate change is causing changes in weather patterns such as the occurrence of warmer periods during late autumn or in winter. An increase in temperature after cold acclimation can decrease frost tolerance, which is particularly dangerous for winter crops. The aim of this study was to investigate the role of brassinosteroids (BRs) and BR analogues as protective agents against the negative results of deacclimation. Plants were cold-acclimated (3 weeks, 4 °C) and deacclimated (1 week, 16/9 °C d/n). Deacclimation generally reversed the cold-induced changes in the level of the putative brassinosteroid receptor protein (BRI1), the expression of BR-induced COR, and the expression of SERK1, which is involved in BR signal transduction. The deacclimation-induced decrease in frost tolerance in oilseed rape could to some extent be limited by applying steroid regulators. The deacclimation in plants could be detected using non-invasive measurements such as leaf reflectance, chlorophyll a fluorescence, and gas exchange monitoring.

Salicylic Acid Is Involved in the Growth Inhibition Caused by Excessive Ammonium in Oilseed Rape (Brassica napus L.).

Rapeseed (Brassica napus L.) is extremely sensitive to excessive NH4+ toxicity. There remains incomplete knowledge of the causal factors behind the growth suppression in NH4+-nourished plants, with limited studies conducted specifically on field crop plants. In this study, we found that NH4+ toxicity significantly increased salicylic acid (SA) accumulation by accelerating the conversion of SA precursors. Moreover, exogenous SA application significantly aggravated NH4+ toxicity symptoms in the rapeseed shoots. Genome-wide differential transcriptomic analysis showed that NH4+ toxicity increased the expression of genes involved in the biosynthesis, transport, signaling transduction, and conversion of SA. SA treatment significantly increased shoot NH4+ concentrations by reducing the activities of glutamine synthase and glutamate synthase in NH4+-treated rapeseed plants. The application of an SA biosynthesis inhibitor, ABT, alleviated NH4+ toxicity symptoms. Furthermore, SA induced putrescine (Put) accumulation, resulting in an elevated ratio of Put to [spermidine (Spd) + spermine (Spm)] in the NH4+-treated plants, while the opposite was true for ABT. The application of exogenous Put and its biosynthesis inhibitor DFMA induced opposite effects on NH4+ toxicity in rapeseed shoots. These results indicated that the increased endogenous SA contributed noticeably to the toxicity caused by the sole NH4+-N supply in rapeseed shoots. This study provided fresh perspectives on the mechanism underlying excessive NH4+-induced toxicity and the corresponding alleviating strategies in plants.

Characteristics of duplicated gene expression and DNA methylation regulation in different tissues of allopolyploid Brassica napus.

Plant polyploidization increases the complexity of epigenomes and transcriptional regulation, resulting in genome evolution and enhanced adaptability. However, few studies have been conducted on the relationship between gene expression and epigenetic modification in different plant tissues after allopolyploidization. In this study, we studied gene expression and DNA methylation modification patterns in four tissues (stems, leaves, flowers and siliques) of Brassica napusand its diploid progenitors. On this basis, the alternative splicing patterns and cis-trans regulation patterns of four tissues in B. napus and its diploid progenitors were also analyzed. It can be seen that the number of alternative splicing occurs in the B. napus is higher than that in the diploid progenitors, and the IR type increases the most during allopolyploidy. In addition, we studied the fate changes of duplicated genes after allopolyploidization in B. napus. We found that the fate of most duplicated genes is conserved, but the number of neofunctionalization and specialization is also large. The genetic fate of B. napus was classified according to five replication types (WGD, PD, DSD, TD, TRD). This study also analyzed generational transmission analysis of expression and DNA methylation patterns. Our study provides a reference for the fate differentiation of duplicated genes during allopolyploidization.

Genome-Wide Identification and Multi-Stress Response Analysis of the DABB-Type Protein-Encoding Genes in Brassica napus.

