Project description:Alfalfa is the most produced perennial forage crop in Canada. Drought stress is a major form of abiotic stress, affecting its productivity and annual yield. A small RNA, miR156, plays a major role in drought tolerance by downregulating downstream SPL genes, but its effects at the proteome level are unknown. In this study, the protein level perturbations of miR156 overexpression (A8) and empty vector (EV) control genotypes were compared under drought stress. Using label-free quantification, 3,000 protein groups were identified, of which 68 were upregulated in A8 and 84 were downregulated relative to EV under control conditions. Conversely, under drought stress, 610 proteins were upregulated and only 52 proteins were downregulated in A8 relative to EV. Functional analysis using PlantRegMap showed that the enriched proteins are likely involved in biological and molecular processes including antioxidant response, response to stress, signal transduction and biosynthesis of secondary metabolites. These proteins/pathways might be involved in the enhancement of drought stress tolerance mediated by miR156. Protein groups related to signaling, such as MAP kinase, calcium-dependent protein kinase, protein phosphatase 2C, and transcriptional regulators including bZIP and zinc finger proteins were found to be differentially expressed when a search was conducted against a drought stress gene database. The proteomic dataset was validated by immunoblotting of selected proteins. The results of this study provide a better understanding and insight into the role of miR156 in drought stress tolerance in alfalfa at the proteomic level.
Project description:Two alfalfa varieties, 'Chilean' (M. sativa ssp. sativa var. Chilean, drought sensitive) and 'Wisfal' (M. sativa ssp. falcata var. Wisfal, drought tolerant), with contrasting water use efficiency were subjected to water withholding for 11 days followed by re-watering. Samples were taken for well-watered plants and plants after five, eight, eleven days of drought stress as well as plants after recovery for one day following drought stress. Roots and shoots were sampled and analyzed separately by expression profiling using Affymetrix Medicago GeneChip.
Project description:OsNAC6 is a stress responsive NAC transcription factor in rice known as a regulator for the transcriptional networks of the drought tolerance mechanisms. However, little is known about the associated molecular mechanisms for drought tolerance. Here, we identified OsNAC6-mediated root structural adaptation such as increased root number and root diameter that was sufficient to confer drought tolerance. Multiyear (5 years) drought field tests clearly demonstrated that OsNAC6 overexpression in roots produced higher grain yield under drought conditions. Genome-wide analyses revealed that OsNAC6 directly up-regulated 13 genes. Taken together, OsNAC6 is a valuable candidate for genetic engineering of drought-tolerant high-yielding crops.
Project description:Brachypodium distachyon as an annual species that colonises broken ground is a highly appropriate model to define drought tolerance mechanisms. We have previously identified accessions exhibiting drought tolerance, high susceptibility and intermediate tolerance to drought; respectively, ABR8, KOZ1 and ABR4, from a screen of 138 genotypes. To define the mechanisms of tolerance, the responses to drought were assessed using transcriptomic and metabolomic approaches. Analysis of RNA-seq before and following drought suggested relatively few differentially expressed genes in ABR8. Linking these to gene ontology (GO) terms revealed an enrichment for biological processes related to “regulated stress response”, “plant cell wall” and “oxidative stress”. Interestingly, drought tolerance also appeared to correlate with pre-existing differences in gene expression linked to GO terms associated with the cell wall. These included glycoside hydrolases involved in cell wall remodelling, pectin methylesterases, expansins and a pectin acetylesterase. Metabolomic assessments of the same samples also indicated few statistically significant changes in ABR8 with drought. Instead, pre-existing differences in the cell-wall associated metabolites appeared to correlate with drought tolerance. Our data suggests two different modes/levels at which cell wall characteristics can play a role in conferring drought tolerance: (i) an active response mode/level which involves stress induced changes in cell wall features to help the plant cope with drought and (ii) an intrinsic mode in which innate differences in cell wall composition and architecture between genotypes are important in determining tolerance to drought stress. Both modes seem to contribute to the drought tolerance of ABR8. Identification of the exact mechanisms through which the cell wall confers drought tolerance will be important to fully exploit the contribution of the cell wall in the development of drought tolerant cereals and other grasses.
