The synthesis of triacylglycerol by diacylglycerol acyltransferases (CsDGAT1A and CsDGAT2D) is essential for tolerance of cucumber's resistance to low-temperature stress.

KEY MESSAGE: CsDGAT1A and CsDGAT2D play a positive regulatory role in cucumber's response to low-temperature stress and positively regulate the synthesis of triacylglycerol (TAG). Triacylglycerol (TAG), a highly abundant and significant organic compound in plants, plays crucial roles in plant growth, development, and stress responses. The final acetylation step of TAG synthesis is catalyzed by diacylglycerol acyltransferases (DGATs). However, the involvement of DGATs in cucumber's low-temperature stress response remains unexplored. This study focused on two DGAT genes, CsDGAT1A and CsDGAT2D, investigating their function in enhancing cucumber's low-temperature stress tolerance. Our results revealed that both proteins were the members of the diacylglycerol acyltransferase family and were predominantly localized in the endoplasmic reticulum. Functional analysis demonstrated that transient silencing of CsDGAT1A and CsDGAT2D significantly compromised cucumber's low-temperature stress tolerance, whereas transient overexpression enhanced it. Furthermore, the TAG content quantification indicated that CsDGAT1A and CsDGAT2D promoted TAG accumulation. In conclusion, this study elucidates the lipid metabolism mechanism in cucumber's low-temperature stress response and offers valuable insights for the cultivation of cold-tolerant cucumber plants.

Nucleotide-Binding Leucine-Rich Repeat Genes CsRSF1 and CsRSF2 Are Positive Modulators in the Cucumis sativus Defense Response to Sphaerotheca fuliginea.

Cucumber powdery mildew caused by Sphaerotheca fuliginea is a leaf disease that seriously affects cucumber's yield and quality. This study aimed to report two nucleotide-binding site-leucine-rich repeats (NBS-LRR) genes CsRSF1 and CsRSF2, which participated in regulating the resistance of cucumber to S. fuliginea. The subcellular localization showed that the CsRSF1 protein was localized in the nucleus, cytoplasm, and cell membrane, while the CsRSF2 protein was localized in the cell membrane and cytoplasm. In addition, the transcript levels of CsRSF1 and CsRSF2 were different between resistant and susceptible cultivars after treatment with exogenous substances, such as abscisic acid (ABA), methyl jasmonate (MeJA), salicylic acid (SA), ethephon (ETH), gibberellin (GA) and hydrogen peroxide (H2O2). The expression analysis showed that the transcript levels of CsRSF1 and CsRSF2 were correlated with plant defense response against S. fuliginea. Moreover, the silencing of CsRSF1 and CsRSF2 impaired host resistance to S. fuliginea, but CsRSF1 and CsRSF2 overexpression improved resistance to S. fuliginea in cucumber. These results showed that CsRSF1 and CsRSF2 genes positively contributed to the resistance of cucumber to S. fuliginea. At the same time, CsRSF1 and CsRSF2 genes could also regulate the expression of defense-related genes. The findings of this study might help enhance the resistance of cucumber to S. fuliginea.

ThDof1.4 and ThZFP1 constitute a transcriptional regulatory cascade involved in salt or osmotic stress in Tamarix hispida.

Identification of the upstream regulators of a gene is important to characterize the transcriptional pathway and the function of the gene. Previously, we found that a zinc finger protein (ThZFP1) is involved in abiotic stress tolerance of Tamarix hispida. In the present study, we further investigated the transcriptional pathway of ThZFP1. Dof motifs are abundant in the ThZFP1 promoter; therefore, we used them to screen for transcriptional regulators of ThZFP1. A Dof protein, ThDof1.4, binds to the Dof motif specifically, and was hypothesized as the upstream regulator of ThZFP1. Further study showed that overexpression of ThDof1.4 in T. hispida activated the expression of GUS controlled by the ThZFP1 promoter. In T. hispida, transient overexpression of ThDof1.4 increased the transcripts of ThZFP1; conversely, transient RNAi-silencing of ThDof1.4 reduced the expression of ThZFP1. Chromatin immunoprecipitation indicated that ThDof1.4 binds to the ThZFP1 promoter. Additionally, ThDof1.4 and ThZFP1 share similar expression patterns in response to salt or drought stress. Furthermore, like ThZFP1, ThDof1.4 could increase the proline level and enhance ROS scavenging capability to improve salt and osmotic stress tolerance. Together, these results suggested that ThDof1.4 and ThZFP1 form a transcriptional regulatory cascade involved in abiotic stress resistance in T. hispida.

