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The WRKY transcription factor family has been associated with a variety of plant biological processes, such as biotic and abiotic stress responses. In this study, 13 wheat TaWRKY DEGs in transcriptome data before and after drought stress, namely TaWRKY1 to TaWRKY8, including various copies, were identified and classified as Group I, II, or III. TaWRKY1-2D overexpression enhanced drought tolerance in transgenic Arabidopsis. Moreover, the AtRD29A, AtP5CS1, AtPOD1, AtCAT1, and AtSOD (Cu/Zn) genes, which are related to the stress response and antioxidant system, were significantly upregulated in TaWRKY1-2D transgenic Arabidopsis under drought stress. TaWRKY1-2 silencing in wheat increases the MDA content, reduces the contents of proline and chlorophyll and the activities of antioxidant enzymes, and inhibits the expression levels of antioxidant (TaPOD, TaCAT, and TaSOD (Fe))- and stress-related genes (TaP5CS) under drought stress. Yeast two-hybrid screening revealed TaDHN3 as an interaction partner of TaWRKY1-2D; their interaction was further confirmed using yeast two-hybrid and bimolecular fluorescence complementation. Furthermore, TaWRKY1-2D may play essential roles in wheat drought tolerance through posttranslational regulation of TaDHN3. Overall, these findings contribute to our knowledge of the WRKY family in wheat and identify TaWRKY1-2D as a promising candidate gene for improving wheat breeding to generate drought-tolerant wheat., 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 Elsevier B.V. All rights reserved.)

AP2/ERF (APETALA2/ethylene responsive factor) is a family of plant-specific transcription factors whose members are widely involved in many biological processes, such as growth, development, and biotic and abiotic stress responses. Here, 20 AP2/ERF genes were identified based on wheat RNA-seq data before and after drought stress, and classified as AP2, ERF, DREB, and RAV. The analysis of gene structure revealed that about 85% of AP2/ERF family members had lost introns, which are presumed to have been lost during the formation and evolution of the wheat genome. The expression of 20 AP2/ERF family genes could be verified by qRT-PCR, which further supported the validity of the RNA-seq data. Subsequently, subcellular localization and transcriptional activity experiments showed that the ERF proteins were mainly located in the nucleus and were self-activating, which further supports their functions as transcription factors. Furthermore, we isolated a novel ERF gene induced by drought, salt, and cold stresses and named it TaERF-6-3A . TaERF-6-3A overexpression increased sensitivity to drought and salt stresses in Arabidopsis , which was supported by physiological and biochemical indices. Moreover, the expression of stress- and antioxidant-related genes was downregulated in TaERF-6-3A -overexpressing plants. Overall, these results contribute to the further understanding of the TaERF-6-3A gene function in wheat.

Flowering time is one of the most important agronomic traits in wheat production. A proper flowering time might contribute to the reduction or avoidance of biotic and abiotic stresses, adjust plant architecture, and affect the yield and quality of grain. In this study, TaTOE1-B1 in wheat produced three transcripts ( TaTOE1-B1-1 , TaTOE1-B1-2 , and TaTOE1-B1-3 ) by alternative splicing. Compared to the longest transcript, TaTOE1-B1-1 , TaTOE1-B1-3 has a deletion in the sixth exon (1219-1264 bp). Under long-day conditions, the heterologous overexpression of the TaTOE1-B1-3 gene delayed flowering, prolonged the vegetative growth time, and enlarged the vegetative body of Arabidopsis , but that of TaTOE1-B1-1 did not. As typical AP2 family members, TaTOE1-B1-1 and TaTOE1-B1-3 are mainly located in the nucleus and have transcriptional activation activities; the transcriptional activation region of TaTOE1-B1-3 is located in the C-terminal. In TaTOE1-B1-3 overexpression lines, the expression of flowering-related AtFT and AtSOC1 genes is significantly downregulated. In addition, this study confirms the protein-protein interaction between TaTOE1-B1-3 and TaPIFI, which may play an important role in flowering inhibition. These results provide a theoretical basis for the precise regulation of wheat flowering time.

Background: Cloning and characterizing the drought-inducible promoters is essential for their use in crop resistance's genetic improvement. Previous studies have shown that the TaNRX1-D gene participates in regulating the response of wheat to drought stress. However, its promoter has not yet been identified., Objective: In this study, we aimed to characterize the promoter of the TaNRX1-D gene., Methods: The promoter of TaNRX1-D (named P0, 2081 bp) was isolated from common wheat with several cis-acting elements that regulate in response to abiotic stresses and some core cis-acting elements. Functional verification of the promoter, eight 5'-deletion fragments of TaNRX1-D promoter, was fused to the β-glucuronidase (GUS) gene P0::GUS ~ P7::GUS and transformed into Arabidopsis, respectively. Agrobacterium-mediated GUS transient assay the P6a and P6b promoter regions in tobacco leaves under normal, osmotic or ABA stress., Results: Activity analysis of the full-length promoter (P0) showed that the intensity of stronger β-glucuronidase (GUS) staining in the roots and leaves was obtained during the growth of transgenic Arabidopsis. P0::GUS displayed the GUS activity was much higher in the roots and leaves than in other parts of the transgenic plant under normal conditions, which was similarly within wheat. Analysis of the 5'-deletion fragments revealed that P0::GUS ~ P6::GUS responded well upon exposure to osmotic (polyethylene glycol-6000, PEG6000) and abscisic acid (ABA) stress treatments and expressed significantly higher GUS activity than the CaMV35S promoter (35S::GUS), while P7::GUS did not. GUS transient assay in tobacco leaves showed that the GUS activities of P6a and P6b were lower than P6 in the PEG6000 and ABA stresses., Conclusion: The 193 bp (P6) segment was considered the core region of TaNRX1-D responding to PEG6000 or ABA treatment. GUS activity assay in transgenic Arabidopsis showed that this segment was sufficient for the PEG6000 or ABA stress response. The identified 193 bp promoter of TaNRX1-D in this study will help breed osmotic or ABA tolerant crops. The 36 bp segment between P6 and P6b (-193 to -157 bp) was considered the critical sequence for the TaNRX1-D gene responding to PEG6000 or ABA treatment., (© 2021. The Genetics Society of Korea.)

Osmotic stresses such as drought, salinity, and cold are major environmental factors that limit agricultural productivity. Transcription factors play essential roles in abiotic stress signaling in plants. Three TaMYB2 members were identified and designated TaMYB2A, TaMYB2B, and TaMYB2D based on their genomic origins. The cis-regulatory elements in the promoter regions were compared, and their diverse expression patterns under different abiotic stress conditions were identified. TaMYB2A was further characterized because of its earlier response to stresses. Subcellular localization revealed that TaMYB2A localized in the nucleus. To examine the role of TaMYB2A under various environmental stresses, transgenic Arabidopsis plants carrying TaMYB2A controlled by the CaMV 35S promoter were generated and subjected to severe abiotic stress. TaMYB2A transgenics had enhanced tolerance to drought, salt, and freezing stresses, which were confirmed by the enhanced expressions of abiotic stress-responsive genes and several physiological indices, including decreased rate of water loss, enhanced cell membrane stability, improved photosynthetic potential, and reduced osmotic potential. TaMYB2A is a multifunctional regulatory factor. Its overexpression confers enhanced tolerance to multiple abiotic stresses while having no obvious negative effects on phenotype under well-watered and stressed conditions; thus, TaMYB2A has the potential for utilization in transgenic breeding to improve abiotic stress tolerances in crops.