WRKY Transcription Factors Shared by BTH-Induced Resistance and NPR1-Mediated Acquired Resistance Improve Broad-Spectrum Disease Resistance in Wheat.

In Arabidopsis, both pathogen invasion and benzothiadiazole (BTH) treatment activate the nonexpresser of pathogenesis-related genes 1 (NPR1)-mediated systemic acquired resistance, which provides broad-spectrum disease resistance to secondary pathogen infection. However, the BTH-induced resistance in Triticeae crops of wheat and barley seems to be accomplished through an NPR1-independent pathway. In the current investigation, we applied transcriptome analysis on barley transgenic lines overexpressing wheat wNPR1 (wNPR1-OE) and knocking down barley HvNPR1 (HvNPR1-Kd) to reveal the role of NPR1 during the BTH-induced resistance. Most of the previously designated barley chemical-induced (BCI) genes were upregulated in an NPR1-independent manner, whereas the expression levels of several pathogenesis-related (PR) genes were elevated upon BTH treatment only in wNPR1-OE. Two barley WRKY transcription factors, HvWRKY6 and HvWRKY70, were predicted and further validated as key regulators shared by the BTH-induced resistance and the NPR1-mediated acquired resistance. Wheat transgenic lines overexpressing HvWRKY6 and HvWRKY70 showed different degrees of enhanced resistance to Puccinia striiformis f. sp. tritici pathotype CYR32 and Blumeria graminis f. sp. tritici pathotype E20. In conclusion, the transcriptional changes of BTH-induced resistance in barley were initially profiled, and the identified key regulators would be valuable resources for the genetic improvement of broad-spectrum disease resistance in wheat.[Formula: see text] Copyright © 2020 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Molecular mapping of major QTL conferring resistance to orange wheat blossom midge (Sitodiplosis mosellana) in Chinese wheat varieties with selective populations.

KEY MESSAGE: Two novel midge resistance QTL were mapped to a 4.9-Mb interval on chromosome arm 4AL based on the genetic maps constructed with SNP markers. Orange wheat blossom midge (OWBM) is a devastating insect pest affecting wheat production. In order to detect OWBM resistance genes and quantitative trait loci (QTL) for wheat breeding, two recombinant inbred line (RIL) populations were established and used for molecular mapping. A total of seven QTL were detected on chromosomes 2D, 4A, 4D and 7D, respectively, of which positive alleles were all from the resistant parents except for the QTL on 7D. Two stable QTL (QSm.hbau-4A.2-1 and QSm.hbau-4A.2-2) were detected in both populations with the LOD scores ranging from 5.58 to 29.22 under all three environments, and they explained a combined phenotypic variation of 24.4-44.8%. These two novel QTL were mapped to a 4.9-Mb physical interval. The single-nucleotide polymorphism (SNP) markers AX-109543456, AX-108942696 and AX-110928325 were closely linked to the QTL and could be used for marker-assisted selection (MAS) for OWBM resistance in wheat breeding programs.

Screening for differential expression of genes for resistance to Sitodiplosis mosellana in bread wheat via BSR-seq analysis.

KEY MESSAGE: Five putative candidate genes for OWBM resistance in Chinese winter wheat 'Jimai 24' were identified via BSR-seq and differential expression analyses. Orange wheat blossom midge (OWBM), Sitodiplosis mosellana, is one of the most serious threats to wheat production worldwide. Conventional gene mapping methods to identify genes require significant amounts of financial support and time. Here, bulked segregant RNA-seq (BSR-seq) was applied to profile candidate genes and develop associated markers for OWBM resistance. Previously, we identified a major QTL (QSm.hebau-4A) for OWBM resistance on the long arm of chromosome 4A. In this study, we aimed at screening differentially expressed resistance genes associated with this QTL. Twelve differentially expressed genes (DEGs) were obtained based on BSR-seq and differential expression analyses. Among them, four were confirmed to be associated with OWBM resistance via quantitative reverse transcription PCR, using an additional set of wheat samples subjected to OWBM invasion. One SPI-like gene and one Malectin-like gene were revealed by gene annotation, respectively. Sequencing results confirmed that the four DEGs and the SPI gene had SNP polymorphisms between wheat parents. All these five resistance-related genes for OWBM were located in the same genomic region with QSm.hebau-4A. Furthermore, six new markers developed based on sequences of the five genes were also mapped in the same genomic region using genetic population. These five genes may be the candidate genes for OWBM resistance in Chinese wheat 'Jimai 24' and should be the targets for further positional isolation.

