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We previously reported a structural rearrangement between wheat (Triticum aestivum) and rye (Secale cereale) chromosomes 1BS/1RS that increased the dosage of 12-OXOPHYTODIENOATE REDUCTASE III (OPRIII) genes involved in jasmonate biosynthesis (henceforth, 1RW line), and that drastically reduced primary root growth relative to a control line with the intact 1RS chromosome (henceforth, 1RS). In this study, we show that the increased gene-dosage of this region is associated with increases in the shoot-root partitioning of magnesium (Mg). Moreover, both a CRISPR-edited 1RW line with reduced OPRIII dosage and the 1RW line treated with the jasmonate biosynthesis inhibitor ibuprofen showed reduced differences in shoot-root Mg partitioning than 1RW. The observed differences in Mg partitioning between 1RS and 1RW plants occur over a wide range of external Mg supplies and imply opposite trends of Mg accumulation in roots and shoots. Furthermore, we show an association between the increase of shoot-root Mg partitioning and increased tolerance of the 1RW line to low levels of Mg supply. In summary, our results provide evidence of the role of the jasmonate pathway on the dynamics of Mg accumulation in roots and shoots, which correlates with the performance of wheat plants under conditions of Mg scarcity., Competing Interests: Declaration of competing interest The authors have no conflict of interest to declare., (Copyright © 2024 The Authors. Published by Elsevier GmbH.. All rights reserved.)

APX proteins are H 2 O 2 -scavenging enzymes induced during oxidative stress. In the first part of this study, we provided an extensive knowledge on the APX family of Triticum durum, TdAPX and their related TdAPX-R, via the genome wide analysis. The outcomes showed that these proteins are clustered into four major subgroups. Furthermore, the exon-intron structure and the synteny analyses revealed that during evolution the genes TdAPX and TdAPX-R are relatively conserved. Besides, during their evolution, these genes underwent purifying selection pressure and were duplicated in segmental. In parallel, the analysis of the conserved motifs and the multiple sequence alignment demonstrated that the residues involved in the active sites, heme- and cations-binding are conserved only in TdAPX proteins. Following the RNA-seq data and the regulatory elements analyses, we focused in the second part of this study on the functional characterization of TdAPX7B-2. The qRT-PCR data showed the upregulation of TdAPX7B-2 essentially in leaves of durum wheat exposed to salt, cold, drought, metals and ABA treatments. The tolerance phenotype of the TdAPX7B-2-expressing Arabidopsis lines to salt, direct-induced oxidative stress and heavy metals was manifested by the development of root system, proline accumulation and induction of the antioxidant CAT, SOD and POD enzymes to maintain the non-toxic H 2 O 2 levels. Likewise, the response to salt stress and direct-oxidative stress of the transgenic lines was accompanied mainly by the induction of AtNCED3, AtRD29A/B and AtERD1., Competing Interests: Declarations. Ethical approval: The study does not require ethics approval as we have not used human or animal subject. Conflict of interest: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Lipophilic cuticles mainly composed of wax mixtures and cutin matrices seal the plant epidermis and control plant development and environmental adaptation. Although cuticle-associated traits have been selected in the breeding of agronomically important cereal bread wheat, the biosynthesis of wheat cuticular wax and cutin remains poorly understood. Herein, wheat topoisomerase VI was characterized as an essential activator of cuticular wax and cutin biosynthesis. Knock-down of wheat TaTOP6A , TaTOP6B , TaRHL1 , or TaBIN4 gene encoding component of topoisomerase VI resulted in decreased loads of leaf cuticular wax and cutin, as well as increased leaf cuticle permeability. Moreover, TaCYP86A2 was identified as a key component of the wheat cutin biosynthetic machinery. Reduction of wheat TaCYP86A2 expression led to decreased cutin accumulation and enhanced cuticle permeability. In addition, TaTOP6A, TaTOP6B, TaRHL1, or TaBIN4 was shown to enrich at the promoter regions of the wax biosynthesis gene TaKCS1 and the cutin biosynthesis gene TaCYP86A2 . Importantly, chromatin at TaKCS1 and TaCYP86A2 promoters is marked by high nucleosome occupancy and low histone acetylation in TaTOP6A -, TaTOP6B -, TaRHL1 -, or TaBIN4 -silenced wheat leaves. These results collectively support that wheat topoisomerase VI positively regulates the biosynthesis of cuticular wax and cutin probably via maintaining a permissive chromatin state at TaKCS1 and TaCYP86A2 genes.

Background: Sulphotransferase (SOT) enzyme (encoded by a conserved family of SOT genes) is involved in sulphonation of a variety of compounds, through transfer of a sulphuryl moiety from 3'phosphoadenosine- 5'phosphosulphate (PAPS) to a variety of secondary metabolites. The PAPS itself is derived from 3'adenosine-5'phosphosulphate (APS) that is formed after uptake of sulphate ions from the soil. The process provides tolerance against abiotic stresses like drought and heat in plants. Therefore, a knowledge of SOT genes in any crop may help in designing molecular breeding methods for improvement of tolerance for drought and heat., Methods: Sequences of rice SOT genes and SOT domain (PF00685) of corresponding proteins were both used for identification of SOT genes in wheat and six related species (T. urartu, Ae. tauschii, T. turgidum, Z. mays, B. distachyon and Hordeum vulgare), although detailed analysis was conducted only in wheat. The wheat genes were mapped on individual chromosomes and also subjected to synteny and collinearity analysis. The proteins encoded by these genes were examined for the presence of a complete SOT domain using 'Conserved Domain Database' (CDD) search tool at NCBI., Results: In wheat, 107 TaSOT genes, ranging in length from 969 bp to 7636 bp, were identified and mapped onto individual chromosomes. SSRs (simple sequence repeats), microRNAs, long non-coding RNAs (lncRNAs) and their target sites were also identified in wheat SOT genes. SOT proteins were also studied in detail. An expression assay of TaSOT genes via wheat RNA-seq data suggested engagement of these genes in growth, development and responses to various hormones and biotic/abiotic stresses., Conclusions: The results of the present study should help in further functional characterization of SOT genes in wheat and other related crops., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The regulation solutions and mechanisms of reducing pesticide phytotoxicity to nontarget plants are not well-defined and detailed. Here, we have proposed a new detoxification strategy to control the toxic effects of herbicide imazethapyr (IM) induced in wheat seedlings from the perspective of the plasma membrane (PM) H + -ATPase. We found that the changes in PM H + -ATPase activity have a regulatory effect on the phytotoxic effects induced by IM in plants. Treatment with PM H + -ATPase activators restored the reduced auxin content and photosynthetic efficiency caused by IM, thereby promoting plant growth. Application of a PM H + -ATPase inhibitor further reduced phosphorus content and significantly increased 2,4-dihydroxy-7-methoxy-2 H ,1,4-benzoxazin-3(4 H )one (DIMBOA) and jasmonic acid levels. These effects indicate that auxin and DIMBOA may regulate plant growth trends and detoxification effects mediated by PM H + -ATPase. This work opens a new strategy for regulating herbicide toxicity to nontarget plants from the PM H + -ATPase.

