Genomic profiling and expression analysis of the diacylglycerol kinase gene family in heterologous hexaploid wheat

The inositol phospholipid signaling system mediates plant growth, development, and responses to adverse conditions. Diacylglycerol kinase (DGK) is one of the key enzymes in the phosphoinositide-cycle (PI-cycle), which catalyzes the phosphorylation of diacylglycerol (DAG) to form phosphatidic acid (PA). To date, comprehensive genomic and functional analyses of DGKs have not been reported in wheat. In this study, 24 DGK gene family members from the wheat genome (TaDGKs) were identified and analyzed. Each putative protein was found to consist of a DGK catalytic domain and an accessory domain. The analyses of phylogenetic and gene structure analyses revealed that each TaDGK gene could be grouped into clusters I, II, or III. In each phylogenetic subgroup, the TaDGKs demonstrated high conservation of functional domains, for example, of gene structure and amino acid sequences. Four coding sequences were then cloned from Chinese Spring wheat. Expression analysis of these four genes revealed that each had a unique spatial and developmental expression pattern, indicating their functional diversification across wheat growth and development processes. Additionally, TaDGKs were also prominently up-regulated under salt and drought stresses, suggesting their possible roles in dealing with adverse environmental conditions. Further cis-regulatory elements analysis elucidated transcriptional regulation and potential biological functions. These results provide valuable information for understanding the putative functions of DGKs in wheat and support deeper functional analysis of this pivotal gene family. The 24 TaDGKs identified and analyzed in this study provide a strong foundation for further exploration of the biological function and regulatory mechanisms of TaDGKs in response to environmental stimuli.

Discovery of a Promising Susceptibility Factor for Fusarium Head Blight in Wheat

Fusarium head blight (FHB) disease of wheat caused by Fusarium spp. deteriorates both quantity and quality of the crop. Manipulation of susceptibility factors, the genes facilitating disease development in plants, offers a novel and alternative strategy for enhancing FHB resistance in plants. In this study, a major effect susceptibility gene for FHB was identified on the short arm of chromosome 7A (7AS). Nullisomic-tetrasomic lines for homoeologous group-7 of wheat revealed dosage effect of the gene, with tetrasomic 7A being more susceptible than control Chinese Spring wheat, qualifying it as a bonafide susceptibility factor. The gene locus was conserved in six chromosome 7A inter-varietal wheat substitution lines of diverse origin and a tetraploid Triticum dicoccoides genotype. The susceptibility gene was named as SF7ASFHB and mapped on chromosome 7AS to 48.5-50.5 Mb peri-centromeric region between del7AS-3 and del7AS-8. Our results showed that deletion of SF7ASFHB imparts ~ 50-60% type 2 FHB resistance (against the spread of the fungal pathogen) and its manipulation may lead to enhanced resistance against FHB in wheat.HighlightDiscovery and mapping of a conserved susceptibility factor located on the short arm of wheat chromosome 7A whose deletion makes plants resistant to Fusarium Head Blight.

Genomic profiling and expression analysis of the diacylglycerol kinase gene family in heterologous hexaploid wheat

Background The inositol phospholipid signaling system, which is based on the metabolism of phosphoinositide (PI), mediates plant growth, development, and responses to adversity. Diacylglycerol kinase (DGK) is one of the key enzymes in the PI-cycle, which catalyzes the phosphorylation of diacylglycerol (DAG) to form phosphatidic acid (PA). To date, comprehensive genomic and functional analyses of DGK genes have not been reported in wheat. Results In this study, 20 DGK gene family members from the heterologous hexaploid wheat genome (TaDGKs) were identified and analyzed. Each putative protein was found to consist of a DGK catalytic domain and a accessory domain. The analyses of phylogenetic and gene structure revealed that each TaDGK gene could be grouped to clusters I, II, or III. In each phylogenetic subgroup, the TaDGKs demonstrated high conservation in functional domains, for example gene structure and amino acid sequences. By cloning, four coding sequences were ascertained from Chinese spring wheat. Expression analysis of these four genes revealed that each had a unique spatial and developmental expression pattern, indicating their functional diversification in wheat growth and development processes. Additionally, TaDGKs were also prominently up-regulated express under salt and drought stresses, suggesting their possible roles in dealing with adversity environment. Further cis-regulatory elements analysis elucidated transcriptional regulation and potential biological functions. Conclusions These results provide valuable information for understanding the putative functions of DGK genes in wheat, and conduce to ulterior functional analysis of this pivotal gene family. The 20 TaDGKs identified and analyzed in this study provide a strong foundation for further exploration of the biological function and regulatory mechanisms of TaDGKs in response to environmental stimuli.

