ABSTRACT Heat stress increasingly threatens global wheat production as extreme temperature events become more frequent under climate change. To better understand the molecular basis of thermotolerance, we evaluated the heat responses of 30 elite wheat accessions and identified substantial natural variation in seedling survival (26.7%), plant height (26.0%–68.8%), and shoot fresh weight (5.7%–86.2%). We subsequently conducted time‐resolved transcriptome profiling of two contrasting genotypes, the heat‐resistant Huaimai211 and the heat‐susceptible Chuke316, across five heat‐stress durations. RNA‐seq analysis revealed 8517 and 4155 genes that were consistently responsive to heat stress in the HR and HS genotypes, respectively, with 3028 core genes shared by both genotypes. These core genes were further enriched in pathways related to Hsp90 binding, protein folding, the unfolded protein response, and glutathione metabolism. Conversely, genes associated with photosynthesis, chlorophyll‐binding proteins, and Calvin‐cycle enzymes were persistently downregulated across all time points, indicating sustained repression of photosynthetic processes during prolonged heat exposure. Genotype‐specific analyses revealed that HR uniquely activated metabolic and protein‐folding pathways related to amino‐acid biosynthesis, fructose‐bisphosphate aldolase activity, and peptidyl‐prolyl isomerase function. Co‐expression network analysis identified several heat‐associated modules, among which the firebrick3 module showed the strongest association with thermotolerance and highlighted TaRbcS‐2B.3 as a hub gene linking photosynthetic adjustment to stress adaptation. Together, these results reveal a coordinated heat‐response framework involving persistent photosynthetic suppression, reinforcement of proteostasis, and genotype‐specific metabolic plasticity. The identified pathways and candidate genes provide valuable targets for marker‐assisted selection, gene editing, and genomic prediction efforts aimed at improving wheat thermotolerance, thereby supporting yield stability under increasingly frequent heat events.

Plant growth and production are strongly affected by water deficit. Mitogen-activated protein kinase kinase kinases (MAP3Ks) are essential signaling components for responses to abiotic stress in many plants. However, the molecular mechanisms underlying their roles in drought-stress tolerance in wheat (Triticum aestivum) remain unclear. In this study, we identified TaMAP3K17 as a drought-induced MAP3K gene in wheat and demonstrated that it encodes a protein localized to both the plasma membrane and nucleus. TaMAP3K17-silenced plants generated by virus-induced gene silencing (VIGS) were hypersensitive to drought stress and accumulated higher levels of reactive oxygen species (ROS) compared to the wild type. TaMAP3K17 overexpression enhanced drought-stress tolerance in transgenic wheat plants. Yeast one-hybrid and dual-luciferase reporter assays showed that TaMYB2 activates the transcription of TaMAP3K17. Suppressing TaMYB2 expression by VIGS resulted in ROS generation and malondialdehyde accumulation in wheat under water-deficit conditions. These findings suggest that a TaMYB2-TaMAP3K17 regulatory module is involved in ROS homeostasis and drought responses in wheat.

Fusarium crown rot (FCR) is a hard-to-control wheat disease prevalent in arid and semi-arid regions. However, the relationship between arid conditions and FCR disease remains unclear. Here, we confirm that drought stress exacerbates the severity of FCR, and that FCR intensifies drought-induced damage. Integrated transcriptome analysis indicates that TaMPK3 gene exhibits distinctly opposite expression patterns under these two stress conditions. Functional identification verifies that TaMPK3 positively regulate FCR resistance while negatively influencing drought tolerance. Upon TaMPK3 gene knockout, the mutually reinforcing effect between drought and FCR is eliminated. TaMPK3 is found to modulate TaWRKY26 activity to regulate the expression of sterol synthesis gene clusters, thereby influencing FCR resistance. Within this cluster, the key gene TaCYP51H37 significantly enhances FCR resistance. Combined with our findings that TaMPK3 decreases drought tolerance through ABA signaling pathway, this study proposes a molecular model in which TaMPK3 mediates the synergistic damage caused by drought and FCR.