Seed inoculation with entomopathogenic fungi (EPF) causes plant-mediated effects against arthropod herbivores, but the responses vary among EPF isolates. We used a wheat model system with three isolates representing Beauveria bassiana and Metarhizium spp. causing either negative or positive effects against the aphid Rhopalosiphum padi. Activities of six carbohydrate enzymes increased in plants showing biomass build-up after EPF inoculations. However, only aldolase activity showed positive correlation with R. padi numbers. Plants inoculated with M. robertsii hosted fewest aphids and showed increased activity of superoxide dismutase, implying a defense strategy of resistance towards herbivores. In M. brunneum-inoculated plants, hosting most R. padi, activities of catalase and glutathione reductase were increased suggesting enhanced detoxification responses towards aphids. However, M. brunneum simultaneously increased plant growth indicating that this isolate may cause the plant to tolerate herbivory. EPF seed inoculants may therefore mediate either tolerance or resistance towards biotic stress in plants in an isolate-dependent manner., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

The alternative oxidase (AOX), a common terminal oxidase in the electron transfer chain (ETC) of plants, plays a crucial role in stress resilience and plant growth and development. Oat ( Avena sativa ), an important crop with high nutritional value, has not been comprehensively studied regarding the AsAOX gene family. Therefore, this study explored the responses and potential functions of the AsAOX gene family to various abiotic stresses and their potential evolutionary pathways. Additionally, we conducted a genome-wide analysis to explore the evolutionary conservation and divergence of AOX gene families among three Avena species ( Avena sativa , Avena insularis , Avena longiglumis ) and four Poaceae species ( Avena sativa , Oryza sativa , Triticum aestivum , and Brachypodium distachyon ). We identified 12 AsAOX, 9 AiAOX, and 4 AlAOX gene family members. Phylogenetic, motif, domain, gene structure, and selective pressure analyses revealed that most AsAOXs, AiAOXs, and AlAOXs are evolutionarily conserved. We also identified 16 AsAOX segmental duplication pairs, suggesting that segmental duplication may have contributed to the expansion of the AsAOX gene family, potentially preserving these genes through subfunctionalization. Chromosome polyploidization, gene structural variations, and gene fragment recombination likely contributed to the evolution and expansion of the AsAOX gene family as well. Additionally, we hypothesize that AsAOX2 may have potential function in resisting wounding and heat stresses, while AsAOX4 could be specifically involved in mitigating wounding stress. AsAOX11 might contribute to resistance against chromium and waterlogging stresses. AsAOX8 may have potential fuction in mitigating ABA-mediated stress. AsAOX12 and AsAOX5 are most likely to have potential function in mitigating salt and drought stresses, respectively. This study elucidates the potential evolutionary pathways of the AsAOXs gene family, explores their responses and potential functions to various abiotic stresses, identifies potential candidate genes for future functional studies, and facilitates molecular breeding applications in A. sativa .

Background: CINNAMYL ALCOHOL DEHYDROGENASE (CAD) catalyzes the NADPH-dependent reduction of cinnamaldehydes into cinnamyl alcohols and is a key enzyme found at the final step of the monolignol pathway. Cinnamyl alcohols and their conjugates are subsequently polymerized in the secondary cell wall to form lignin. CAD genes are typically encoded by multi-gene families and thus traditionally organized into general classifications of functional relevance., Results: In silico analysis of the hexaploid Triticum aestivum genome revealed 47 high confidence TaCAD copies, of which three were determined to be the most significant isoforms (class I) considered bone fide CADs. Class I CADs were expressed throughout development both in RNAseq data sets as well as via qRT-PCR analysis. Of the 37 class II TaCADs identified, two groups were observed to be significantly co-expressed with class I TaCADs in developing tissue and under chitin elicitation in RNAseq data sets. These co-expressed class II TaCADs were also found to be phylogenetically unrelated to a separate clade of class II TaCADs previously reported to be an influential resistance factor to pathogenic fungal infection. Lastly, two groups were phylogenetically identified as class III TaCADs, which possess distinct conserved gene structures. However, the lack of data supporting their catalytic activity for cinnamaldehydes and their bereft transcriptional presence in lignifying tissues challenges their designation and function as CADs., Conclusions: Taken together, our comprehensive transcriptomic analyses suggest that TaCAD genes contribute to overlapping but nonredundant functions during T. aestivum growth and development across a wide variety of agroecosystems and provide tolerance to various stressors., (© 2024. The Author(s).)

Sitobion miscanthi , the main species of wheat aphids, is one kind of harmful pest. Chemical insecticides are the important agrochemical products to effectively control wheat aphids. However, the broad application has led to serious resistance of pests to several insecticides, and understanding insecticide resistance mechanisms is critical for integrated pest management. In this study, SmUGGT1 , a new uridine diphosphate (UDP)-glycosyltransferase ( UGT ) gene, was cloned and more strongly expressed in the SM-R (the resistant strain to imidacloprid) than in the SM-S (the susceptible strain to imidacloprid). The increased susceptibility to imidacloprid was observed after silencing SmUGGT1 , indicating that it can be related to the resistance to imidacloprid. Subsequently, SmUGGT1 regulated post-transcriptionally in the coding sequences (CDs) by miR-81 was verified and involved in the resistance to imidacloprid in S. miscanthi . This finding is crucial in the roles of UGT involved in insecticide resistance management in pests.

Plant fungal parasites manipulate host metabolism to support their own survival. Among the many central metabolic pathways altered during infection, the glyoxylate cycle is frequently upregulated in both fungi and their host plants. Here, we examined the response of the glyoxylate cycle in bread wheat (Triticum aestivum) to infection by the obligate biotrophic fungal pathogen Puccinia striiformis f. sp. tritici (Pst). Gene expression analysis revealed that wheat genes encoding the two unique enzymes of the glyoxylate cycle, isocitrate lyase (TaICL) and malate synthase, diverged in their expression between susceptible and resistant Pst interactions. Focusing on TaICL, we determined that the TaICL B homoeolog is specifically upregulated during early stages of a successful Pst infection. Furthermore, disruption of the B homoeolog alone was sufficient to significantly perturb Pst disease progression. Indeed, Pst infection of the TaICL-B disruption mutant (TaICL-B Y400 *) was inhibited early during initial penetration, with the TaICL-B Y400 * line also accumulating high levels of malic acid, citric acid, and aconitic acid. Exogenous application of malic acid or aconitic acid also suppressed Pst infection, with trans-aconitic acid treatment having the most pronounced effect by decreasing fungal biomass 15-fold. Thus, enhanced TaICL-B expression during Pst infection may lower accumulation of malic acid and aconitic acid to promote Pst proliferation. As exogenous application of aconitic acid and malic acid has previously been shown to inhibit other critical pests and pathogens, we propose TaICL as a potential target for disruption in resistance breeding that could have wide-reaching protective benefits for wheat and beyond., (© 2024 The Author(s). The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

Background: Superoxide dismutase (SOD) can greatly scavenge reactive oxygen species (ROS) in plants. SOD activity is highly related to plant stress tolerance that can be improved by overexpression of SOD genes. Identification of SOD activity-related loci and potential candidate genes is essential for improvement of grain quality in wheat breeding. However, the loci and candidate genes for relating SOD in wheat grains are largely unknown. In the present study, grain SOD activities of 309 recombinant inbred lines (RILs) derived from the 'Berkut' × 'Worrakatta' cross were assayed by photoreduction method with nitro-blue tetrazolium (NBT) in four environments. Quantitative trait loci (QTL) of SOD activity were identified using inclusive composite interval mapping (ICIM) with the genotypic data of 50 K single nucleotide polymorphism (SNP) array., Results: Six QTL for SOD activity were mapped on chromosomes 1BL, 4DS, 5AL (2), and 5DL (2), respectively, explaining 2.2 ~ 7.4% of the phenotypic variances. Moreover, QSOD.xjau-1BL, QSOD.xjau-4DS, QSOD.xjau-5 A.1, QSOD.xjau-5 A.2, and QSOD.xjau-5DL.2 identified are likely to be new loci for SOD activity. Four candidate genes TraesCS4D01G059500, TraesCS5A01G371600, TraesCS5D01G299900, TraesCS5D01G343100LC, were identified for QSOD.xjau-4DS, QSOD.xjau-5AL.1, and QSOD.xjau-5DL.1 (2), respectively, including three SOD genes and a gene associated with SOD activity. Based on genetic effect analysis, this can be used to identify desirable alleles and excellent allele variations in wheat cultivars., Conclusion: These candidate genes are annotated for promoting SOD production and inhibiting the accumulation of ROS during plant growth. Therefore, lines with high SOD activity identified in this study may be preferred for future wheat breeding., (© 2024. The Author(s).)

