Potassium (K) is essential to plant growth and development. Foxtail millet (Setaria italic L.) is an important fodder grain crop in arid and semi-arid regions of Asia and Africa because of its strong tolerance to drought and barren stresses. The molecular mechanisms of physiological and biochemical responses and regulations to various abiotic stresses such as low potassium conditions in foxtail millet are not fully understood, which hinders the research and exploitation of this valuable resource. In this research, we demonstrated that the millet variety Longgu 25 was the most insensitive variety to low potassium stress among other five varieties. The transcriptome analysis of Longgu 25 variety revealed a total of 26,192 and 26,849 genes from the K+-deficient and normal transcriptomic libraries by RNA-seq, respectively. A total of 1982 differentially expressed genes (DEGs) were identified including 866 up-regulated genes and 1116 down-regulated genes. We conducted a comparative analysis of these DEGs under low-K+ stress conditions and discovered 248 common DEGs for potassium deprivation among foxtail millet, rice and Arabidopsis. Further Gene Ontology (GO) enrichment analysis identified a series of candidate genes that may involve in K+-deficient response and in intersection of molecular functions among foxtail millet, rice and Arabidopsis. The expression profiles of randomly selected 18 candidate genes were confirmed as true DEGs with RT-qPCR. Furthermore, one of the 18 DEGs, SiMYB3, is specifically expressed only in the millet under low-K+ stress conditions. Overexpression of SiMYB3 promoted the main root elongation and improved K+ deficiency tolerance in transgenic Arabidopsis plants. The fresh weight of the transgenic plants was higher, the primary root length was longer and the root surface-area was larger than those of control plants after K+ deficiency treatments. This study provides a global view of transcriptomic resources relevant to the K+-deficient tolerance in foxtail millet, and shows that SiMYB3 is a valuable genetic resource for the improvement of K+ deficiency tolerance in foxtail millet.

Wheat Avenin-like proteins (TaALP) are atypical storage proteins belonging to the Prolamin superfamily. Previous studies on ALPs have focused on the proteins’ positive effects on dough strength, whilst no correlation has been made between TaALPs and the plant immune system. Here, we performed genome-wide characterization of ALP encoding genes in bread wheat. In silico analyses indicated the presence of critical peptides in TaALPs that are active in the plant immune system. Pathogenesis-related nucleotide motifs were also identified in the putative promoter regions of TaALP encoding genes. RT-PCR was performed on TaALP and previously characterised pathogenesis resistance genes in developing wheat caryopses under control and Fusarium graminearum infection conditions. The results showed that TaALP and NMT genes were upregulated upon F. graminearum inoculation. mRNA insitu hybridization showed that TaALP genes were expressed in the embryo, aleurone and sub-aleurone layer cells. Seven TaALP genes were cloned for the expression of recombinant proteins in Escherichia coli , which displayed significant inhibitory function on F. graminearum under anti-fungal tests. In addition, FHB index association analyses showed that allelic variations of two ALP genes on chromosome 7A were significantly correlated with FHB symptoms. Over-expression of an ALP gene on chromosome 7A showed an enhanced resistance to FHB. Yeast two Hybridization results revealed that ALPs have potential proteases inhibiting effect on metacaspases and beta-glucosidases. A vital infection process related pathogen protein, F. graminearum Beta-glucosidase was found to interact with ALPs. Our study is the first to report a class of wheat storage protein or gluten protein with biochemical functions. Due to its abundance in the grain and the important multi-functions, the results obtained in the current study are expected to have a significant impact on wheat research and industry.

