| Wheat is the second largest staple food in China, which is closely related to the food security and people’s living standards. Salt stress is one of the major limiting factors influencing wheat production. It negatively affects wheat quality and quantity through different pathways, e.g. photosynthesis, respiration, traspiration etc. via transcriptional and translational level. Therefore, it it has important theoretical and practical significance to identify key salt-tolerant genes and its protein modifications and study the molecular mechanisms underlying salt tolerance of wheat, which will provide useful information for the further analysis of salt tolerance.To explore the wheat transcriptional response to salt stress, we performed high-throughput transcriptome sequencing of 1-week-old wheat roots under normal and high-salinity conditions at 6,12,24 and 48 h in both a salt-tolerant cultivar (Qing Mai 6, QM) and salt-sensitive cultivar (Chinese Spring, CS), and presented a global gene expression reprograms in response to salt stress. Additionally, to reveal post-translational modifications role in wheat, we studied the range of physiological processes regulated by lysine acetylation and succinylation by using LC-MS/MS analysis and bioinformatics analysis, main results are as follows:1. We identified 36,804 genes that were up-or down-regulated compared with the controls in response to salinity in QM and CS.. Approximately 55.4% and 50.1% of the triplets exhibited biased expression patterns among the homeologs in the untreated QM and CS samples, whereas the proportions increased to 62.6% and 59.9%, respectively after salt stress, and q RT-PCR results confirmed our analysis. The proportion of up-regulated genes was higher in CS than in QM, while down-regulated genes was higher in QM than in CS. In addition, the proportion of up-regulated genes was lower in QM compared to that in CS at 6 and 12 salt stress, but significantly higher at 48 salt stress. In contrast, the proportion of down-regulated genes was higher in CS than in QM at 6 and 12 salt stress, while it was lower at 48 salt stress. The GO (Gene Ontology) enrichment analysis confirmed our observation that in the processes of cell growth, response to ABA stimulus, potassium ion transport, establishment of localization and response to salt stress they were significantly enriched in QM, while in the processes of cell death and jasmonic acid biosynthesis they were significantly enriched in CS.2. Totally,3,718 transcription factor genes (TFs) were identified from wheat genome by using domain searching method, distributing among 51 gene families. Among the genes, 1,583 were differentially expressed under salt stress. Stress response transcription factor genes were unevenly distributed in different family, with seven largest TF families accounted for 47% of stress responsive TF genes, including Myb, bZIP, bHLH, NAC, C2H2, AP2/ERF and WRKY. The differentially expressed TFs could be classified into two groups or 20 clusters based on their expression patterns under salt stress.3. We mapped the salt responsive genes of wheat onto the reference genome released by IWGSC, and found these genes were distributed non-randomly on the chromosomes. In total, 142 gene clusters were identified, with the D genome (57) exhibiting more salt-responsive hotspots than the A (36) and B genomes (49). Further analysis found that the salt responsive genes identified in one cluster encoding similar proteins, which indicated that tandem duplication was one of causes resulting in salt responsive hotspot formation.4. We performed the first global analysis on the overlap between the two types of acylation in wheat, in total,416 lysine acetylated modification sites were identified on 277 proteins and 330 lysine succinylated modification sites were identified on 173 proteins. Altogether, the enrichment analyses based on GO, KEGG pathway and domain GO annotation suggested lysine acetylation and succinylation at proteins associated with energy metabolic process and central carbon metabolism are particularly essential to the physiological functions of wheat. Experimental results also showed that there were 26 proteins involved in photosynthesis and Calvin-Benson cycle happen these two lysine acylations, with 21 succinyl-lysine sites also found to be acetylated at the same position and 33 proteins modified by both acetylation and succinylation, which were accounting for 7.9% of the total succinylated proteins, suggesting extensive overlap between succinylation and acetylation in wheat. Seven enzymes of these 33 proteins were involved in Calvin-Benson cycle, suggesting that both types of modification may play important roles in regulating carbon fixation metabolism process. This systematic analysis provides an important reference for exploring the physiological role of lysine acetylation and succinylation in this cereal crop and likely in all plants. |