Background: Environmental stresses, including high salinity and drought, severely diminish wheat yield and quality globally. The xyloglucan endotransglucosylase/hydrolase (XTH) family represents a class of cell wall-modifying enzymes and plays important roles in plants growth, development and stress adaptation. However, systematic analyses of XTH family genes and their functions under salt and drought stresses have not been undertaken in wheat., Results: In this study, we identified a total of 135 XTH genes in wheat, which were clustered into three evolutionary groups. These TaXTHs were unevenly distributed on 21 chromosomes of wheat with a majority of TaXTHs located on homelogous groups 2, 3 and 7. Gene duplication analysis revealed that segmental and tandem duplication were the main reasons for the expansion of XTH family in wheat. Interaction network predictions indicated that TaXTHs could interact with multiple proteins, including three kinases, one methyltransferase and one gibberellin-regulated protein. The promoters of the TaXTH genes harbored various cis-acting elements related to stress and hormone responses. RNA-seq data analyses showed that some TaXTH genes were induced by salt and drought stresses. Furthermore, we verified that TaXTH17 was induced by abiotic stresses and phytohormone treatments, and demonstrated that TaXTH17 was localized in the secretory pathway and cell wall. Functional analyses conducted in heterologous expression systems and in wheat established that TaXTH17 plays a negative role in plant resistance to salt and drought., Conclusions: We identified 135 XTH genes in wheat and conducted comprehensive analyses of their phylogenetic relationships, gene structures, conserved motifs, gene duplication events, chromosome locations, interaction networks, cis-acting elements and gene expression patterns. Furthermore, we provided solid evidence supporting the notion that TaXTH17 plays a negative role in plant resistance to salt and drought stresses. Collectively, our results provide valuable insights into understanding wheat XTHs, particularly their involvement in plant stress responses, and establish a foundation for further functional and mechanistic studies of TaXTHs., (© 2024. The Author(s).)

Although forward-genetics-metabolomics methods such as mGWAS and mQTL have proven effective in providing myriad loci affecting metabolite contents, they are somehow constrained by their respective constitutional flaws such as the hidden population structure for GWAS and insufficient recombinant rate for QTL. Here, the combination of mGWAS and mQTL was performed, conveying an improved statistical power to investigate the flavonoid pathways in common wheat. A total of 941 and 289 loci were, respectively, generated from mGWAS and mQTL, within which 13 of them were co-mapped using both approaches. Subsequently, the mGWAS or mQTL outputs alone and their combination were, respectively, utilized to delineate the metabolic routes. Using this approach, we identified two MYB transcription factor encoding genes and five structural genes, and the flavonoid pathway in wheat was accordingly updated. Moreover, we have discovered some rare-activity-exhibiting flavonoid glycosyl- and methyl-transferases, which may possess unique biological significance, and harnessing these novel catalytic capabilities provides potentially new breeding directions. Collectively, we propose our survey illustrates that the forward-genetics-metabolomics approaches including multiple populations with high density markers could be more frequently applied for delineating metabolic pathways in common wheat, which will ultimately contribute to metabolomics-assisted wheat crop improvement., (© 2024 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.)

Fusarium head blight (FHB), mainly incited by Fusarium graminearum , has caused great losses in grain yield and quality of wheat globally. Fhb7 , a major gene from 7E chromosome of Thinopyrum ponticum , confers broad resistance to multiple Fusarium species in wheat and has recently been cloned and identified as encoding a glutathione S-transferase ( GST ). However, some recent reports raised doubt about whether GST is the causal gene of Fhb7 . To resolve the discrepancy and validate the gene function of GST in wheat, we phenotyped Fhb7 near-isogenic lines (Jimai22- Fhb7 versus Jimai22) and GST overexpressed lines for FHB resistance. Jimai22- Fhb7 showed significantly higher FHB resistance with a lower percentage of symptomatic spikelets, Fusarium -damaged kernels, and deoxynivalenol content than susceptible Jimai22 in three experiments. All the positive GST transgenic lines driven by either the maize ubiquitin promoter or its native promoter with high gene expression in the wheat cultivar 'Fielder' showed high FHB resistance. Only one maize ubiquitin promoter-driven transgenic line showed low GST expression and similar susceptibility to Fielder, suggesting that high GST expression confers Fhb7 resistance to FHB. Knockout of GST in the Jimai22- Fhb7 line using CRISPR-Cas9-based gene editing showed significantly higher FHB susceptibility compared with the nonedited control plants. Therefore, we confirmed GST as the causal gene of Fhb7 for FHB resistance. Considering its major effect on FHB resistance, pyramiding Fhb7 with other quantitative trait loci has a great potential to create highly FHB-resistant wheat cultivars., Competing Interests: The author(s) declare no conflict of interest.

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 H 2 O 2 homeostasis by scavenging excess H 2 O 2 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., (© 2024. The Author(s).)

Mitogen-activated protein kinases (MAPKs) play important roles in plant stress response. As a major member of the MAPK family, MPK3 has been reported to participate in the regulation of chilling stress. However, the regulatory function of wheat (Triticum aestivum ) mitogen-activated protein kinase TaMPK3 in freezing tolerance remains unknown. Dongnongdongmai No.1 (Dn1) is a winter wheat variety with strong freezing tolerance; therefore, it is important to explore the mechanisms underlying this tolerance. In this study, the expression of TaMPK3 in Dn1 was detected under low temperature and hormone treatment. Gene cloning, bioinformatics and subcellular localisation analyses of TaMPK3 in Dn1 were performed. Overexpressed TaMPK3 in Arabidopsis thaliana was obtained, and freezing tolerance phenotype observations, physiological indices and expression levels of ICE-C-repeat binding factor (CBF)-COR -related genes were determined. In addition, the interaction between TaMPK3 and TaICE41 proteins was detected. We found that TaMPK3 expression responds to low temperatures and hormones, and the TaMPK3 protein is localised in the cytoplasm and nucleus. Overexpression of TaMPK3 in Arabidopsis significantly improves freezing tolerance. TaMPK3 interacts with the TaICE41 protein. In conclusion, TaMPK3 is involved in regulating the ICE-CBF-COR cold resistance module through its interaction with TaICE41, thereby improving freezing tolerance in Dn1 wheat.

Multiple genetic factors control tillering, a key agronomy trait for wheat ( Triticum aestivum L.) yield. Previously, we reported a dwarf-monoculm mutant ( dmc ) derived from wheat cultivar Guomai 301, and found that the contents of gibberellic acid 3 (GA 3 ) in the tiller primordia of dmc were significantly higher. Transcriptome analysis indicated that some wheat gibberellin oxidase ( TaGAox ) genes TaGA20ox-A2 , TaGA20ox-B2 , TaGA3ox-A2 , TaGA20ox-A4 , TaGA2ox-A10 and TaGA2ox-B10 were differentially expressed in dmc . Therefore, this study systematically analyzed the roles of gibberellin oxidase genes during wheat tillering. A total of 63 TaGAox genes were identified by whole genome analysis. The TaGAoxs were clustered to four subfamilies, GA20oxs, GA2oxs, GA3oxs and GA7oxs, including seven subgroups based on their protein structures. The promoter regions of TaGAox genes contain a large number of cis -acting elements closely related to hormone, plant growth and development, light, and abiotic stress responses. Segmental duplication events played a major role in TaGAoxs expansion. Compared to Arabidopsis , the gene collinearity degrees of the GAoxs were significantly higher among wheat, rice and maize. TaGAox genes showed tissue-specific expression patterns. The expressions of TaGAox genes ( TaGA20ox-B2 , TaGA7ox-A1 , TaGA2ox10 and TaGA3ox-A2 ) were significantly affected by exogenous GA 3 applications, which also significantly promoted tillering of Guomai 301, but didn't promote dmc . TaGA7ox-A1 overexpression transgenic wheat lines were obtained by Agrobacterium mediated transformation. Genomic PCR and first-generation sequencing demonstrated that the gene was integrated into the wheat genome. Association analysis of TaGA7ox-A1 expression level and tiller number per plant demonstrated that the tillering capacities of some TaGA7ox-A1 transgenic lines were increased. These data demonstrated that some TaGAoxs as well as GA signaling were involved in regulating wheat tillering, but the GA signaling pathway was disturbed in dmc . This study provided valuable clues for functional characterization of GAox genes in wheat., Competing Interests: The authors declare that they have no competing interests., (© 2023 Wang et al.)