The Fantastic Four gene family encodes small, plant-specific regulatory proteins involved in developmental control; however, their roles in wheat remain poorly understood. In this study, we conducted a comprehensive genome-wide analysis of the Fantastic Four gene family in wheat. A total of 42 TaFAF genes were identified and systematically characterized in terms of their chromosomal distribution, phylogenetic relationships, gene structures, conserved motifs, and promoter cis-regulatory elements. Phylogenetic analysis classified TaFAF genes into four distinct clades, which exhibit high structural conservation but show divergent motif compositions. Expression profiling revealed tissue-specific expression patterns and suggested that a subset of TaFAF genes responded transcriptionally to heat stress in a genotype-dependent manner. Subcellular localization assays showed that representative Fantastic Four proteins were localized in the cytoplasm. Protein-protein interaction analyses indicated that TaFAF-1A.1 and TaFAF-5D.5 physically interact with the key flowering regulator TaFT1. Furthermore, haplotype analysis of TaFAF-5D.5 across 145 wheat accessions revealed a significant association with wheat growth habit, with a favorable haplotype preferentially enriched in winter wheat. Together, these results provide insights into the evolutionary diversification and functional relevance of the Fantastic Four genes and identify TaFAF-5D.5 as a candidate gene potentially associated with developmental adaptation and heat stress responses in wheat.

The wheat (Triticum aestivum) brassinazole-resistant 2 (TaBZR2) gene is identified as significantly associated with drought tolerance by genome-wide association study (GWAS), and a chloroplast pentatricopeptide repeat (PPR) protein gene TaPPR13 functioned as a positive drought stress regulator downstream of TaBZR2. Overexpression of TaPPR13 enhanced the antioxidative defense system, whereas knockdown of TaPPR13 led to the accumulation of reactive oxygen species (ROS) and caused abnormalities in chloroplast thylakoids under drought stress conditions. RNA-seq analysis showed that overexpression of TaPPR13 significantly upregulated the expression of nuclear-encoded genes involved in ROS scavenging and the abscisic acid (ABA) signaling pathway. Furthermore, TaPPR13 interacted with TaAOR1 and TaSIG5 to facilitate detoxification and regulate chloroplast gene expression, thereby enhancing drought tolerance. Overexpression of TaPPR13 and TaAOR1 mediated stomatal closure to reduce water loss, improving photosynthetic capacity and conferring a yield advantage under drought stress. These findings show that TaPPR13 promotes retrograde signaling to alter nuclear gene expression, with the TaBZR2-TaPPR13-TaAOR1/TaSIG5 module representing a novel signaling pathway that likely plays a pivotal role in drought stress response.

Heat stress caused by increasing global temperature has become a major factor limiting yield in wheat. Heat shock transcription factors (Hsfs), as the primary regulators in plant responses to heat stress, play essential roles in modulating both basal and acquired thermotolerance in plants. However, the underlying molecular mechanisms remain to be elucidated. By analysing the wheat transcriptome after subjecting wheat to heat treatments for different time intervals, we identified gene TaHsfA2h that showed a significant positive regulatory response to heat stress. Heat stress tolerance was enhanced by overexpression of TaHsfA2h and constrained by its RNA interference. RNA-seq analysis demonstrated that the overexpression of TaHsfA2h significantly enhanced the expression levels of genes involved in ABA and ROS signalling pathways. Additionally, we identified TaABF5b, a critical regulatory factor in the ABA signalling pathway, as being capable of modulating the expression of TaHsfA2h. Notably, TaHsfA2h interacted with TaHsfC2a both in vivo and in vitro. Similarly, overexpression of TaHsfC2a significantly enhanced heat stress tolerance, whereas knockout dramatically reduced tolerance. The presence of TaHsfC2a significantly enhanced the regulatory activity of TaHsfA2h. TaHsfA2h and TaHsfC2a can co-regulate the expression levels of heat stress tolerance-related genes, including TaNCED2B, TaPOD4 and TaHSP26, thereby enhancing wheat's tolerance to heat stress. Overall, our findings revealed a positive regulatory function of the ABF5b-HsfA2h/HsfC2a-NCED2b/POD4/HSP26 module on wheat heat stress tolerance. This discovery further expanded the functionality of a plant heat stress response model, providing a theoretical foundation for the development of heat-tolerant wheat varieties.