Haynaldia villosa NAM-V1 is linked with the powdery mildew resistance gene Pm21 and contributes to increasing grain protein content in wheat.

BACKGROUND: The 6AL/6VS translocation lines, carrying the wheat powdery mildew resistance gene Pm21, are planted on more than 3.4 million hectares. The NAM-A1 gene, located on chromosome 6AS of hexaploid wheat, has been implicated with increased wheat grain protein content (GPC). However, the NAM-A1 gene was removed from the 6AL/6VS translocation lines after the original chromosome 6AS was replaced by chromosome 6VS of Haynaldia villosa. The present study aimed to clone the NAM homologous gene from chromosome 6VS, to analyze the changes of GPC in the 6AL/6VS translocation lines, and to develop related molecular markers for wheat molecular breeding. RESULTS: A new NAM family gene, NAM-V1, was cloned from 6VS of H. villosa (GenBank ACC. no. KR873101). NAM-V1 contained an intact open reading frame (ORF) and putatively encodes a protein of 407 amino acids. Phylogenetic analysis indicated that NAM-V1 was an orthologous gene of NAM-A1, B1, and D1. The determination of GPC in four Pm21 F2 segregation populations demonstrated that the replacement of NAM-A1 by NAM-V1 confers increased GPC in hexaploid wheat. Multiple sequence alignment of NAM-A1, B1, B2, D1, D2, and V1 showed the single nucleotide polymorphism (SNP) sites for each of the NAM genes, allowing us to develop a molecular marker, CauNAM-V1, for the specific detection of NAM-V1 gene. Our results indicate that CauNAM-V1 can be used as a novel DNA marker for NAM-V1, and can also be used for selecting Pm21 in wheat breeding programs. Further, we developed a marker, CauNAM-ABD, for the amplification and simultaneously distinguish among the NAM-A1, NAM-B1, NAM-B2, NAM-D1, and NAM-D2 genes in a single step. CauNAM-ABD enabled us to develop an efficient "one-marker-for-five-genes" procedure for identifying genes and its copy numbers related with grain protein content. CONCLUSION: Here, we report the isolation of the NAM-V1 gene of H. villosa. This gene contributes to increasing GPC in 6AL/6VS translocation wheat lines. We developed a molecular marker for the specific detection of NAM-V1 and a molecular marker that can be used to simultaneously distinguished among the NAM-A1, NAM-B1, NAM-B2, NAM-D1, and NAM-D2 genes in a single step.

Fine Mapping of the Wheat Leaf Rust Resistance Gene LrLC10 (Lr13) and Validation of Its Co-segregation Markers.

Wheat leaf rust, caused by the fungus Puccinia triticina Eriks. (Pt), is a destructive disease found throughout common wheat production areas worldwide. At its adult stage, wheat cultivar Liaochun10 is resistant to leaf rust and the gene for that resistance has been mapped on chromosome 2BS. It was designated LrLC10 and is the same gene as cataloged gene Lr13 by pedigree analysis and allelism test. We fine-mapped it using recessive class analysis (RCA) of the homozygous susceptible F2 plants derived from crosses using Liaochun10 as the resistant, male parent. Taking advantage of the re-sequencing data of Liaochun10 and its counterpart susceptible parent, we converted nucleotide polymorphisms in the LrLC10 interval between the resistant and susceptible parents into molecular markers to saturate the LrLC10 genetic linkage map. Four indel markers were added in the 1.65 cM map of LrLC10 flanked by markers CAUT163 and Lseq22. Thirty-two recombinants were identified by those two markers from the 984 F2 homozygous susceptible plants and were further genotyped with additional ten markers. LrLC10 was finally placed in a 314.3 kb region on the Chinese Spring reference sequence (RefSeq v1.0) that contains three high confidence genes: TraesCS2B01G182800, TraesCS2B01G182900, and TraesCS2B01G183000. Sequence analysis showed several variations in TraesCS2B01G182800 and TraesCS2B01G183000 between resistant and susceptible parents. One KASP marker and an indel marker were designed based on the differences in those two genes, respectively, and were validated to be diagnostic co-segregating markers for LrLC10. Our results both improve marker-assisted selection and help with the map-based cloning of LrLC10.