Recombination UVB (sensitivity) like (RuvBL) helicase genes represent a conserved family of genes, which are known to be involved in providing tolerance against abiotic stresses like heat and drought. We identified nine wheat RuvBL genes, one each on nine different chromosomes, belonging to homoeologous groups 2, 3, and 4. The lengths of genes ranged from 1647 to 2197 bp and exhibited synteny with corresponding genes in related species including Ae. tauschii, Z. mays, O. sativa, H. vulgare, and B. distachyon. The gene sequences were associated with regulatory cis-elements and transposable elements. Two genes, namely TaRuvBL1a-4A and TaRuvBL1a-4B, also carried targets for a widely known miRNA, tae-miR164. Gene ontology revealed that these genes were closely associated with ATP-dependent formation of histone acetyltransferase complex. Analysis of the structure and function of RuvBL proteins revealed that the proteins were localized mainly in the cytoplasm. A representative gene, namely TaRuvBL1a-4A, was also shown to be involved in protein-protein interactions with ten other proteins. On the basis of phylogeny, RuvBL proteins were placed in two sub-divisions, namely RuvBL1 and RuvBL2, which were further classified into clusters and sub-clusters. In silico studies suggested that these genes were differentially expressed under heat/drought. The qRT-PCR analysis confirmed that expression of TaRuvBL genes differed among wheat cultivars, which differed in the level of thermotolerance. The present study advances our understanding of the biological role of wheat RuvBL genes and should help in planning future studies on RuvBL genes in wheat including use of RuvBL genes in breeding thermotolerant wheat cultivars., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Chlorophyll pigments are used by photosynthetic organisms to facilitate light capture and mediate the conversion of sunlight into chemical energy. Due to the indispensable nature of this pigment and its propensity to form reactive oxygen species, organisms heavily invest in its biosynthesis, recycling, and degradation. One key enzyme implicated in these processes is chlorophyllase, an α/β hydrolase that hydrolyzes the phytol tail of chlorophyll pigments to produce chlorophyllide molecules. This enzyme was discovered a century ago, but despite its importance to diverse photosynthetic organisms, there are still many missing biochemical details regarding how chlorophyllase functions. Here, we present the 4.46-Å resolution crystal structure of chlorophyllase from Triticum aestivum. This structure reveals the dimeric architecture of chlorophyllase, the arrangement of catalytic residues, an unexpected divalent metal ion-binding site, and a substrate-binding site that can accommodate a diverse range of pigments. Further, this structure exhibits the existence of both intermolecular and intramolecular disulfide bonds. We investigated the importance of these architectural features using enzyme kinetics, mass spectrometry, and thermal shift assays. Through this work, we demonstrated that the oxidation state of the Cys residues is imperative to the activity and stability of chlorophyllase, illuminating a biochemical trigger for responding to environmental stress. Additional bioinformatics analysis of the chlorophyllase enzyme family reveals widespread conservation of key catalytic residues and the identified "redox switch" among other plant chlorophyllase homologs, thus revealing key details regarding the structure-function relationships in chlorophyllase., Competing Interests: Conflict of interests The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Lead (Pb 2+ ) pollution in the soil sub-ecosystem has been a continuously growing problem due to economic development and ever-increasing anthropogenic activities across the world. In this study, the photosynthetic performance and antioxidant capacity of Triticeae cereals (rye, wheat and triticale) were compared to assess the activities of antioxidants, the degree of oxidative damage, photochemical efficiency and the levels of photosynthetic proteins under Pb stress (0.5 mM, 1 mM and 2 mM Pb (NO 3 ) 2 ). Compared with triticale, Pb treatments imposed severe oxidative damage in rye and wheat. In addition, the highest activity of major antioxidant enzymes (SOD, POD, CAT, and GPX) was also found to be elevated. Triticale accumulated the highest Pb contents in roots. The concentration of mineral ions (Mg, Ca, and K) was also high in its leaves, compared with rye and wheat. Consistently, triticale showed higher photosynthetic activity under Pb stress. Immunoblotting of proteins revealed that rye and wheat have significantly lower levels of D1 (photosystem II subunit A, PsbA) and D2 (photosystem II subunit D, PsbD) proteins, while no obvious decrease was noticed in triticale. The amount of light-harvesting complex II b6 (Lhcb6; CP24) and light-harvesting complex II b5 (Lhcb5; CP26) was significantly increased in rye and wheat. However, the increase in PsbS (photosystem II subunit S) protein only occurred in wheat and triticale exposed to Pb treatment. Taken together, these findings demonstrate that triticale shows higher antioxidant capacity and photosynthetic efficiency than wheat and rye under Pb stress, suggesting that triticale has high tolerance to Pb and could be used as a heavy metal-tolerant plant., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: TILLING (Targeting Induced Local Lesions in Genomes) is a reverse-genetic strategy that is used to locate an allelic series of induced point mutations in genes of interest. High-throughput TILLING allows the rapid and cost-effective detection of induced point mutations in populations of chemically mutagenized individuals. Grain amylose content is the major constraints for its nutritional quality and have drawn research interest. Identification of allelic variations in genes involved in starch biosynthesis in wheat endosperm is pre-requisite to amenable for nutritional quality improvement., Methods and Results: In this study, 44 EMS-induced (M4 generation) mutant lines having variation for amylose content were used for TILLING sequencing. Overall 2098.08 kb of the sequence was analyzed, and the average mutation density was 1/65.56 kb. In analysis, at the high depth score a total of 32 variations were identified including three natural variations, 76% transitions, 10% transversions, and 14% InDels respectively. The substitutions led to intronic variants, UTRs and up-downstream gene variants in Alpha-amylase, TabZIP77.1, TabZIP1 and Myb respectively. In the Myb transcription factor two missense mutations recorded namely Myb_7B c.680G > A and c.1358 T > C led to p.Gly227Asp and p.Met453Thr and c.1390G > A one substitution in Myb_7D led to p.Val464Ile., Conclusion: The identified missense substitutions were predicted to affect the protein function; hence they may have a probable role in context to the amylose content in mutants. The mutations ascertained in the current study will help in gene discovery in wheat and identified mutants can be used as genetic resources to improve nutritional quality of wheat., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Background: Phosphatidylinositol 4 phosphate 5-kinase (PIP5K) plays a key enzyme role in the inositol signal transduction system and has essential functions in plants in terms of growth, development, and stress responses. However, systematic studies on the wheat PIP5K gene family and its relation to male sterility have not been reported yet., Results: Sixty-four TaPIP5K genes were identified. The TaPIP5K genes contained similar gene structures and conserved motifs on the same branches of the evolutionary tree, and their cis-regulatory elements were related to MeJA-responsiveness. Furthermore, 49 pairs of collinearity genes were identified and mainly subjected to purification selection during evolution. Synteny analyses showed that some PIP5K genes in wheat and the other four species shared a relatively conserved evolutionary process. The expression levels of many conservative TaPIP5K genes in HT-ms anthers were significantly lower than that in Normal anthers. In addition, HT-ms anthers have no dehiscence, and levels of OPDA and JA-ILE are significantly lower at the trinucleus stage., Conclusion: These results indicate that the PIP5K gene family may be associated with male sterility induced by HT, and the reduction of JA-ILE levels and the abnormal levels of these genes expression may be one reason for the HT-ms anthers having no dehiscence, ultimately leading to the abortion of the anthers., (© 2021. The Author(s).)