Phosphate deficiency and drought are significant environmental constraints that impact both the productivity and quality of wheat. The interaction between phosphorus and water facilitates their mutual absorption processes in plants. Under conditions of both phosphorus deficiency and drought stress, we observed a significant upregulation in the expression of wheat MYB-CC transcription factors through the transcriptome analysis. 52 TaMYB-CC genes in wheat were identified and analyzed their evolutionary relationships, structures, and expression patterns. The TaMYB-CC5 gene exhibited specific expression in roots and demonstrated significant upregulation under phosphorus deficiency and drought stress compared to other TaMYB-CC genes. The overexpression of TaMYB-CC5A in Arabidopsis resulted in a significant increase of root length under stress conditions, thereby enhancing tolerance to phosphate starvation and drought stress. The wheat lines with silenced TaMYB-CC5 genes exhibited reduced root length under stress conditions and increased sensitivity to phosphate deficiency and drought stress. In addition, silencing the TaMYB-CC5 genes resulted in altered phosphorus content in leaves but did not lead to a reduction in phosphorus content in roots. Enrichment analysis the co-expression genes of TaMYB-CC5 transcription factors, we found the zinc-induced facilitator-like (ZIFL) genes were prominent associated with TaMYB-CC5 gene. The TaZIFL1, TaZIFL2, and TaZIFL5 genes were verified specifically expressed in roots and regulated by TaMYB-CC5 transcript factor. Our study reveals the pivotal role of the TaMYB-CC5 gene in regulating TaZIFL genes, which is crucial for maintaining normal root growth under phosphorus deficiency and drought stress, thereby enhanced resistance to these abiotic stresses in wheat.

Sterols have long been associated with diverse fields, such as cancer treatment, drug development, and plant growth; however, their underlying mechanisms and functions remain enigmatic. Here, we unveil a critical role played by a GmNF-YC9-mediated CCAAT-box transcription complex in modulating the steroid metabolism pathway within soybeans. Specifically, this complex directly activates squalene monooxygenase (GmSQE1), which is a rate-limiting enzyme in steroid synthesis. Our findings demonstrate that overexpression of either GmNF-YC9 or GmSQE1 significantly enhances soybean stress tolerance, while the inhibition of SQE weakens this tolerance. Field experiments conducted over two seasons further reveal increased yields per plant in both GmNF-YC9 and GmSQE1 overexpressing plants under drought stress conditions. This enhanced stress tolerance is attributed to the reduction of abiotic stress-induced cell oxidative damage. Transcriptome and metabolome analyses shed light on the upregulation of multiple sterol compounds, including fucosterol and soyasaponin II, in GmNF-YC9 and GmSQE1 overexpressing soybean plants under stress conditions. Intriguingly, the application of soybean steroids, including fucosterol and soyasaponin II, significantly improves drought tolerance in soybean, wheat, foxtail millet, and maize. These findings underscore the pivotal role of soybean steroids in countering oxidative stress in plants and offer a new research strategy for enhancing crop stress tolerance and quality from gene regulation to chemical intervention.

Wheat growth process has been experiencing severe challenges arising from the adverse environment. Notably, the incidence of Fusarium crown rot (FCR), a severe soil-borne disease caused by Fusarium pseudograminearum (Fp), has significantly intensified in various wheat-growing regions, resulting in a decline in grain yield. However, the identification of wheat varieties and the exploration of effective gene resources resistant to FCR have not yet been accomplished. Here, we screened and identified the tryptophan metabolism pathway to participate in wheat resistance to FCR by correlation analysis between transcriptome and metabolome, and found that indole-3-acetaldehyde (IAAld) and melatonin, two key metabolites in the tryptophan metabolic pathway, were significantly accumulated in Fp-induced wheat stem bases. Interestingly, exogenous application of these two metabolites could significantly enhance wheat resistance against Fp. Additionally, we observed that the activity of TaALDHase, a crucial enzyme responsible for catalyzing IAAld to produce indole-3-acetic acid (IAA), was inhibited. Conversely, the activity of TaMTase, a rate-limiting involved in melatonin biosynthesis, was enhanced in the Fp-induced wheat transcriptome. Further analysis showed that TaWRKY24 could regulate IAA and melatonin biosynthesis by inhibiting the expression of TaALDHase and enhancing the transcription of TaMTase, respectively. Silencing of TaALDHase could significantly increase wheat resistance to FCR. However, interference with TaWRKY24 or TaMTase could decrease wheat resistance to FCR. Collectively, our findings demonstrate the crucial role of the tryptophan metabolism pathway in conferring resistance against FCR in wheat, thereby expanding its repertoire of biological functions within the plant system.

