Perennial ryegrass (Lolium perenne) is one of the most widely used forage and turf grasses in the world due to its desirable agronomic qualities. However, as a cool-season perennial grass species, high temperature is a major factor limiting its performance in warmer and transition regions. In this study, a de novo transcriptome was generated using a cDNA library constructed from perennial ryegrass leaves subjected to short-term heat stress treatment. Then the expression profiling and identification of perennial ryegrass heat response genes by digital gene expression analyses was performed. The goal of this work was to produce expression profiles of high temperature stress responsive genes in perennial ryegrass leaves and further identify the potentially important candidate genes with altered levels of transcript, such as those genes involved in transcriptional regulation, antioxidant responses, plant hormones and signal transduction, and cellular metabolism. The de novo assembly of perennial ryegrass transcriptome in this study obtained more total and annotated unigenes compared to previously published ones. Many DEGs identified were genes that are known to respond to heat stress in plants, including HSFs, HSPs, and antioxidant related genes. In the meanwhile, we also identified four gene candidates mainly involved in C4 carbon fixation, and one TOR gene. Their exact roles in plant heat stress response need to dissect further. This study would be important by providing the gene resources for improving heat stress tolerance in both perennial ryegrass and other cool-season perennial grass plants.
As a conserved microRNA (miRNA) family in plants, miR408 is known to be involved in different abiotic stress responses, including drought. Interestingly, some studies indicated a species-and/or cultivar-specific drought-responsive characteristic of miR408 in plant drought stress. Moreover, the functions of miR408 in perennial grass species are unknown. In this study, we investigated the role of miR408 in perennial ryegrass (Lolium perenne L.) by withholding water for 10 days for both wild type and transgenic plants with heterologous expression of rice (Oryza sativa L.) miR408 gene, Os-miR408. The results showed that transgenic perennial ryegrass plants displayed morphological changes under normal growth conditions, such as curl leaves and sunken stomata, which could be related to decreased leaf water loss. Moreover, transgenic perennial ryegrass exhibited improved drought tolerance, as demonstrated by maintaining higher leaf relative water content (RWC), lower electrolyte leakage (EL), and less lipid peroxidation compared to WT plants under drought stress. Furthermore, the transgenic plants showed higher antioxidative capacity under drought.These results showed that the improved drought tolerance in Os-miR408 transgenic plants could be due to leaf morphological changes favoring the maintenance of water status and to increased antioxidative capacity protecting against the reactive oxygen species damages under stress. These findings implied that miR408 could serve as a potential target for genetic manipulations to engineer perennial grass plants for improved water stress tolerance. | INTRODUCTIONAs sessile organisms living in constantly changing environment, plants are often exposed to abiotic stress, such as drought, salt, and temperature stresses (Zhu, 2016), which leads to more than 50% yield loss in major crops worldwide each year (Boyer, 1982;Lobell et al., 2011).Meanwhile, due to global climate change and world population growth, water scarcity is already a critical concern in many parts of the world, and drought has become one of the most adverse factors of plant growth and productivity (Shao et al., 2009). Therefore, it is critical to breed drought-tolerant crops and study plants' mechanism in response to drought stress.Plants can perceive abiotic stress signals and elicit appropriate responses by altering metabolism, growth, and development (Bartels & Sunkar, 2005). The primary signal caused by drought is hyperosmotic stress, which causes the accumulation of reactive oxygen species (ROS), damage to cellular components such as membrane lipids, proteins, nucleic acids, and metabolic dysfunction (Zhu, 2016).
With global warming, high temperature stress has become a main threat to the growth of cool-season turfgrasses, including perennial ryegrass. As one of the conserved plant microRNA families, miR408s are known to play roles in various abiotic stresses, including cold, drought, salinity, and oxidative stress, but no report, thus far, was found for heat. Here, perennial ryegrass plants overexpressing rice Os-miR408 were used to investigate the role of miR408 in plant heat tolerance. Both wild type (WT) and miR408 transgenic perennial ryegrass plants (TG) were subjected to short-term heat stress at 38 °C for 72 h (experiment 1) or at 42 °C for 48 h (experiment 2), and then let recover for 7 days at optimum temperature. Morphological changes and physiological parameters, including antioxidative responses of TG and WT plants, were compared. The results showed that miR408 downregulated the expression of two putative target genes, PLASTOCYANIN and LAC3. Additionally, overexpression of Os-miR408 improved thermo-tolerance of perennial ryegrass, demonstrated by lower leaf lipid peroxidation and electrolyte leakage, and higher relative water content after both 38 and 42 °C heat stresses. In addition, the enhanced thermotolerance of TG plants could be associated with its morphological changes (e.g., narrower leaves, smaller tiller angles) and elevated antioxidative capacity. This study is the first that experimentally reported a positive role of miR408 in plant tolerance to heat stress, which provided useful information for further understanding the mechanism by which miR408 improved plant high-temperature tolerance, and offered a potential genetic resource for breeding heat-resistant cool-season turfgrass in the future.
Experiments were conducted to study the relationship between autumn (fall) dormancy rating (FDR) and salt tolerance of lucerne (alfalfa, Medicago sativa L.). Seeds of eight cultivars with different FDRs (range 2.0–10.2) were germinated in a growth chamber under five concentrations of NaCl (range 0–1.2%). A subordinate function value method was adopted for comprehensive evaluation of salt tolerance of the lucerne cultivars. Seed germination rate and germination potential, and seedling electrolyte leakage, shoot length, root length, fresh weight, root : shoot length ratio, Na+ and K+ concentrations and K+ : Na+ ratio were used as the indices of evaluation. Under 1.2% salt treatment, salt-tolerance ranking of the cultivars was WL656HQ > WL440HQ > WL712HQ > WL525HQ > WL343HQ > WL319HQ > WL363HQ > WL168HQ, with respective FDRs of 9.3, 6.0, 10.2, 8.2, 3.9, 2.8, 4.9 and 2.0. The results of this study show that non-dormant cultivars were generally more salt tolerant than dormant cultivars.
Watermelon fruit contains a high percentage of amino acid citrulline (Cit) and arginine (Arg). Cit and Arg accumulation in watermelon fruit are most likely mediated by both de novo synthesis from other amino acids within fruits and direct import from source tissues (leaves) through the phloem. The amino acid transporters involved in the import of Cit, Arg, and their precursors into developing fruits of watermelon have not been reported. In this study, we have compiled the list of putative amino acid transporters in watermelon and characterized transporters that are expressed in the early stage of fruit development. Using the yeast complementation study, we characterized ClAAP3 (Cla023187) and ClAAP6 (Cla023090) as functional amino acid transporters belonging to the family of amino acid permease (AAP) genes. The yeast growth and uptake assays of radiolabeled amino acid suggested that ClAAP3 and ClAAP6 can transport a broad spectrum of amino acids. Expression of translational fusion proteins with a GFP reporter in Nicotiana benthamiana leaves confirmed the ER- and plasma membrane-specific localization, suggesting the role of ClAAP proteins in the cellular import of amino acids. Based on the gene expression profiles and functional characterization, ClAAP3 and ClAAP6 are expected to play a major role in regulation of amino acid import into developing watermelon fruits.
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