Background: Peroxidase (POD) activity plays an important role in flour-based product quality, which is mainly associated with browning and bleaching effects of flour. Here, we performed a genome-wide association study (GWAS) on POD activity using an association population consisted with 207 wheat world-wide collected varieties. Our study also provide basis for the genetic improvement of flour color-based quality in wheat., Results: Twenty quantitative trait loci (QTLs) were detected associated with POD activity, explaining 5.59-12.67% of phenotypic variation. Superior alleles were positively correlated with POD activity. In addition, two SNPs were successfully developed to KASP (Kompetitive Allele-Specific PCR) markers. Two POD genes, TraesCS2B02G615700 and TraesCS2D02G583000, were aligned near the QTLs flanking genomic regions, but only TraesCS2D02G583000 displayed significant divergent expression levels (P < 0.001) between high and low POD activity varieties in the investigated association population. Therefore, it was deduced to be a candidate gene. The expression level of TraesCS2D02G583000 was assigned as a phenotype for expression GWAS (eGWAS) to screen regulatory elements. In total, 505 significant SNPs on 20 chromosomes (excluding 4D) were detected, and 9 of them located within 1 Mb interval of TraesCS2D02G583000., Conclusions: To identify genetic loci affecting POD activity in wheat grain, we conducted GWAS on POD activity and the candidate gene TraesCS2D02G583000 expression. Finally, 20 QTLs were detected for POD activity, whereas two QTLs associated SNPs were converted to KASP markers that could be used for marker-assisted breeding. Both cis- and trans-acting elements were revealed by eGWAS of TraesCS2D02G583000 expression. The present study provides genetic loci for improving POD activity across wide genetic backgrounds and largely improved the selection efficiency for breeding in wheat., (© 2021. The Author(s).)

The production of heterologous proteins is an important procedure for biologists in basic and applied sciences. A variety of cell-based and cell-free protein expression systems are available to achieve this. The expression system must be selected carefully, especially for target proteins that require post-translational modifications. In this study, human Src family kinases were prepared using six different protein expression systems: 293 human embryonic kidney cells, Escherichia coli, and cell-free expression systems derived from rabbit reticulocytes, wheat germ, insect cells, or Escherichia coli . The phosphorylation status of each kinase was analyzed by Phos-tag SDS-PAGE. The kinase activities were also investigated. In the eukaryotic systems, multiple phosphorylated forms of the expressed kinases were observed. In the rabbit reticulocyte lysate system and 293 cells, differences in phosphorylation status between the wild-type and kinase-dead mutants were observed. Whether the expressed kinase was active depended on the properties of both the kinase and each expression system. In the prokaryotic systems, Src and Hck were expressed in autophosphorylated active forms. Clear differences in post-translational phosphorylation among the protein expression systems were revealed. These results provide useful information for preparing functional proteins regulated by phosphorylation.

Salicylic acid (SA), a phytohormone, has been considered to be a key regulator mediating plant defence against pathogens. It is still vague how SA activates plant defence against herbivores such as chewing and sucking pests. Here, we used an aphid-susceptible wheat variety to investigate Sitobion avenae response to SA-induced wheat plants, and the effects of exogenous SA on some defence enzymes and phenolics in the plant immune system. In SA-treated wheat seedlings, intrinsic rate of natural increase (rm), fecundity and apterous rate of S. avenae were 0.25, 31.4 nymphs/female and 64.4%, respectively, and significantly lower than that in the controls (P < 0.05). Moreover, the increased activities of phenylalanine-ammonia-lyase, polyphenol oxidase (PPO) and peroxidase in the SA-induced seedlings obviously depended on the sampling time, whereas activities of catalase and 4-coumarate:CoA ligase were suppressed significantly at 24, 48 and 72 h in comparison with the control. Dynamic levels of p-coumaric acid at 96 h, caffeic acid at 24 and 72 h and chlorogenic acid at 24, 48 and 96 h in wheat plants were significantly upregulated by exogenous SA application. Nevertheless, only caffeic acid content was positively correlated with PPO activity in SA-treated wheat seedlings (P = 0.031). These findings indicate that exogenous SA significantly enhanced the defence of aphid-susceptible wheat variety against aphids by regulating the plant immune system, and may prove a potential application of SA in aphid control.

Background: Phosphoinositide-specific phospholipase C proteins mediate environmental stress responses in many plants. However, the potential of PI-PLC genes involved with abiotic stress tolerance in wheat remains un-explored., Objective: To study TaPLC1 genetic relation with wheat drought and heat resistance., Methods: The seedlings were treated with PI-PLC inhibitor U73122 at the single leaf stage. The seedlings were treated with drought and heat stress at the two leaf stage, and some physiological indexes and the expression profile of TaPLC1 gene were determined. And the TaPLC1 overexpression vector was transferred to Arabidopsis and selected to T3 generation for drought and heat stress treatment., Results: After 4 h of drought and heat stress, the SOD activity, MDA and soluble sugar content of the two cultivars with inhibitor were higher than those without inhibitor, the chlorophyll content decreased. CS seedlings showed significant wilting phenomenon, and TAM107 showed slight wilting. After the elimination of drought and heat stress, all seedling wilting gradually recovered, while the leaf tips of the two varieties treated with inhibitors began to wilt and turn yellow, which was more significant 5 days after the drought and heat stress, while the degree of spring wilting and yellow in CS was earlier than that in TAM107. The expression patterns of TaPLC1 gene were different in the two cultivars, but the expression levels reached the maximum at 30 min of heat stress. The change of TaPLC1 expression in TAM107 without inhibitor treatment was significantly greater than that in CS. The expression level of TaPLC1 in the two cultivars under stress was significantly different between the two cultivars treated with inhibitor and untreated, and was lower than that of the normal plants under normal conditions. These results indicated that inhibition of TaPLC1 gene expression could enhance the sensitivity of seedlings to stress. In Arabidopsis, the root lengths of transgenic and wild-type seedlings were shortened after drought stress treatment, but the root lengths of transgenic plants decreased slightly. And the expression of TaPLC1 gene was significantly increased after drought and heat stress. This indicated that overexpression of TaPLC1 improved drought resistance of Arabidopsis., Conclusions: The results of this study suggest that TaPLC1 may be involved in the regulation mechanism of drought and heat stress in wheat., (© 2021. The Genetics Society of Korea.)

Plant-pathogen interactions result in disease development in a susceptible host. Plants actively resist pathogens via a complex immune system comprising both surface-localized receptors that sense the extracellular space as well as intracellular receptors recognizing pathogen effectors. To date, the majority of cloned resistance genes encode intracellular nucleotide-binding leucine-rich repeat receptor proteins. Recent discoveries have revealed tandem kinase proteins (TKPs) as another important family of intracellular proteins involved in plant immune responses. Five TKP genes-barley Rpg1 and wheat WTK1 ( Yr15 ), WTK2 ( Sr60 ), WTK3 ( Pm24 ), and WTK4 -protect against devastating fungal diseases. Moreover, a large diversity and numerous putative TKPs exist across the plant kingdom. This review explores our current knowledge of TKPs and serves as a basis for future studies that aim to develop and exploit a deeper understanding of innate plant immunity receptor proteins.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Brassinosteroids (BRs) play key roles in diverse plant growth processes through a complex signaling pathway. Components orchestrating the BR signaling pathway include receptors such as kinases, transcription factors, protein kinases and phosphatases. The proper functioning of the receptor kinase BRI1 and the transcription factors BES1/BZR1 depends on their dephosphorylation by type 2A protein phosphatases (PP2A). In this work, we report that an additional phosphatase family, type one protein phosphatases (PP1), contributes to the regulation of the BR signaling pathway. Co-immunoprecipitation and BiFC experiments performed in Arabidopsis plants overexpressing durum wheat TdPP1 showed that TdPP1 interacts with dephosphorylated BES1, but not with the BRI1 receptor. Higher levels of dephosphorylated, active BES1 were observed in these transgenic lines upon BR treatment, indicating that TdPP1 modifies the BR signaling pathway by activating BES1. Moreover, ectopic expression of durum wheat TdPP1 lead to an enhanced growth of primary roots in comparison to wild-type plants in presence of BR. This phenotype corroborates with a down-regulation of the BR-regulated genes CPD and DWF4. These data suggest a role of PP1 in fine-tuning BR-driven responses, most likely via the control of the phosphorylation status of BES1.