INTRODUCTION:Heat stress poses a severe threat to the growth and production of soybean (Glycine max). Brassinosteroids (BRs) actively participate in plant responses to abiotic stresses, however, the role of BR signaling pathway genes in response to heat stress in soybean remains poorly understood. OBJECTIVES:In this study, we investigate the regulatory mechanisms of GmBSK1 and GmBES1.5 in response to heat stress and the physiological characteristics and yield performance under heat stress conditions. METHODS:Transgenic technology and CRISPR/Cas9 technology were used to generated GmBSK1-OE, GmBES1.5-OE and gmbsk1 transgenic soybean plants, and transcriptome analysis, LUC activity assay and EMSA assay were carried out to elucidate the potential molecular mechanism underlying GmBSK1-GmBES1.5-mediated heat stress tolerance in soybean. RESULTS:CRISPR/Cas9-generated gmbsk1 knockout mutants exhibited increased sensitivity to heat stress due to a reduction in their ability to scavenge reactive oxygen species (ROS). The expression of GmBES1.5 was up-regulated in GmBSK1-OE plants under heat stress conditions, and it directly binds to the E-box motif present in the promoters of abiotic stress-related genes, thereby enhancing heat stress tolerance in soybean plants. Furthermore, we identified an interaction between GmGSK1 and GmBES1.5, while GmGSK1 inhibits the transcriptional activity of GmBES1.5. Interestingly, the interaction between GmBSK1 and GmGSK1 promotes the localization of GmGSK1 to the plasma membrane and releases the transcriptional activity of GmBES1.5. CONCLUSION:Our findings suggest that both GmBSK1 and GmBES1.5 play crucial roles in conferring heat stress tolerance, highlighting a potential strategy for breeding heat-tolerant soybean crops involving the regulatory module consisting of GmBSK1-GmGSK1-GmBES1.5.

Global climate change has resulted in increasingly harsh environments that have become the restriction factors for plants growth and development. Calcium-dependent protein kinases (CDPKs) in plants play critical roles in resisting various abiotic stresses, such as heat and cold stresses. Using the expression pattern analysis of foxtail millet CDPK family members under heat- and cold-treatments, respectively, and the SiCDPK7 was found to respond to extreme temperature stress and was selected for further study. The results suggested that overexpression of SiCDPK7 in Arabidopsis conferred tolerance to heat stress by increasing seedling survival rates and hypocotyl elongation compared with wild type (WT) plants, and also can enhance heat stress tolerance in foxtail millet. Analysis of physiological and biochemical indexes showed that SiCDPK7 transgenic plant lines had the markedly higher catalase (CAT) activity and the significantly lower malonaldehyde (MDA) content than WT plants. In addition, qRT-PCR analysis showed that the transcription levels of heat and cold stress-responsive genes were significantly increased in SiCDPK7 transgenic Arabidopsis and foxtail millet under stress conditions. This supporting evidence suggests that SiCDPK7 facilitated the extreme temperature tolerance capabilities in plants

SGT1（Suppressor of the G2 Allele of skP1） is an inhibitor of skp1-4, which plays an important role in the abiotic stress response of plants. Based on the early transcriptomics and proteinomics analyses of common buckwheat under drought stress, a FeSGT1 gene was screened and cloned, which contained a 1086 bp open reading frame encoding 361 amino acids and 3 domains including TPR, CS, and SGS. Homologous protein comparison showed that Fe SGT1 was closely related to CqSGT1（XP＿021726759.1）, BvSGT1（XP＿010671588.1）, and SoSGT1（XP＿021839743.1）. Besides, FeSGT1 gene encoded membrane localization protein. The relative expression levels revealed that FeSGT1 tended to be up-regulated within 24 hours of drought stress. The expression of FeSGT1 gene peaked at 12 hours and began to decline after 24 hours under salt, low temperature（4℃）,and ABA treatments. Overexpression of FeSGT1 gene in transgenic Arabidopsis not only conferred drought and salt tolerance, but also significantly increased root length, fresh weight, and survival rate compared with the wild type（WT） plant, accompanied by the elevated activities of catalase（CAT）, the lowered malonaldehyde（MDA） and H2O2 contents, thus allowing plants to better adapt to adverse environments. Our results provided information in the exploring of the molecular regulation mechanism responding to drought tolerance in common buckwheat.