Function of MYB8 in larch under PEG simulated drought stress.

Larch, a prominent afforestation, and timber species in northeastern China, faces growth limitations due to drought. To further investigate the mechanism of larch's drought resistance, we conducted full-length sequencing on embryonic callus subjected to PEG-simulated drought stress. The sequencing results revealed that the differentially expressed genes (DEGs) primarily played roles in cellular activities and cell components, with molecular functions such as binding, catalytic activity, and transport activity. Furthermore, the DEGs showed significant enrichment in pathways related to protein processing, starch and sucrose metabolism, benzose-glucuronic acid interconversion, phenylpropyl biology, flavonoid biosynthesis, as well as nitrogen metabolism and alanine, aspartic acid, and glutamic acid metabolism. Consequently, the transcription factor T_transcript_77027, which is involved in multiple pathways, was selected as a candidate gene for subsequent drought stress resistance tests. Under PEG-simulated drought stress, the LoMYB8 gene was induced and showed significantly upregulated expression compared to the control. Physiological indices demonstrated an improved drought resistance in the transgenic plants. After 48 h of PEG stress, the transcriptome sequencing results of the transiently transformed LoMYB8 plants and control plants exhibited that genes were significantly enriched in biological process, cellular component and molecular function. Function analyses indicated for the enrichment of multiple KEGG pathways, including energy synthesis, metabolic pathways, antioxidant pathways, and other relevant processes. The pathways annotated by the differential metabolites mainly encompassed signal transduction, carbohydrate metabolism, amino acid metabolism, and flavonoid metabolism.

Transcriptional regulatory cascade of LcMYB71 and LcNAC73 affects low-temperature and drought stress response in Lonicera caerulea.

The development of stress tolerance is regulated via the transcriptional regulatory networks involving regulatory homeostasis mediated by protein-DNA interactions. LcNAC73 from Lonicera caerulea was characterized to understand the underlying mechanism of low-temperature and drought stress response in L. caerulea. To better understand the transcription pathway of LcNAC73, we cloned the promoter and screened proteins that could interact with the promoter. Using Yeast one-hybrid, electrophoretic mobility shift, and chromatin immunoprecipitation assays, we found that the LcMYB71 protein specifically bound to the promoter of LcNAC73. The transient transformation and stable transgenic system were used to produce transgenic L. caerulea plants with overexpressed and silenced LcNAC73, elucidating the effect of LcNAC73 on low-temperature and drought stress tolerance. LcNAC73 positively regulated the proline content and enhanced the scavenging of reactive oxygen species, thus improving tolerance to low-temperature and drought stress. Further studies revealed that LcMYB71 and LcNAC73 had similar functions and could improve plant low-temperature and drought tolerance. It is necessary to identify the upstream regulators of a specific gene to characterize gene functions and the associated transcriptional pathways.

An ERF transcription factor from Tamarix hispida, ThCRF1, can adjust osmotic potential and reactive oxygen species scavenging capability to improve salt tolerance.

Ethylene-Responsive Factors (ERFs) are plant-specific transcription factors (TFs) involved in multiple biological processes, especially in abiotic stress tolerance. However, the ERFs from woody halophytes that are involved in salt stress have been little studied. In the present investigation, we characterized a subfamily member of ERF TFs from Tamarix hispida, ThCRF1, which responds to salt stress. ThCRF1 is a nuclear protein that binds to the motifs including TTG, DRE and GCC-box. Transient transformation was performed to generate T. hispida overexpressing ThCRF1 and RNA interference (RNAi)-silenced ThCRF1 to analyze its function using gain- and loss-of-function methods. Overexpression of ThCRF1 in T. hispida significantly improved tolerance to salt-shock-induced stress; by contrast, RNAi-silence of ThCRF1 significantly decreased tolerance to salt-shock-induced stress. Further experiments showed that ThCRF1 induces the expression of genes including those encoding pyrroline-5-carboxylate synthetase (P5CS), trehalose-6-phosphate synthase (TPS), trehalose-6-phosphate phosphatase (TPP), superoxide dismutase (SOD) and peroxidase (POD), which lead to enhanced proline and trehalose levels and increased SOD and POD activities. These results were further confirmed by studying transgenic Arabidopsis plants overexpressing ThCRF1. Therefore, the results suggested that ThCRF1 improves tolerance to salt-shock-induced stress by enhancing trehalose and proline biosynthesis to adjust the osmotic potential, and by improving SOD and POD activities to increase reactive oxygen species scavenging capability.