The current study evaluates the role of phenylalanine ammonia-lyase (PAL) and the associated metabolic complex in the accumulation of lignin in common wheat plants ( Tríticum aestívum L.) at the early stages of ontogenesis. The data analysis was performed using plant samples that had reached Phases 4 and 5 on the Feekes scale-these phases are characterized by a transition to the formation of axial (stem) structures in cereal plants. We have shown that the substrate stimulation of PAL with key substrates, such as L-phenylalanine and L-tyrosine, leads to a significant increase in lignin by an average of 20% in experimental plants compared to control plants. In addition, the presence of these compounds in the nutrient medium led to an increase in the number of gene transcripts associated with lignin synthesis ( PAL6 , C4H1 , 4CL1 , C3H1 ). Inhibition was the main tool of the study. Potential competitive inhibitors of PAL were used: the optical isomer of L-phenylalanine-D-phenylalanine-and the hydroxylamine equivalent of phenylalanine-O-Benzylhydroxylamine. As a result, plants incubated on a medium supplemented with O-Benzylhydroxylamine were characterized by reduced PAL activity (almost one third). The lignin content of the cell wall in plants treated with O-Benzylhydroxylamine was almost halved. In contrast, D-phenylalanine did not lead to significant changes in the lignin-associated metabolic complex, and its effect was similar to that of specific substrates.

Studying and understanding the genetic basis of polyphenol oxidases (PPO)-related traits plays a crucial role in genetic improvement of crops. A tetraploid wheat collection (T. turgidum ssp., TWC) was analyzed using the 90K wheat SNP iSelect assay and phenotyped for PPO activity. A total of 21,347 polymorphic SNPs were used to perform genome-wide association analysis (GWA) in TWC and durum wheat sub-groups, detecting 23 and 85 marker-trait associations (MTA). In addition, candidate genes responsible for PPO activity were predicted. Based on the 23 MTAs detected in TWC, two haplotypes associated with low and high PPO activity were identified. Four SNPs were developed and validated providing one reliable marker (IWB75732) for marker assisted selection. The 23 MTAs were used to evaluate the genetic divergence (F ST  > 0.25) between the T. turgidum subspecies, providing new information important for understanding the domestication process of Triticum turgidum ssp. and in particular of ssp. carthlicum., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Polyploidy occurs prevalently and plays an important role during plant speciation and evolution. This phenomenon suggests polyploidy could develop novel features that enable them to adapt wider range of environmental conditions compared with diploid progenitors. Bread wheat (Triticum aestivum L., BBAADD) is a typical allohexaploid species and generally exhibits greater salt tolerance than its tetraploid wheat progenitor (BBAA). However, little is known about the underlying molecular basis and the regulatory pathway of this trait. Here, we show that the histone acetyltransferase TaHAG1 acts as a crucial regulator to strengthen salt tolerance of hexaploid wheat. Salinity-induced TaHAG1 expression was associated with tolerance variation in polyploidy wheat. Overexpression, silencing, and CRISPR-mediated knockout of TaHAG1 validated the role of TaHAG1 in salinity tolerance of wheat. TaHAG1 contributed to salt tolerance by modulating reactive oxygen species (ROS) production and signal specificity. Moreover, TaHAG1 directly targeted a subset of genes that are responsible for hydrogen peroxide production, and enrichment of TaHAG1 triggered increased H3 acetylation and transcriptional upregulation of these loci under salt stress. In addition, we found the salinity-induced TaHAG1-mediated ROS production pathway is involved in salt tolerance difference of wheat accessions with varying ploidy. Our findings provide insight into the molecular mechanism of how an epigenetic regulatory factor facilitates adaptability of polyploidy wheat and highlights this epigenetic modulator as a strategy for salt tolerance breeding in bread wheat., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Small GTP-binding proteins, also known as ROPs (Rho of Plants), are a subfamily of the Ras superfamily of signaling G-proteins and are required for numerous signaling processes, ranging from growth and development to biotic and abiotic signaling. In this study, we cloned and characterized wheat TaRop10, a homolog of Arabidopsis ROP10 and member of the class II ROP, and uncovered a role for TaRop10 in wheat response to Puccinia striiformis f. sp. tritici (Pst). TaRop10 was downregulated by actin depolymerization and was observed to be differentially induced by abiotic stress and the perception of plant hormones. A combination of yeast two-hybrid and bimolecular fluorescence complementation assays revealed that TaRop10 interacted with a h-type thioredoxin (TaTrxh9). Knocking-down of TaRop10 and TaTrxh9 was performed using the BSMV-VIGS (barley stripe mosaic virus-based virus-induced gene silencing) technique and revealed that TaRop10 and TaTrxh9 play a role in the negative regulation of defense signaling in response to Pst infection. In total, the data presented herein further illuminate our understanding of how intact plant cells accommodate fungal infection structures, and furthermore, support the function of TaRop10 and TaTrxh9 in negative modulation of defense signaling in response to stripe rust infection., (Copyright © 2021 Elsevier B.V. All rights reserved.)

In this study, the effect of microwave-ultrasound or/and toughening treatment on the physicochemical, structural properties, and in vitro digestibility of A- and B-type granules isolated from wheat starch were investigated. From the SEM, microwave-ultrasound and toughening treatment (MU-T) led to the appearance of irregular and disrupted structure significantly and an increment in the resistant starch content of A- and B-type granule. Furthermore, the MU-T starch possessed the lowest swelling power, light transmittance, and gelatinization temperature range (T c -T o ) and the highest ΔH. After MU-T, the relative crystallinity (RC) of X-ray pattern, Fourier transform infrared ratio of 1047/1022 cm -1 , and the content of double helix and single helix of 13 C CP/MAS NMR had increased significantly. In particular, there was a difference in the content of RS and SDS between A-starch granules and B-starch granules as well as their changes after modification (from 69.305% to 82.93 for A-starch and form 74.97% to 88.17 for B-starch, respectively), which was a similar trend with RC and helix content. This study indicated that, for both A-type granule and B-type granule starches, microwave-ultrasound and toughening treated samples had unique properties compared to singly modified starches., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Hypergravity-an evolutionarily novel environment has been exploited to comprehend the response of living organisms including plants in the context of extra-terrestrial applications. Recently, researchers have shown that hypergravity induces desired phenotypic variability in seedlings. In the present study, we tested the utility of hypergravity as a novel tool in inducing reliable phenotype/s for potential terrestrial crop improvement applications. To investigate, bread wheat seeds (UAS-375 genotype) were subjected to hypergravity treatment (10×g for 12, and 24 h), and evaluated for seedling vigor and plant growth parameters in both laboratory and greenhouse conditions. It was also attempted to elucidate the associated biochemical and hormonal changes at different stages of vegetative growth. Resultant data revealed that hypergravity treatment (10×g for 12 h) significantly enhanced root length, root volume, and root biomass in response to hypergravity. The robust seedling growth phenotype may be attributed to increased alpha-amylase and TDH enzyme activities observed in seeds treated with hypergravity. Elevated total chlorophyll content and Rubisco (55 kDa) protein expression across different stages of vegetative growth in response to hypergravity may impart physiological benefits to wheat growth. Further, hypergravity elicited robust endogenous phytohormones dynamics in root signifying altered phenotype/s. Collectively, this study for the first time describes the utility of hypergravity as a novel tool in inducing reliable root phenotype that could be potentially exploited for improving wheat varieties for better water usage management., (© 2021. The Author(s).)