The DABB proteins, which are characterized by stress-responsive dimeric A/B barrel domains, have multiple functions in plant biology. In Arabidopsis thaliana, these proteins play a crucial role in defending against various pathogenic fungi. However, the specific roles of DABB proteins in Brassica napus remain elusive. In this study, 16 DABB encoding genes were identified, distributed across 10 chromosomes of the B. napus genome, which were classified into 5 branches based on phylogenetic analysis. Genes within the same branch exhibited similar structural domains, conserved motifs, and three-dimensional structures, indicative of the conservation of BnaDABB genes (BnaDABBs). Furthermore, the enrichment of numerous cis-acting elements in hormone induction and light response were revealed in the promoters of BnaDABBs. Expression pattern analysis demonstrated the involvement of BnaDABBs, not only in the organ development of B. napus but also in response to abiotic stresses and Sclerotinia sclerotiorum infection. Altogether, these findings imply the significant impacts of BnaDABBs on plant growth and development, as well as stress responses.

Genetic control of root morphology in response to nitrogen across rapeseed diversity.

Rapeseed (Brassica napus L.) is an oil-containing crop of great economic value but with considerable nitrogen requirement. Breeding root systems that efficiently absorb nitrogen from the soil could be a driver to ensure genetic gains for more sustainable rapeseed production. The aim of this study is to identify genomic regions that regulate root morphology in response to nitrate availability. The natural variability offered by 300 inbred lines was screened at two experimental locations. Seedlings grew hydroponically with low or elevated nitrate levels. Fifteen traits related to biomass production and root morphology were measured. On average across the panel, a low nitrate level increased the root-to-shoot biomass ratio and the lateral root length. A large phenotypic variation was observed, along with important heritability values and genotypic effects, but low genotype-by-nitrogen interactions. Genome-wide association study and bulk segregant analysis were used to identify loci regulating phenotypic traits. The first approach nominated 319 SNPs that were combined into 80 QTLs. Three QTLs identified on the A07 and C07 chromosomes were stable across nitrate levels and/or experimental locations. The second approach involved genotyping two groups of individuals from an experimental F2 population created by crossing two accessions with contrasting lateral root lengths. These individuals were found in the tails of the phenotypic distribution. Co-localized QTLs found in both mapping approaches covered a chromosomal region on the A06 chromosome. The QTL regions contained some genes putatively involved in root organogenesis and represent selection targets for redesigning the root morphology of rapeseed.

The fungus Acremonium alternatum enhances salt stress tolerance by regulating host redox homeostasis and phytohormone signaling.

While endophytic fungi offer promising avenues for bolstering plant resilience against abiotic stressors, the molecular mechanisms behind this biofortification remain largely unknown. This study employed a multifaceted approach, combining plant physiology, proteomic, metabolomic, and targeted hormonal analyses to illuminate the early response of Brassica napus to Acremonium alternatum during the nascent stages of their interaction. Notably, under optimal growth conditions, the initial reaction to fungus was relatively subtle, with no visible alterations in plant phenotype and only minor impacts on the proteome and metabolome. Interestingly, the identified proteins associated with the Acremonium response included TUDOR 1, Annexin D4, and a plastidic K+ efflux antiporter, hinting at potential processes that could counter abiotic stressors, particularly salt stress. Subsequent experiments validated this hypothesis, showcasing significantly enhanced growth in Acremonium-inoculated plants under salt stress. Molecular analyses revealed a profound impact on the plant's proteome, with over 50% of salt stress response proteins remaining unaffected in inoculated plants. Acremonium modulated ribosomal proteins, increased abundance of photosynthetic proteins, enhanced ROS metabolism, accumulation of V-ATPase, altered abundances of various metabolic enzymes, and possibly promoted abscisic acid signaling. Subsequent analyses validated the accumulation of this hormone and its enhanced signaling. Collectively, these findings indicate that Acremonium promotes salt tolerance by orchestrating abscisic acid signaling, priming the plant's antioxidant system, as evidenced by the accumulation of ROS-scavenging metabolites and alterations in ROS metabolism, leading to lowered ROS levels and enhanced photosynthesis. Additionally, it modulates ion sequestration through V-ATPase accumulation, potentially contributing to the observed decrease in chloride content.