The calcineurin B-like protein (CBL)-CBL interacting protein kinase (CIPK) signaling pathway is important for plants to response to environmental stresses. However, few academic studies have elucidated CIPKs in soybean, and the mechanism by which CIPKs promote drought resistance is unclear. In this study, the GmCIPK29 was found to interact with GmCBL1, expression analysis and GUS staining showed that transcript level of GmCIPK29 was induced by drought and exogenous abscisic acid (ABA). GmCIPK29 increased ABA sensitivity and conferred drought resistance in Arabidopsis. Moreover, GmCIPK29-overexpression (GmCIPK29-OE) Arabidopsis plants showed increased stomatal movement and reduced water loss under water deficit condition, and the content of malondialdehyde (MDA) decreased under drought stress in the GmCIPK29-OE soybean hairy roots. In addition, soybean hairy roots with GmCIPK29 knocked down via RNA interference showed higher levels of reactive oxygen species (ROS) under water deficit conditions. Gene expression analysis showed that stress-related genes, including GmCAT5, GmPOD4, and GmNCED1, were induced in GmCIPK29-OE soybean plants under drought stress. This study revealed an important role of soybean GmCIPK29 in responses to abiotic stress and these findings provide a foundation for additional inquiry into the complex mechanisms of the CBL-CIPK signaling pathway.

A soybean elongation factor Tu family (EF-Tu) protein, GmEF8, was determined to interact with GmCBL1, and GmEF8 expression was found to be induced by various abiotic stresses such as drought and heat. An ortholog of GmEF8 was identified in Arabidopsis, a T-DNA knockout line for which exhibited hypersensitivity to drought and heat stresses. Complementation with GmEF8 rescued the sensitivity of the Arabidopsis mutant to drought and heat stresses, and GmEF8 overexpression conferred drought and heat tolerance to transgenic Arabidopsis plants. In soybean, plants with GmEF8-overexpressing hairy roots (OE-GmEF8) exhibited enhanced drought and heat tolerance and had higher proline levels compared to plants with RNAi GmEF8-knockdown hairy roots (MR-GmEF8) and control hairy roots (EV). A number of drought-responsive genes, such as GmRD22 and GmP5CS, were induced in the OE-GmEF8 line compared to MR-GmEF8 and EV under normal growth conditions. These results suggest that GmEF8 has a positive role in regulating drought and heat stresses in Arabidopsis and soybean. This study reveals a potential role of the soybean GmEF8 gene in response to abiotic stresses, providing a foundation for further investigation into the complexities of stress signal transduction pathways.

为了更准确、高效地揭示小麦不同器官的叶绿素含量,本研究以小麦的旗叶、小花和花药为材料,分别采用80％丙酮研磨法(Arnon法)、丙酮乙醇研磨法、丙酮乙醇浸提法来提取小麦不同组织器官的叶绿素.结果表明:不同提取方法提取的叶绿素溶液的吸收光谱基本一致,但在提取效率上存在差异,其中丙酮乙醇研磨法为旗叶和小花叶绿素提取的最佳方法,而用丙酮乙醇浸提法(浸提18 h)提取小麦花药的效率更高,这为准确提取和检测小麦不同组织器官叶绿素提供了必要的技术支持.

