Drought, a major consequence of climate change, initiates molecular interactions among genes, proteins, and metabolites. Agropyron mongolicum a high-quality perennial grass species, exhibits robust drought resistance. However, the molecular mechanism underlying this resistance remaining largely unexplored. In this study, we performed an integrated analysis of the transcriptome, proteome, and metabolome of A. mongolicum under optimal and drought stress conditions. This combined analysis highlighted the pivotal role of transporters in responding to drought stress. Moreover, metabolite profiling indicated that arginine and proline metabolism, as well as the pentose phosphate pathway, are significantly involved in the drought response of A. mongolicum. Additionally, the integrated analysis suggested that proline metabolism and the pentose phosphate pathway are key elements of the drought resistance strategy in A. mongolicum plants. In summary, our research elucidates the drought adaptation mechanisms of A. mongolicum and identifies potential candidate genes for further study.

The improvement of crop yield and quality is an eternal theme to face climate change and population growth. The key to improving crop varieties lies in precisely manipulating gene expression. Recent advancements in CRISPR/Cas9 technology have made gene knockout increasingly straightforward, yet for genes related to important agronomic traits, it is crucial to regulate their expression levels appropriately. Complete knockout often results in defects in other aspects. In addition, many agronomic traits require upregulation of target gene expression for their improvement. Therefore, the development of novel methods for precise upregulation or downregulation of gene expression, without altering gene protein sequences or introducing new genome fragments, will significantly bolster the technical foundation for crop genetic improvement.N 6 -methyladenosine (m 6 A) is the most abundant and reversible internal chemical modification in eukaryotic mRNA, which is installed, removed, and recognized by methyltransferases (writers), demethylases (erasers), and m 6 A-binding proteins (readers), respectively (Tang et al., 2023). Currently, two types of m 6 A methyltransferases have been identified in plants: multiprotein complexes and a single protein. The multiprotein complex includes MTA, MTB, FIP37, VIRILIZER (VIR), HAKAI, and HIZ2 (HAKAI interacting zinc finger protein 2), which catalyze the majority of m 6 A modifications on mRNA (Parker et al., 2021;Ruzicka et al., 2017;Shen et al., 2016;Zhang et al., 2022;Zhong et al., 2008). The single protein FIONA1 also exhibits methyltransferase activity in Arabidopsis (Wang et al., 2022;Xu et al., 2022), catalyzing approximately 10% of m 6 A modifications on mRNA. Several m 6 A demethylases, which belong to the Fe (II)/α -kg dependent dioxygenase superfamily, have been identified in plants, including Arabidopsis AtALKBH10B and AtALKBH9B, rice OsALKBH9, and tomato SlALKBH2. m 6 A is recognized by m 6 Abinding proteins, such as ECTs in Arabidopsis, which contains the YTH domain. In plants, the ratio of m 6 A/A in poly A + RNA varied among different tissues, with a range of 0.36-0.75% in Arabidopsis and 0.52-0.67% in rice, suggesting its high abundance (Wang et al., 2024). At the transcriptome level, m 6 A sites are primarily enriched within the 3′-untranslated region (3′ UTR), followed by the coding DNA sequence (CDS) and 5′-untranslated region (5′UTR). Recent studies have demonstrated the crucial roles of m 6 A in regulating gene expression in plants, primarily by influencing mRNA stability, translation, and 3′UTR processing (Tang et al., 2023). Among them, mRNA stability regulation is one of the primary functions of m 6 A, which involves two aspects: acceleration of RNA decay or preservation of RNA stability, depending on the specific m 6 A-binding proteins. Recent studies combining proteomics and m 6 A analysis have shown that m 6 A in the untranslated regions is negatively correlated with protein abundance, suggesting that m 6 A in UTR is likely to inhibit protein abundance in plants. Therefore, manipulating m 6 A could lead to an increase in protein abundance (Li et al., 2024). m 6 A modifications have also been found to play crucial roles in plant biology, such as embryo development, floral transition, stem cell fate determination, pollen development, fruit ripening, photomorphogenesis, circadian clock, nitrate signaling, and responses to biotic and abiotic stress (Tang et al., 2023). Given the high abundance and crucial roles of m 6 A modifications in mRNA, altering m 6 A modifications in genes holds promise as a strategy for enhancing crop agronomic traits.In mammals, the m 6 A demethylase FTO, known as an obesity gene, plays a crucial role in regulating body weight. Researchers have genetically engineered rice and potatoes to express FTO. In field experiments, the yield and biomass of genetically engineered rice and potatoes increased by approximately 50% (Yu et al., 2021).Further research indicates that the expression of FTO in rice promotes root growth, tiller bud formation, photosynthetic efficiency, and drought resistance, and these phenotypes are dependent on the m 6 A demethylase activity of FTO (Yu et al., 2021).In strawberries, inhibiting the expression of m 6 A methyltransferase genes FveMTA or FveMTB can delay fruit ripening, while upregulating the expression of either gene can accelerate fruit ripening, suggesting that manipulating m 6 A modification can regulate the ripening time of strawberries. (Zhou et al., 2021). The above research suggests that manipulating the m 6 A modification level overall can enhance the agronomic traits of crops.In addition to altering the overall m 6 A modification level, researchers have also endeavored to edit m 6 A modification levels on individual genes. In plants, a precise editing system for plant mRNA m 6 A has been successfully developed by fusing dCas13a with the plant m 6 A methyltransferase core complex MTA-MTB or the mammalian demethylase ALKBH5 (Shi et al., 2024). By specifically editing the m 6 A modification on the SHR transcript, which is a key gene for root development, it was found that an increase in the m 6 A modification level of the SHR transcript can promote the enlargement of the aboveground and root parts of plants, increase leaf area, plant height, biomass, and grain yield, thereby promoting plant growth (Shi et al., 2024). In cotton, similar m 6 A editing tools have also been developed, combining CRISPR/dCas13(Rx) with the methyltransferase GhMTA (Targeted RNA Methylation Editor, TME) or the demethyltransferase GhALKBH10 (Targeted RNA Demethylation Editor, TDE) (Yu et al., 2024). Using TME editor, the m 6 A level of GhDi19 transcript increased, and the plants with increased m 6 A levels results in a significant increase in root length and enhanced drought resistance. Both works indicated that manipulating the m 6 A modification level of key genes...