Plant pathogens suppress defense responses to evade recognition and promote successful colonization. Although identifying the genes essential for pathogen ingress has traditionally relied on screening mutant populations, the post-genomic era provides an opportunity to develop novel approaches that accelerate identification. Here, RNA-seq analysis of 68 pathogen-infected bread wheat (Triticum aestivum) varieties, including three (Oakley, Solstice and Santiago) with variable levels of susceptibility, uncovered a branched-chain amino acid aminotransferase (termed TaBCAT1) as a positive regulator of wheat rust susceptibility. We show that TaBCAT1 is required for yellow and stem rust infection and likely functions in branched-chain amino acid (BCAA) metabolism, as TaBCAT1 disruption mutants had elevated BCAA levels. TaBCAT1 mutants also exhibited increased levels of salicylic acid (SA) and enhanced expression of associated defense genes, indicating that BCAA regulation, via TaBCAT1, has a key role in SA-dependent defense activation. We also identified an association between the levels of BCAAs and resistance to yellow rust infection in wheat. These findings provide insight into SA-mediated defense responses in wheat and highlight the role of BCAA metabolism in the defense response. Furthermore, TaBCAT1 could be manipulated to potentially provide resistance to two of the most economically damaging diseases of wheat worldwide., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Background: Understanding the determinants of free asparagine concentration in wheat grain is necessary to reduce levels of the processing contaminant acrylamide in baked and toasted wheat products. Although crop management strategies can help reduce asparagine concentrations, breeders have limited options to select for genetic variation underlying this trait. Asparagine synthetase enzymes catalyse a critical step in asparagine biosynthesis in plants and, in wheat, are encoded by five homeologous gene triads that exhibit distinct expression profiles. Within this family, TaASN2 genes are highly expressed during grain development but TaASN-B2 is absent in some varieties., Results: Natural genetic diversity in the asparagine synthetase gene family was assessed in different wheat varieties revealing instances of presence/absence variation and other polymorphisms, including some predicted to affect the function of the encoded protein. The presence and absence of TaASN-B2 was determined across a range of UK and global common wheat varieties and related species, showing that the deletion encompassing this gene was already present in some wild emmer wheat genotypes. Expression profiling confirmed that TaASN2 transcripts were only detectable in the grain, while TaASN3.1 genes were highly expressed during the early stages of grain development. TaASN-A2 was the most highly expressed TaASN2 homeologue in most assayed wheat varieties. TaASN-B2 and TaASN-D2 were expressed at similar, lower levels in varieties possessing TaASN-B2. Expression of TaASN-A2 and TaASN-D2 did not increase to compensate for the absence of TaASN-B2, so total TaASN2 expression was lower in varieties lacking TaASN-B2. Consequently, free asparagine concentrations in field-produced grain were, on average, lower in varieties lacking TaASN-B2, although the effect was lost when free asparagine accumulated to very high concentrations as a result of sulphur deficiency., Conclusions: Selecting wheat genotypes lacking the TaASN-B2 gene may be a simple and rapid way for breeders to reduce free asparagine concentrations in commercial wheat grain.

The effects of argon and nitrogen cold plasma treatments on the lipolytic enzymes activity in wheat germ were investigated. Using argon as plasma gas, the residual activity of lipase and lipoxygenase decreased to 42.50% and 87.72%, respectively after 30 min. Switching plasma input gas to nitrogen, the residual activities of lipase and lipoxygenase after the same time of atmospheric cold plasma (ACP) treatment were 77.50% and 92.52%, respectively. The antioxidant potential and phenolic compounds show no significant difference during ACP duration. However, the remaining activities of lipase and lipoxygenase after 30 min steam autoclaving were 6.25% and 18.60%, respectively. Also, the antioxidant activity and total phenolic content reduced by 14.70% and 30.80%, respectively. In brief, the ACP treatment efficiency was function of the input gas and the treatment time. The presented results about the input gas impacts would be useful in industrial development of ACP application for wheat germ stabilization., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

We expressed a bacterial glucan synthase (Agrobacterium GlgA) in the cytosol of developing endosperm cells in wheat grains, to discover whether it could generate a glucan from cytosolic ADP-glucose. Transgenic lines had high glucan synthase activity during grain filling, but did not accumulate glucan. Instead, grains accumulated very high concentrations of maltose. They had large volumes during development due to high water content, and very shrivelled grains at maturity. Starch synthesis was severely reduced. We propose that cytosolic glucan synthesized by the glucan synthase was immediately hydrolysed to maltose by cytosolic β-amylase(s). Maltose accumulation resulted in a high osmotic potential in developing grain, drawing in excess water that stretched the seed coat and pericarp. Loss of water during grain maturation then led to shrinkage when the grains matured. Maltose accumulation is likely to account for the reduced starch synthesis in transgenic grains, through signalling and toxic effects. Using bioinformatics, we identify an isoform of β-amylase likely to be responsible for maltose accumulation. Removal of this isoform through identification of TILLING mutants or genome editing, combined with co-expression of heterologous glucan synthase and a glucan branching enzyme, may in future enable elevated yields of carbohydrate through simultaneous accumulation of starch and cytosolic glucan., (© 2021 The Authors. The Plant Journal Published by Society for Experimental Biology and John Wiley & Sons Ltd.)

Xyloglucan endotransglycosylase/hydrolase (XTH) enzymes play important roles in cell wall remodelling. Although previous studies have shown a pathway of evolution for XTH genes from bacterial licheninases, through plant endoglucanases (EG16), the order of development within the phylogenetic clades of true XTHs is yet to be elucidated. In addition, recent studies have revealed interesting and potentially useful patterns of transglycosylation beyond the standard xyloglucan-xyloglucan donor/acceptor substrate activities. To study evolutionary relationships and to search for enzymes with useful broad substrate specificities, genes from the 'ancestral' XTH clade of two monocots, Brachypodium distachyon and Triticum aestivum, and two eudicots, Arabidopsis thaliana and Populus tremula, were investigated. Specific activities of the heterologously produced enzymes showed remarkably broad substrate specificities. All the enzymes studied had high activity with the cellulose analogue HEC (hydroxyethyl cellulose) as well as with mixed-link β-glucan as donor substrates, when compared with the standard xyloglucan. Even more surprising was the wide range of acceptor substrates that these enzymes were able to catalyse reactions with, opening a broad range of possible roles for these enzymes, both within plants and in industrial, pharmaceutical and medical fields. Genome screening and expression analyses unexpectedly revealed that genes from this clade were found only in angiosperm genomes and were predominantly or solely expressed in reproductive tissues. We therefore posit that this phylogenetic group is significantly different and should be renamed as the group-IV clade., (© 2021 Society for Experimental Biology and John Wiley & Sons Ltd.)

Sharp eyespot, caused by necrotrophic fungus Rhizoctonia cerealis , is a serious fungal disease in wheat ( Triticum aestivum ). Certain wall-associated receptor kinases (WAK) mediate resistance to diseases caused by biotrophic/hemibiotrophic pathogens in several plant species. Yet, none of wheat WAK genes with positive effect on the innate immune responses to R. cerealis has been reported. In this study, we identified a WAK gene TaWAK7D , located on chromosome 7D, and showed its positive regulatory role in the defense response to R. cerealis infection in wheat. RNA-seq and qRT-PCR analyses showed that TaWAK7D transcript abundance was elevated in wheat after R. cerealis inoculation and the induction in the stem was the highest among the tested organs. Additionally, TaWAK7D transcript levels were significantly elevated by pectin and chitin treatments. The knock-down of TaWAK7D transcript impaired resistance to R. cerealis and repressed the expression of five pathogenesis-related genes in wheat. The green fluorescent protein signal distribution assays indicated that TaWAK7D localized on the plasma membrane in wheat protoplasts. Thus, TaWAK7D, which is induced by R. cerealis , pectin and chitin stimuli, positively participates in defense responses to R. cerealis through modulating the expression of several pathogenesis-related genes in wheat.

Lignocellulose, the structural component of plant cells, is a major agricultural byproduct and the most abundant terrestrial source of biopolymers on Earth. The complex and insoluble nature of lignocellulose limits its conversion into value-added commodities, and currently, efficient transformation requires expensive pretreatments and high loadings of enzymes. Here, we report on a fungus from the Parascedosporium genus, isolated from a wheat-straw composting community, that secretes a large and diverse array of carbohydrate-active enzymes (CAZymes) when grown on lignocellulosic substrates. We describe an oxidase activity that cleaves the major β-ether units in lignin, thereby releasing the flavonoid tricin from monocot lignin and enhancing the digestion of lignocellulose by polysaccharidase mixtures. We show that the enzyme, which holds potential for the biorefining industry, is widely distributed among lignocellulose-degrading fungi from the Sordariomycetes phylum., Competing Interests: Competing interest statement: A patent application has been filed by the University of York on technology described in this research.