BnaPLDα1-BnaMPK6 Involved in NaCl-Mediated Overcoming of Self-Incompatibility in Brassica napus L.

Self-incompatibility (SI) is an important genetic mechanism exploited by numerous angiosperm species to prevent inbreeding. This mechanism has been widely used in the breeding of SI trilinear hybrids of Brassica napus. The SI responses in these hybrids can be overcome by using a salt (NaCl) solution, which is used for seed propagation in SI lines. However, the mechanism underlying the NaCl-induced breakdown of the SI response in B. napus remains unclear. Here, we investigated the role of two key proteins, BnaPLDα1 and BnaMPK6, in the breakdown of SI induced by NaCl. Pollen grain germination and seed set were reduced in BnaPLDα1 triple mutants following incompatible pollination with NaCl treatment. Conversely, SI responses were partially abolished by overexpression of BnaC05.PLDα1 without salt treatment. Furthermore, we observed that phosphatidic acid (PA) produced by BnaPLDα1 bound to B. napus BnaMPK6. The suppression and enhancement of the NaCl-induced breakdown of the SI response in B. napus were observed in BnaMPK6 quadruple mutants and BnaA05.MPK6 overexpression lines, respectively. Moreover, salt-induced stigmatic reactive oxygen species (ROS) accumulation had a minimal effect on the NaCl-induced breakdown of the SI response. In conclusion, our results demonstrate the essential role of the BnaPLDα1-PA-BnaMPK6 pathway in overcoming the SI response to salt treatment in SI B. napus. Additionally, our study provides new insights into the relationship between SI signaling and salt stress response. SIGNIFICANCE STATEMENT: A new molecular mechanism underlying the breakdown of the NaCl-induced self-incompatibility (SI) response in B. napus has been discovered. It involves the induction of BnaPLDα1 expression by NaCl, followed by the activation of BnaMPK6 through the production of phosphatidic acid (PA) by BnaPLDα1. Ultimately, this pathway leads to the breakdown of SI. The involvement of the BnaPLDα1-PA-BnaMPK6 pathway in overcoming the SI response following NaCl treatment provides new insights into the relationship between SI signalling and the response to salt stress.

Rapeseed PP2C37 Interacts with PYR/PYL Abscisic Acid Receptors and Negatively Regulates Drought Tolerance.

Global water deficit is a severe abiotic stress threatening the yielding and quality of crops. Abscisic acid (ABA) is a phytohormone that mediates drought tolerance. Protein kinases and phosphatases function as molecular switches in eukaryotes. Protein phosphatases type 2C (PP2Cs) are a major family that play essential roles in ABA signaling and stress responses. However, the role and underlying mechanism of PP2C in rapeseed (Brassica napus L.) mediating drought response has not been reported yet. Here, we characterized a PP2C family member, BnaPP2C37, and its expression level was highly induced by ABA and dehydration treatments. It negatively regulates drought tolerance in rapeseed. We further identified that BnaPP2C37 interacted with multiple PYR/PYL receptors and a drought regulator BnaCPK5 (calcium-dependent protein kinase 5) through yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays. Specifically, BnaPYL1 and BnaPYL9 repress BnaPP2C37 phosphatase activity. Moreover, the pull-down assay and phosphatase assays show BnaPP2C37 interacts with BnaCPK5 to dephosphorylate BnaCPK5 and its downstream BnaABF3. Furthermore, a dual-luciferase assay revealed BnaPP2C37 transcript level was enhanced by BnaABF3 and BnaABF4, forming a negative feedback regulation to ABA response. In summary, we identified that BnaPP2C37 functions negatively in drought tolerance of rapeseed, and its phosphatase activity is repressed by BnaPYL1/9 whereas its transcriptional level is upregulated by BnaABF3/4.

Functional and regulatory diversity of homeobox-leucine zipper transcription factors BnaHB6 under dehydration and salt stress in Brassica napus L.