Project description:Plant stress response and tolerance mechanisms are controlled by diverse genes. Transcription factors have been implicated in drought tolerance under drought stress conditions. Identification of target genes of such transcription factors could offer molecular regulatory networks by which the tolerance mechanisms orchestrated. Previously, we generated transgenic rice plants with 4 rice transcription factors OsNAC5, 6, 9, and 10 under the root-specific promoter RCc3 that were tolerant to drought stress with less loss of grain yield under drought conditions. To understand the molecular mechanisms of drought tolerance, we performed ChIP-Seq and RNA-Seq analyses to identify direct target genes of the OsNACs using the RCc3:MYC-OsNACs roots. A total of 475 binding loci of 4 OsNACs were identified by cross-referencing the binding occupancy of OsNACs at promoter regions and expression levels of corresponding genes. The binding loci are distributed on promoter regions of 391 target genes that were directly up-regulated by OsNACs in four RCc3:MYC-OsNAC transgenic roots. The direct target genes were related to transmembrane/transporter activity, vesicle, plant hormone, carbohydrate metabolism, and transcription factors. The direct targets of each OsNAC are in a range of 4.0 to 8.7% of the genes up-regulated in RNA-Seq data sets. Thus, each OsNAC up-regulates of corresponding direct target genes that alters root system architectures of RCc3:OsNACs for drought tolerance. Our results provide valuable resources for functional dissection of the molecular mechanisms for plant drought tolerance.
Project description:To identify novel microRNAs that are associated with drought tolerance in two different cowpea genotypes, we generated small RNA sequences from adult cowpea plants under control and dought stress treatments. Over 79 million raw reads were generated and numerous novel microRNAs are identified, including some associated with drought tolerance.
Project description:Plants have evolved to possess adaptation mechanism to cope with drought stress by reprograming transcriptional networks through drought responsive transcription factors, which in turn mediate morphological and physiological changes. NAM, ATAF1-2, and CUC2 (NAC) transcription factors are known to be associated with various developmental processes and stress tolerance. In this study, we functionally characterized the rice drought responsive NAC transcription factor OsNAC14. OsNAC14 was predominantly induced at meiosis stage, and induced by drought, high salinity, ABA and low temperature in leaves than roots. Overexpression of OsNAC14 resulted in drought tolerance at the vegetative growth stage and enhanced filling rate at vegetative growth. OsNAC14 overexpression elevated expression of genes related to DNA damage repair, defense response, strigolactone biosynthesis, which correlated with resistance to drought tolerance. Furthermore, OsNAC14 directly bound to the promoter of drought inducible OsRAD51A1, a key component in homologous recombination in DNA repair system. These results indicate that OsNAC14 mediate drought tolerance by recruiting factors involved in DNA damage repair and defense response to enable plant to protect from cellular damage caused by drought stress, thereby provide mechanism for drought tolerance.
Project description:Drought is a major environmental constraint affecting physiological, biochemical and molecular changes of crops, causing loss in crop productivities. Understanding the molecular mechanisms of drought tolerance is important for crop biotechnology. Here, we report that the rice (Oryza sativa) homeodomain-leucine zipper class IV transcription factor gene, Rice outermost cell-specific gene 10 (Roc10), improves drought tolerance and grain yield by increasing lignin accumulation in ground tissues of rice plants. Overexpression of Roc10 significantly enhanced drought tolerance of transgenic rice plants at the vegetative stages of growth with highly effective photosystem and reduction of water loss rate as compared with non-transgenic control and RNAi plants. More importantly, Roc10 overexpression plants had higher drought tolerance at the reproductive stage of growth with higher grain yield over controls under field-drought conditions. We identified downstream and putative target genes of Roc10 by using RNA-seq and ChIP-seq data of rice shoots. Roc10 overexpression elevated the expression levels of lignin biosynthetic genes in shoots with a concomitant increase in accumulation of lignin. The overexpression and RNAi lines showed opposite patterns of lignin accumulation. The Roc10 is mainly expressed in the outer cell layers including epidermis and vasculature of shoots that coincides with areas of increased lignification. Furthermore, the Roc10 was found to directly bind to the promoter of PEROXIDASEN/PEROXIDASE38, a key gene in lignin biosynthesis. Together, our findings suggested that the Roc10 confers drought stress tolerance by enhancing lignin biosynthesis in ground tissues of rice plants.