Nuclear proteins play critical roles in regulating mRNA transcription and processing, DNA replication, and epigenetic genome modification. Therefore, the ability to monitor changes in nuclear proteins is helpful not only to identify important regulatory proteins but also to study the mechanisms of actions of nuclear proteins. However, no effective methods have been developed yet. Rye is strongly resistant to various biotic and abiotic stresses; however, few genes have been functionally characterized to date due to the complexity of its genome and a lack of genomic sequence information. We developed an integrative Nuclear Transportation Trap (iNTT) system that includes an improved nuclear transportation trap and utilizes the “after suppression subtraction” method. Oligonucleotides encoding a nuclear localization signal (NLS) or a transcription factor, GmAREB, were inserted into pLexAD or pLexAD-NES, respectively, and then transformed into yeast cells (EGY48). We showed that the pLexAD vector expressing a cDNA library in the iNTT system was more efficient for screening than the vector pLexAD-NES, which has previously been used in an NTT system. We used the iNTT system to screen a cDNA library of cold-treated rye. A total of 241 unique genes were identified, including 169 differentially expressed proteins; of these, 106 were of known and 63 were of unknown function. Moreover, 82 genes (49 %) among the 169 differentially expressed genes were predicted to contain an NLS domain. Thirty-three (31 %) of the 106 functionally known proteins have DNA-binding activity. To test the specificity of the nuclear proteins identified using the iNTT screen, four of the proteins differentially expressed in response to temperature stress, ScT1 (a heat shock protein), ScT36 (a MYB-like transcription factor), ScT133 (an ERF-like transcription factor) and ScT196 (a protein of unknown function), were studied in more depth. These proteins were shown to exclusively localize to the nucleus, and their expression levels were increased in response to low-temperature stress. To identify the function of these screened nuclear proteins, ScT1- and ScT36-transgenic Arabidopsis plants were constructed, and ScT1 or ScT36 overexpression was found to enhance tolerance to high-temperature or freezing stresses, respectively. The newly developed iNTT system provides an effective method for identifying nuclear-targeted proteins and monitoring induced expression levels. ScT1 and ScT36 might be good candidate genes for improving the stress tolerance of plants by genetic transformation.

Powdery mildew is a severe wheat disease that causes substantial yield losses in wheat production worldwide. The Chinese wheat landrace Changanhongmai (CAHM) exhibits high resistance to the physiological race E09 of powdery mildew. In this study, we characterized the powdery mildew resistance gene in CAHM, and developed molecular markers for wheat marker-assisted selection. To investigate the genetic characteristics of this resistant gene, we developed F₁ plants, F₂ generation population, and F_2:3 families by crossing CAHM with SY225 (Shaanyou ‘225’ as susceptible male parent). Genetic analysis demonstrated that all F₁ plants were resistant to the disease, while the ratio of resistant to susceptible plants was 3: 1 in both the F₂ population and F_2:3 families, indicating that CAHM is inherited in a manner of a single dominant powdery mildew resistance gene, which was tentatively designated as PmCAHM. By using bulk segregation analysis, we constructed a genetic map encompassing Xgwm273, Xwmc626, Xgwm11, Xgwm18, Xgdm28, Xgpw7812, Xgpw5195, Xwmc694, and PmCAHM. Among these markers, Xgpw7812 and Xgpw5195 are flanking markers that are tightly linked to PmCAHM at a genetic distance of 2.5 cM and 8.4 cM, respectively. Furthermore, nullisomic-tetrasomic analysis revealed that PmCAHM is located on chromosome 1B. These results indicate that PmCAHM differs from the internationally recognized powdery mildew resistance genes in both location and source. In addition, a new germplasm/line NW1748 with stronger powdery mildew resistance and large grains was developed from the cross and backcross populations of Fengyou1718 (FY1718)/CAHM/5/FY 1718. Therefore, PmCAHM can serve as a novel powdery mildew resistance source for breeding of wheat by using NW1748 as the donor in the future.

Powdery mildew is a severe disease causing substantial yield losses in wheat production worldwide. The Chinese wheat landrace Changanhongmai (CAHM) exhibits high resistance to the physiological race E09 of powdery mildew. This study aims to characterize the powdery mildew resistance gene in CAHM, and develop molecular markers for marker-assisted selection of wheat. To investigate the genetic characteristics of this resistant gene, we developed F1 plants, F2 generation population, and F2:3 families from a cross between CAHM and SY225. Genetic analysis demonstrated that all F1 plants were resistant to the disease, while the ratio of resistant to susceptible plants was 3: 1 in both the F2 population and F2:3 families, indicating that CAHM is inherited in a manner of single dominant powdery mildew resistance gene, which was tentatively designated as PmCAHM. By using bulk segregation analysis, we constructed a genetic map encompassing Xgwm273, Xwmc626, Xgwm11, Xgwm18, Xgdm28, Xgpw7812, Xgpw5195, Xwmc694 and PmCAHM. Among these markers, Xgpw7812 and Xgpw5195 were flanking markers tightly linked to PmCAHM at a genetic distance of 2.5 cM and 8.4 cM, respectively. Furthermore, nullisomic-tetrasomic analysis revealed that PmCAHM is located on chromosome 1B. These results indicate that PmCAHM differs from the internationally recognized powdery mildew resistance genes in both location and source. In addition, a new germplasm/line NW1748 with large seeds and powdery mildew resistance was developed from the cross and back cross populations of Fengyou 1718 (FY1718)/CAHM//FY 1718. Therefore, PmCAHM can serve as a novel powdery mildew resistance source for breeding of wheat by using NW1748 as the donor in the future.