Drought stress has been the main abiotic factor affecting the growth, development and production of common buckwheat (Fagopyrum esculentum). To explore the response mechanisms of regulating buckwheat drought stress on the post-transcriptional and translational levels, a comparative proteomic analysis was applied to monitor the short-term proteomic variations under the drought stress in the seedling stage. From which 593 differentially abundant proteins (DAPs) were identified using the TMT-based proteomics analysis. A number of DAPs were found to be intimately correlated with the styrene degradation, phenylpropanoid biosynthesis and stimulus response, within which. The acyl-CoA oxidase 4 (ACX4), a key regulator in plant abiotic stress response, was selected for further elucidation. Overexpression of the FeACX4 not only conferred drought and salt tolerance in the Arabidopsis, but also significantly increased the root length and fresh weight in the overexpression lines plant relative to the wild type (WT) plant, accompanied by the elevated activities of catalase (CAT) and lowered malonaldehyde (MDA) and H2O2 contents, therefore allowing plants to better adapt to adverse environments. Our results provided information in the exploring of the molecular regulation mechanism responding to drought tolerance in common buckwheat.

Plant non-specific lipid transfer proteins (nsLTPs/LTPs) play important roles in plant growth, development and abiotic stress responses. However, little is known about LTPs in soybean. In this study, 131 LTPs were identified and divided into ten types (I, II, III, IV, V, VI, VII, VIII, IX, and Y), and each GmLtp has the characteristic eight-cysteine motif (8 CM) of plant LTPs. RNA-seq data showed that most GmLtps responded to a variety of abiotic stresses. Among the GmLtp members, GmLtpⅠ.3 was significantly induced under drought and salt treatments. Overexpressing GmLtpⅠ.3 improved tolerance to drought and salt stresses in transgenic Arabidopsis and soybean. The results showed that the accumulation of reactive oxygen species (ROS) in the roots of GmLtpⅠ.3-overexpressing (OE-GmLtpⅠ.3) plants decreased significantly, compare with GmLtpⅠ.3 RNA interference (RNAi-GmLtpⅠ.3) and empty vector (EV-Control) plants. Furthermore, compared with EV-Control, the expression level of GmSOD1 and GmSOD2 increased in OE-GmLtpI.3. Contrarily, the expression of GmRBOHA and GmRBOHB decreased in OE-GmLtpI.3. The results of this study provide the basis for further comprehensive analyses of LTP genes and insights into abiotic stress response mechanisms in soybean.

Drought stress impairs crop growth and productivity. Stress-associated proteins (SAPs), a class of zinc finger proteins containing the A20/AN1 domain, function in various stress responses in plants. However, little is known about the function of SAPs in drought-stress responses in soybean, an oil and protein crop. We report that a GmSAP5 protein confers drought tolerance by increasing sensitivity to abscisic acid (ABA) and reducing stomatal aperture. Overexpression and RNA interference of GmSAP5 in soybean hairy roots resulted in elevated resistance and sensitivity to drought stress, respectively. ABA and proline contents increased in GmSAP5-overexpressing plants under water-deficit conditions. Lower water loss rates and higher relative water contents were observed in GmSAP5-overexpressing plants, resulting in increased drought-stress resistance. A yeast one-hybrid assay and luciferase transient transcriptional activity assay showed that GmAREB3, an AREB/ABF transcription factor, could bind to the promoter of GmSAP5 and activate its expression. These results suggest that GmSAP5 acts downstream of GmAREB3 and improves drought-stress resistance by mediating ABA signaling.

Tubby-like proteins (TLPs) are transcription factors that are widely present in eukaryotes and generally participate in growth and developmental processes. Using genome databases, a total of 22 putative TLP genes were identified in the soybean genome, and unevenly distributed across 13 chromosomes. Phylogenetic analysis demonstrated that the predicted GmTLP proteins were divided into five groups (I-V). Gene structure, protein motifs, and conserved domains were analyzed to identify differences and common features among the GmTLPs. A three-dimensional protein model was built to show the typical structure of TLPs. Analysis of publicly available gene expression data showed that GmTLP genes were differentially expressed in response to abiotic stresses. Based on those data, GmTLP8 was selected to further explore the role of TLPs in soybean drought and salt stress responses. GmTLP8 overexpressors had improved tolerance to drought and salt stresses, whereas the opposite was true of GmTLP8 -RNAi lines. 3,3-diaminobenzidine and nitro blue tetrazolium staining and physiological indexes also showed that overexpression of GmTLP8 enhanced the tolerance of soybean to drought and salt stresses; in addition, downstream stress-responsive genes were upregulated in response to drought and salt stresses. This study provides new insights into the function of GmTLPs in response to abiotic stresses.