The plant-specific homeodomain-leucine zipper I subfamily is involved in the regulation of various biological processes, particularly growth, development and stress response. In the present study, we characterized four BnaHB6 homologues from Brassica napus. All BnaHB6 proteins have transcriptional activation activity. Structural and functional data indicate the complex role of BnaHB6 genes in regulating biological processes, with some functions conserved and others diverged. Transcriptional analyzes revealed that they are induced in a similar manner in different tissues but show different expression patterns in response to stress and circadian rhythm. Only the BnaA09HB6 and BnaC08HB6 genes are expressed under dehydration and salt stress, and in darkness. The partial transcriptional overlap of BnaHB6s with the evolutionarily related genes BnaHB5 and BnaHB16 was also observed. Transgenic Arabidopsis thaliana plants expressing a single proBnaHB6::GUS partially confirmed the expression results. Bioinformatic analysis allowed the identification of TF-binding sites in the BnaHB6 promoters that may control their expression under stress and circadian rhythm. ChIP-qPCR analysis revealed that BnaA09HB6 and BnaC08HB6 bind directly to the promoters of the target genes BnaABF4 and BnaDREB2A. Comparison of their expression patterns in the WT plants and the bnac08hb6 mutant showed that BnaC08HB6 positively regulates the expression of the BnaABF4 and BnaDREB2A genes under dehydration and salt stress. We conclude that four BnaHB6 homologues have distinct functions in response to stress despite high sequence similarity, possibly indicating different binding preferences with BnaABF4 and BnaDREB2A. We hypothesize that BnaC08HB6 and BnaA09HB6 function in a complex regulatory network under stress.

Mechanisms of low nighttime temperature promote oil accumulation in Brassica napus L. based on in-depth transcriptome analysis.

Rape (Brassica napus L.; AACC) is an important oil-bearing crop worldwide. Temperature significantly affects the production of oil crops; however, the mechanisms underlying temperature-promoted oil biosynthesis remain largely unknown. In this study, we found that a temperature-sensitive cultivar (O) could accumulate higher seed oil content under low nighttime temperatures (LNT,13°C) compared with a temperature-insensitive cultivar (S). We performed an in-depth transcriptome analysis of seeds from both cultivars grown under different nighttime temperatures. We found that low nighttime temperatures induced significant changes in the transcription patterns in the seeds of both cultivars. In contrast, the expression of genes associated with fatty acid and lipid pathways was higher in the O cultivar than in the S cultivar under low nighttime temperatures. Among these genes, we identified 14 genes associated with oil production, especially BnLPP and ACAA1, which were remarkably upregulated in the O cultivar in response to low nighttime temperatures compared to S. Further, a WGCNA analysis and qRT-PCR verification revealed that these genes were mainly regulated by five transcription factors, WRKY20, MYB86, bHLH144, bHLH95, and NAC12, whose expression was also increased in O compared to S under LNT. These results allowed the elucidation of the probable molecular mechanism of oil accumulation under LNT conditions in the O cultivar. Subsequent biochemical assays verified that BnMYB86 transcriptionally activated BnLPP expression, contributing to oil accumulation. Meanwhile, at LNT, the expression levels of these genes in the O plants were higher than at high nighttime temperatures, DEGs (SUT, PGK, PK, GPDH, ACCase, SAD, KAS II, LACS, FAD2, FAD3, KCS, KAR, ECR, GPAT, LPAAT, PAP, DGAT, STERO) related to lipid biosynthesis were also upregulated, most of which are used in oil accumulation.

BnaC06.WIP2-BnaA09.STM transcriptional regulatory module promotes leaf lobe formation in Brassica napus.