Diurnal rhythms and light availability affect transcription-translation feedback loops that regulate the synthesis of photosynthetic proteins. The CO 2 -fixing enzyme Rubisco is the most abundant protein in the leaves of major crop species and its activity depends on interaction with the molecular chaperone Rubisco activase (Rca). In Triticum aestivum L. (wheat), three Rca isoforms are present that differ in their regulatory properties. Here, we tested the hypothesis that the relative abundance of the redox-sensitive and redox-insensitive Rca isoforms could be differentially regulated throughout light-dark diel cycle in wheat. While TaRca1-β expression was consistently negligible throughout the day, transcript levels of both TaRca2-β and TaRca2-α were higher and increased at the start of the day, with peak levels occurring at the middle of the photoperiod. Abundance of TaRca-β protein was maximal 1.5 h after the peak in TaRca2-β expression, but the abundance of TaRca-α remained constant during the entire photoperiod. The redox-sensitive TaRca-α isoform was less abundant, representing 85% of the redox-insensitive TaRca-β at the transcript level and 12.5% at the protein level. Expression of Rubisco large and small subunit genes did not show a consistent pattern throughout the diel cycle, but the abundance of Rubisco decreased by up to 20% during the dark period in fully expanded wheat leaves. These results, combined with a lack of correlation between transcript and protein abundance for both Rca isoforms and Rubisco throughout the entire diel cycle, suggest that the abundance of these photosynthetic enzymes is post-transcriptionally regulated.

Black point is a cereal disease caused by complex pathogens, of which the pathogenicity of Bipolaris sorokiniana is the most serious in wheat. Resistance to black point is quantitative in nature, and thus the mechanism is poorly understood. We conducted a comparative transcriptome analysis to identify differentially expressed genes (DEGs) in black point-slightly susceptible and -highly susceptible wheat lines at different timepoints following B. sorokiniana inoculation. DEGs associated with photosynthesis were upregulated in black point-slightly susceptible lines. The top Gene Ontology enrichment terms for biological processes were oxidation-reduction, response to cold, salt stress, oxidative stress, and cadmium ion; terms for cellular component genes were mainly involved in plasma membrane and cytoplasmic membrane-bounded vesicle, whereas those for molecular function were heme binding and peroxidase activity. Moreover, activities of antioxidant enzymes superoxide dismutase, catalase, and peroxidase were higher in slightly susceptible lines than those in highly susceptible lines (except peroxidase 12-24 days post-inoculation). Thus, resistance to B. sorokiniana-caused black point in wheat was mainly related to counteracting oxidative stress, although the specific metabolic pathways require further study. This study presents new insights for understanding resistance mechanisms of selected wheat lines to black point.

Wheat germ acid phosphatase (WGAP) is a commercial preparation of partially purified protein commonly used in laboratory settings for non-specific enzymatic dephosphorylation. It is known that these preparations contain multiple phosphatase isozymes and are still relatively crude. This study therefore aimed to identify the protein components of a commercial preparation of wheat germ acid phosphatase using mass spectroscopy and comparative genomics. After one post-purchase purification step, the most prevalent fifteen proteins in the mixture included heat shock proteins, beta-amylases, glucoseribitol dehydrogenases, enolases, and an aminopeptidase. While not among the most abundant components, eight unique dephosphorylation enzymes were also present including three purple acid phosphatases. Furthermore, it is shown that some of these correspond to previously isolated isozymes; one of which has been also previously shown by transcriptome data to be overexpressed in wheat seeds. In summary, this study identified the major components of WGAP including phosphatases and hypothesizes the most active components towards a better understanding of this commonly used laboratory tool., Competing Interests: The authors have declared that no competing interests exist.

Crop breeding for resistance to pathogens largely relies on genes encoding receptors that confer race-specific immunity. Here, we report the identification of the wheat Pm4 race-specific resistance gene to powdery mildew. Pm4 encodes a putative chimeric protein of a serine/threonine kinase and multiple C2 domains and transmembrane regions, a unique domain architecture among known resistance proteins. Pm4 undergoes constitutive alternative splicing, generating two isoforms with different protein domain topologies that are both essential for resistance function. Both isoforms interact and localize to the endoplasmatic reticulum when co-expressed. Pm4 reveals additional diversity of immune receptor architecture to be explored for breeding and suggests an endoplasmatic reticulum-based molecular mechanism of Pm4-mediated race-specific resistance.

The current study involves the biogenesis of titanium dioxide nanoparticles (TiO2 NPs) by using Moringa oleifera Lam. aqueous leaf extract for the reduction of titanium dioxide salt into TiO2 nanoparticles. The biosynthesized TiO2 nanoparticles were observed by using the UV-visible spectrophotometry, SEM, EDX and XRD analytical methods. It was confirmed that the nanoparticles are crystalline and exist in the size range of 10-100 nm. The FTIR analysis confirmed the presence of O-H (hydrogen bonding), N-H (amide), C-C (alkanes) and C-I (Iodo-stretch) functional groups responsible for the stabilization of nanoparticles. Various concentrations (20, 40, 60 and 80 mg/L) of TiO2 NPs were applied exogenously on wheat plants infected with a fungus Bipolaris sorokiniana responsible to cause spot blotch disease at different time intervals. The measurement of disease incidence and percent disease index showed the time-dependent response and 40 mg/L was reported a stable concentration of TiO2 NPs to reduce the disease severity. The effects of biosynthesized TiO2 NPs were also evaluated for agro-morphological (leaf and root surface area, plant fresh and dry weight and yield parameters), physiological (relative water content, membrane stability index and chlorophyll content) and non-enzymatic metabolites (soluble sugar, protein, soluble phenol and flavonoid content) in wheat plants under biotic stress and 40 mg/L concentration of TiO2 NPs was found to be effective to elicit modifications to reduce biotic stress. The current study highlights the significant role of biosynthesized TiO2 NPs in controlling fungal diseases of wheat plants and thus ultimately improving the quality and yield of wheat plants., Competing Interests: The authors have declared that no competing interests exist.

Background: Lack of nutritionally appropriate foods is one of the leading causes of obesity in the US and worldwide. Wheat (Triticum aestivum) provides 20% of the calories consumed daily across the globe. The nutrients in the wheat grain come primarily from the starch composed of amylose and amylopectin. Resistant starch content, which is known to have significant human health benefits, can be increased by modifying starch synthesis pathways. Starch synthase enzyme SSIIa, also known as starch granule protein isoform-1 (SGP-1), is integral to the biosynthesis of the branched and readily digestible glucose polymer amylopectin. The goal of this work was to develop a triple null mutant genotype for SSIIa locus in the elite hard red winter wheat variety 'Jagger' and evaluate the effect of the knock-out mutations on resistant starch content in grains with respect to wild type., Results: Knock-out mutations in SSIIa in the three genomes of wheat variety 'Jagger' were identified using TILLING. Subsequently, these loss-of function mutations on A, B, and D genomes were combined by crossing to generate a triple knockout mutant genotype Jag-ssiia-∆ABD. The Jag-ssiia-∆ABD had an amylose content of 35.70% compared to 31.15% in Jagger, leading to ~ 118% increase in resistant starch in the Jag-ssiia-∆ABD genotype of Jagger wheat. The single individual genome mutations also had various effects on starch composition., Conclusions: Our full null Jag-ssiia-∆ABD mutant showed a significant increase in RS without the shriveled grain phenotype seen in other ssiia knockouts in elite wheat cultivars. Moreover, this study shows the potential for developing nutritionally improved foods in a non-GM approach. Since all the mutants have been developed in an elite wheat cultivar, their adoption in production and supply will be feasible in future.