Mapping of Stripe Rust and Leaf Rust Resistance Quantitative Trait Loci in the Chinese Spring Wheat Line Mianyang351-15

Stripe rust and leaf rust cause wheat yield losses of up to 70% worldwide. The employment of resistant cultivars is the major method to reduce losses from these diseases. The objective of this study was to detect quantitative trait loci (QTL) for stripe rust and leaf rust resistance in 150 F6 recombinant inbred lines (RIL) derived from a cross between Mianyang351-15 and Zhengzhou 5389. Both parents and the RIL population were genotyped with the Wheat55K single nucleotide polymorphism (SNP) array and simple sequence repeat markers, and phenotyped for stripe rust severity at Mianyang in Sichuan Province and Baoding in Hebei Province, and for leaf rust severity at Zhoukou in Henan Province and at Baoding in 2014 to 2017 cropping seasons. Seven and four QTL all contributed from Mianyang351-15 were identified for resistance to stripe rust and leaf rust, respectively. Four of these QTL on chromosomes 1BL, 2AS, 2DS, and 7BL conferred resistance to both stripe rust and leaf rust. The QTL on 1BL, 2AS, and 7BL were identified as Lr46/Yr29, Lr37/Yr17, and Lr68, respectively. QYr.hbau-2DS/QLr.hbau-2DS was detected at similar positions to previously reported loci. QYr.hbau-1DL, QYr.hbau-3AS, and QYr.hbau-3DL are likely to be new. Combined effects of QTL in the RIL population indicated RIL combining all QTL had the highest resistance level compared with those of lower numbers or no QTL. These QTL, with their closely linked SNP markers, are applicable for marker-assisted breeding and candidate gene discovery.

De novo assembly and comparative transcriptome analysis revealed the genes that were potentially involved in defensive terpenoids emission in the wheat against the wheat aphid Sitobion avenae (Fabricius)

Background Terpenoid volatiles play an important role directly or indirectly in the plant defense mechanisms against herbivores, including the gramineous crops such as transgenic rice and corn. The conventional varieties of wheat are important gramineous cereal crops that lack aphid-resistant genes. Therefore, it is necessary to seek the aphid-resistant genes by screening for potential terpenoid synthase genes in the wheat germplasm resources. Results The result showed that aphid-damaged Octoploid Tirtitrigia emitted a higher amount of S-linalool, ent-kaurene, (+)-delta-cadinene, (3S,6E)-nerolidol as compared to the intact plant. In addition, (E)-β-caryophyllene, β‐Myrcene, (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) were new volatile terpenoids emitted by the damaged plant. Further olfactory responses tests showed that S-linalool significantly repelled Sitobion avenae (Fabricius). After de novo assembly and quantitative assessment, a total of 182348 (74.8%) unigenes were annotated by alignment with the public protein databases. Of these unigenes, 2389 differentially expressed genes were identified between the intact and damaged ears of Octoploid Trititrigia. The expression profile of 10 randomly selected TPSs was confirmed with RT-qPCR. Candidate genes involved in terpenes biosynthesis were identified by showing the significant transcript changes between the intact and damaged plant ears of Octoploid Trititrigia. The transcript abundances of terpenes biosynthetic pathway-related genes were also positively correlated with the production of volatile terpenoids in the ears. The unigenes of S-linalool synthase gene was mapped to the cloned cDNA WT008_M07 (AK333728) and WT013_P07 (AK335856) of the Chinese spring wheat cultivar. The predicted protein complete ORF sequence (TaLIS1/2) when compared with the S-linalool synthase gene of other species, contained an aspartate-rich region DDxxD motif. Its function was characterized as coordinating the divalent metal ions involved in substrate binding. Furthermore, the phylogenetic tree results indicated that the TaLIS1 and TuNES1 are highly homologous. Conclusions This assembled transcriptome of S. avenae-damaged Octoploid Trititrigia and the intact ears could provide more molecular resources for the future functional characterization analysis of genomics in volatile terpenoids involved in direct or indirect defenses. Our study describes the metabolic regulation mechanism of the volatile terpenoids in the gramineous crops, which provides support for both breeding and genetic modification of the wheat varieties resistant to wheat aphid.