Sevoflurane Exerts an Anti-depressive Action by Blocking the HMGB1/TLR4 Pathway in Unpredictable Chronic Mild Stress Rats.

This study was performed to investigate whether sevoflurane has an anti-depressive effect and to elucidate its underlying mechanism. Unpredictable chronic mild stress (uCMS)-treated rats were used for inducing depressive-like behavior and subsequently treated with sevoflurane. A forced swimming test was conducted with the rats. An ELISA was performed to detect the levels of brain-derived neurotrophic factor (BDNF) and inflammatory cytokines in the hippocampus of the rats. Differentially expressed genes in uCMS and normal rats were analyzed by microarray. qRT-PCR, western blot, and flow cytometry, and gain and loss of function measurements were carried out to determine the association between sevoflurane and the HMGB1/TLR4 pathway. A forced swimming test with uCMS rats exposed to sevoflurane demonstrated that a 2% sevoflurane treatment resulted in an anti-depressive effect. In addition, ELISAs of TNF-α (tumor necrosis factor-α), IL-1ß (interleukin-1 beta), IL-6 (interleukin-6), and BDNF supported an effect of sevoflurane on inflammatory cytokines and a neurotrophic factor. HMGB1 was dramatically induced in uCMS rats, and the HMGB1/TLR4 pathway was implicated in sevoflurane exposure. A 2% sevoflurane treatment resulted in a restoration of HMGB1/TLR4 signaling and expression of cytokines and BDNF. HMGB1 overexpression partially prevented the protective effect of 2% SF, suggesting sevoflurane protects uCMS rats.

Simultaneous Transfer of Leaf Rust and Powdery Mildew Resistance Genes from Hexaploid Triticale Cultivar Sorento into Bread Wheat.

Wheat powdery mildew, caused by Blumeria graminis f. sp. tritici, and wheat leaf rust, caused by Puccinia triticina Eriks, are two important diseases that severely threaten wheat production. Sorento, a hexaploid triticale cultivar from Poland, shows high resistance to the wheat powdery mildew isolate E09 and the leaf rust isolate PHT in Beijing, China. To introduce resistance genes into common wheat, Sorento was crossed with wheat line Xuezao, which is susceptible to both diseases, and the F1 hybrids were then backcrossed with Xuezao as the recurrent male parent. By marker analysis, we demonstrate that the long arm of the 2R (2RL) chromosome confers resistance to both the leaf rust and powdery mildew isolates at adult-plant and seedling stages, while the long arm of 4R (4RL) confers resistance only to powdery mildew at both stages. The chromosomal composition of BC2F3 plants containing 2R or 2RL and 4R or 4RL in the form of substitution and translocation were confirmed by GISH (genomic in situ hybridization) and FISH (fluorescence in situ hybridization). Monosomic and disomic substitutions of a wheat chromosome with chromosome 2R or 4R, as well as one 4RS-4DL/4DS-4RL reciprocal translocation homozigote and one 2RL-1DL translocation hemizigote, were recovered. Such germplasms are of great value in wheat improvement.

Application of Glycerol for Induced Powdery Mildew Resistance in Triticum aestivum L.

Previous work has demonstrated that glycerol-3-phosphate (G3P) and oleic acid (18:1) are two important signal molecules associated with plant resistance to fungi. In this article, we provide evidence that a 3% glycerol spray application 1-2 days before powdery mildew infection and subsequent applications once every 4 days was sufficient to stimulate the plant defense responses without causing any significant damage to wheat leaves. We found that G3P and oleic acid levels were markedly induced by powdery mildew infection. In addition, TaGLI1 (encoding a glycerol kinase) and TaSSI2 (encoding a stearoylacyl carrier protein fatty acid desaturase), two genes associated with the glycerol and fatty acid (FA) pathways, respectively, were induced by powdery mildew infection, and their promoter regions contain some fungal response elements. Moreover, exogenous application of glycerol increased the G3P level and decreased the level of oleic acid (18:1). Glycerol application induced the expression of pathogenesis-related (PR) genes (TaPR-1, TaPR-2, TaPR-3, TaPR-4, and TaPR-5), induced the generation of reactive oxygen species (ROS) before powdery mildew infection, and induced salicylic acid (SA) accumulation in wheat leaves. Further, we sprayed glycerol in a wheat field and found that it significantly (p < 0.05) reduced the severity of powdery mildew disease and lessened disease-associated kernel weight loss, all without causing any noticeable degradation in wheat seed quality.