The lobed leaves of rapeseed (Brassica napus L.) offer significant advantages in dense planting, leading to increased yield. Although AtWIP2, a C2H2 zinc finger transcription factor, acts as a regulator of leaf development in Arabidopsis thaliana, the function and regulatory mechanisms of BnaWIP2 in B. napus remain unclear. Here, constitutive expression of the BnaC06.WIP2 paralog, predominantly expressed in leaf serrations, produced lobed leaves in both A. thaliana and B. napus. We demonstrated that BnaC06.WIP2 directly repressed the expression of BnaA01.TCP4, BnaA03.TCP4, and BnaC03.TCP4 and indirectly inhibited the expression of BnaA05.BOP1 and BnaC02.AS2 to promote leaf lobe formation. On the other hand, we discovered that BnaC06.WIP2 modulated the levels of endogenous gibberellin, cytokinin, and auxin, and controlled the auxin distribution in B. napus leaves, thus accelerating leaf lobe formation. Meanwhile, we revealed that BnaA09.STM physically interacted with BnaC06.WIP2, and ectopic expression of BnaA09.STM generated smaller and lobed leaves in B. napus. Furthermore, we found that BnaC06.WIP2 and BnaA09.STM synergistically promoted leaf lobe formation through forming transcriptional regulatory module. Collectively, our findings not only facilitate in-depth understanding of the regulatory mechanisms underlying lobed leaf formation, but also are helpful for guiding high-density breeding practices through improving leaf morphology in B. napus.

Genome-Wide Identification and Characterization of the CCT Gene Family in Rapeseed (Brassica napus L.).

The CCT gene family is present in plants and is involved in biological processes such as flowering, circadian rhythm regulation, plant growth and development, and stress resistance. We identified 87, 62, 46, and 40 CCTs at the whole-genome level in B. napus, B. rapa, B. oleracea, and A. thaliana, respectively. The CCTs can be classified into five groups based on evolutionary relationships, and each of these groups can be further subdivided into three subfamilies (COL, CMF, and PRR) based on function. Our analysis of chromosome localization, gene structure, collinearity, cis-acting elements, and expression patterns in B. napus revealed that the distribution of the 87 BnaCCTs on the chromosomes of B. napus was uneven. Analysis of gene structure and conserved motifs revealed that, with the exception of a few genes that may have lost structural domains, the majority of genes within the same group exhibited similar structures and conserved domains. The gene collinearity analysis identified 72 orthologous genes, indicating gene duplication and expansion during the evolution of BnaCCTs. Analysis of cis-acting elements identified several elements related to abiotic and biotic stress, plant hormone response, and plant growth and development in the promoter regions of BnaCCTs. Expression pattern and protein interaction network analysis showed that BnaCCTs are differentially expressed in various tissues and under stress conditions. The PRR subfamily genes have the highest number of interacting proteins, indicating their significant role in the growth, development, and response to abiotic stress of B. napus.

Treatment of marble industry wastewater by Brassica napus (L.) under oxalic acid amendment: efficacy as fodder and carcinogenic risk assessment.

In the present study, Brassica napus, a food plant, was grown for phytoextraction of selected heavy metals (HMs) from marble industry wastewater (WW) under oxalic acid (OA) amendment. The hydroponic experiment was performed under different combination of WW with OA in complete randomized design. Photosynthetic pigments and growth reduction were observed in plants treated with WW alone amendments. The combination of OA in combination with WW significantly enhanced the growth of plants along with antioxidant enzyme activities compared with WW-treated-only plants. HM stress alone enhanced the hydrogen peroxide, electrolyte leakage, and malondialdehyde contents in plants. OA-treated plants were observed with enhanced accumulation of cadmium (Cd), copper (Cu), and lead (Pb) concentrations in the roots and shoots of B. napus. The maximum concentration and accumulation of Cd in root, stem, and leaves was increased by 25%, 30%, and 30%; Cu by 42%, 24%, and 17%; and Pb by 45%, 24%, and 43%, respectively, under OA amendment. Average daily intake and hazard quotient (HQ) were calculated for males, females, and children in two phases of treatments in phytoremediation of metals before and after accumulation into B. napus leaves and stems. HQ of metals in the leaves and stem was < 1 before metal accumulation, whereas > 1 was observed after HM accumulation for all males, females, and children. Similarly, the hazard index of the three study types was found > 1. It was observed that the estimated excess lifetime cancer risk was of grade VII (very high risk), not within the accepted range of 1 × 10-4 to 1 × 10-6. Based on the present study, the increased levels of HMs up to carcinogenicity was observed in the B. napus which is not safe to be consumed later as food.