De novo assembly and comparative transcriptome analysis reveal genes potentially involved in changes of terpenoids in wheat germplasm resource, that defend against the wheat aphid Sitobion avenae (Fabricius)

Background Terpenoid volatiles play an important role in direct and indirect plant defense responses against herbivores, including in gramineous crops such as transgenic rice and corn. The conventional varieties of wheat, an important gramineous cereal crop, lack aphid-resistant genes. It is therefore, necessary to seek aphid-resistant genes by screening for potential terpenoid synthase genes in wheat germplasm resources. Results The result showed that aphid-damaged Octoploid Tirtitrigia emitted a higher amount of S-linalool, ent-kaurene, (+)-delta-cadinene, (3S,6E)- nerolidol compared to intact plant. In addition, (E)-β-caryophyllene, β‐Myrcene, (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT) were new volatile terpenoids emitted by the damaged plant. Further olfactory responses tests showed that S-linalool significantly repelled Sitobion avenae (Fabricius). Using the Illumina sequencing platform, approximately 203.09 million high quality paired-end reads were obtained. After de novo assembly and quantitative assessment, a total of 182,348(74.8%) unigenes were annotated by alignment with public protein databases. Of these unigenes, 2,389 differentially expressed genes were identified between intact and damaged ears of Octoploid Trititrigia. The expression profile of 10 randomly selected TPSs was confirmed with RT-qPCR. Candidate genes involved in terpenes biosynthesis were identified showing significant transcript changes between intact and damaged plant ears of Octoploid Trititrigia. Also transcript abundances of terpenes biosynthetic pathway-related genes were positively correlated with the production of volatile terpenoids in ears. The unigenes of S-linalool synthase gene was mapped to the cloned cDNA WT008_M07 (AK333728) and WT013_P07 (AK335856) of the Chinese spring wheat cultivar. The predicted protein complete ORF sequence (TaLIS1/2) when compared with the S-linalool synthase gene of other species, contained an aspartate-rich region DDxxD motif. Its function was characterized as coordinating the divalent metal ions involved in substrate binding. Furthermore, phylogenetic tree results showed that the TaLIS1 and TuNES1 are highly homologous. Conclusions This assembled transcriptome of S. avenae-damaged Octoploid Trititrigia and intact ears can provide more molecular resources for future functional characterization analysis of genomics in volatile terpenoids involved in direct or indirect defenses. Our study describes the metabolic regulation mechanism of volatile terpenoids in gramineous crops, which provides support for both breeding and genetic modification of wheat varieties resistant to wheat aphids.

Genome-wide identification of and functional insights into the late embryogenesis abundant (LEA) gene family in bread wheat (Triticum aestivum)

Late embryogenesis abundant (LEA) proteins are involved in the responses and adaptation of plants to various abiotic stresses, including dehydration, salinity, high temperature, and cold. Here, we report the first comprehensive survey of the LEA gene family in “Chinese Spring” wheat (Triticum aestivum). A total of 179 TaLEA genes were identified in T. aestivum and classified into eight groups. All TaLEA genes harbored the LEA conserved motif and had few introns. TaLEA genes belonging to the same group exhibited similar gene structures and chromosomal locations. Our results revealed that most TaLEA genes contained abscisic acid (ABA)-responsive elements (ABREs) and various cis-acting elements associated with the stress response in the promoter region and were induced under ABA and abiotic stress treatments. In addition, 8 genes representing each group were introduced into E. coli and yeast to investigate the protective function of TaLEAs under heat and salt stress. TaLEAs enhanced the tolerance of E. coli and yeast to salt and heat, indicating that these proteins have protective functions in host cells under stress conditions. These results increase our understanding of LEA genes and provide robust candidate genes for future functional investigations aimed at improving the stress tolerance of wheat.

The Plastidial Glyceraldehyde-3-Phosphate Dehydrogenase Is Critical for Abiotic Stress Response in Wheat

Plastidial glyceraldehyde-3-phosphate dehydrogenase (GAPDH, GAPCp) are ubiquitous proteins that play pivotal roles in plant metabolism and are involved in stress response. However, the mechanism of GAPCp’s function in plant stress resistance process remains unclear. Here we isolated, identified, and characterized the TaGAPCp1 gene from Chinese Spring wheat for further investigation. Subcellular localization assay indicated that the TaGAPCp1 protein was localized in the plastid of tobacco (Nicotiana tobacum) protoplast. In addition, quantitative real-time PCR (qRT-PCR) unraveled that the expression of TaGAPCp1 (GenBank: MF477938.1) was evidently induced by osmotic stress and abscisic acid (ABA). This experiment also screened its interaction protein, cytochrome b6-f complex iron sulfite subunit (Cyt b6f), from the wheat cDNA library using TaGAPCp1 protein as a bait via the yeast two-hybrid system (Y2H) and the interaction between Cyt b6f and TaGAPCp1 was verified by bimolecular fluorescence complementation assay (BiFC). Moreover, H2O2 could also be used as a signal molecule to participate in the process of Cyt b6f response to abiotic stress. Subsequently, we found that the chlorophyll content in OE-TaGAPCp1 plants was significantly higher than that in wild type (WT) plants. In conclusion, our data revealed that TaGAPCp1 plays an important role in abiotic stress response in wheat and this stress resistance process may be completed by H2O2-mediated ABA signaling pathway.