Crops frequently endure diverse environmental stresses that negatively impact their growth and yield. Understanding the mechanisms underlying stress responses is crucial for developing stress-tolerant crop varieties, thereby improving their adaptability and productivity. N⁶-methyladenosine (m⁶A) is the most prevalent and reversible internal modification found in eukaryotic mRNA, and is installed, removed, and recognized by methyltransferases (writers), demethylases (erasers), and m⁶A-binding proteins (readers), playing crucial roles in regulating plant growth, development, and environmental stress response. However, the role of m⁶A in alfalfa remains largely unexplored. In this work, we conducted comprehensive whole genome identification and expression analysis of the m⁶A gene family in alfalfa under abiotic stress conditions. There are 11 writers, 6 erasers, and 9 readers were identified in the alfalfa genome. Through comparative genomic analysis, we uncovered the evolutionary relationships of the m⁶A gene family across different species. Additionally, we characterized the gene structures, conserved motifs, cis-regulatory elements, chromosomal localization, and expression patterns specific to tissues and under various abiotic stress treatments. Furthermore, we conducted a transcriptome-wide analysis of m⁶A methylome under drought stress. We identified 10,855 hyper-methylated peaks in shoots and 3,262 in roots, along with a global increase in m⁶A levels during drought stress, individually highlighting the significant role of m⁶A modification in drought stress responses. Notably, we found that ABA signaling-related genes, such as ABA2, SnRK2.1, SnRK2.3, and SnRK2.4, exhibited m⁶A hypermethylation in shoots after drought stress treatment. Furthermore, we demonstrated that m⁶A modifications of ABA2, SnRK2.1, SnRK2.3, and SnRK2.4 enhance their mRNA stability, suggesting that m⁶A modification may regulate drought stress responses through the ABA signaling pathway. Above all, these findings provide new insights into the adaptive mechanism of alfalfa under drought stress and highlight the potential of regulating m⁶A modifications as a novel approach for alfalfa stress resistance breeding strategies.

The plant respiratory burst oxidase homologs (RBOHs) are crucial enzymes responsible for the production of reactive oxygen species (ROS) in plants, playing a pivotal role in regulating various aspects of plant growth, development, and stress responses. While RBOH family members have been identified across a wide range of plant species, the functions and characteristics of the RBOH gene family in oats remain poorly understood. In this study, 35 members of the RBOH gene family in the oat genome were identified using bioinformatics approaches. Conserved motif and gene structure analyses revealed that most AsRBOH genes contain Motif4 and Motif5. Phylogenetic tree analysis demonstrated that the AsRBOHs can be classified into five distinct subfamilies. Synteny analysis indicated that AsRBOHs share the highest number of syntenic gene pairs with wheat. Additionally, cis-regulatory element analysis identified several elements associated with drought and hypoxia-specific responses in AsRBOHs. Expression analysis using qRT-PCR showed that 28 AsRBOH genes were upregulated under drought stress, while 18 were downregulated under salt stress. Notably, the genes 7DG1382190 and 7AG1225850 were found to be involved in both drought and salt stress responses. In conclusion, these findings provide a valuable foundation for future functional studies of the AsRBOH gene family in oats, offering insights that could contribute to the improvement and innovation of oat varieties and germplasm.