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The type-B response regulator (ARR) family members represent the elements of two-component regulatory systems and play crucial roles in the cytokinin signaling, plant growth, and stress responses. The objective of this study was to characterize type-B ARR genes from perennial ryegrass, and to investigate their roles in regulating to salt tolerance. Twelve type-B ARRs , namely LpARR1 - 12 , were identified in perennial ryegrass. They were clustered into three subgroups and were unevenly distributed on chromosomes. The stress responsive elements (SREs) and hormone responsive elements (HREs) were identified in LpARRs promoters and the transcription of LpARRs genes were responsive to various abiotic stress (heat, salt, and osmotic) and hormonal (cytokinin, ABA, and ethylene) treatments. LpARR10 was strong induced by salt stress, therefore it was selected for function analysis. Overexpressing LpARR10 in Arabidopsis led to a significant improvement in salt tolerance. The LpARR10 -overexpressing (LpARR10 -OE) lines exhibited lower levels of Na+ content compared to WT plants, and the expression levels of SOS1 and SOS3 were significantly higher in the LpARR10 -OE lines. Results of yeast auto-transactivation and plant cell transcriptional activity assays shows LpARR10 is a nucleus localized transcriptional activator and directly bind and transactivated the promoter of SOS1 and SOS3. Together, LpARR10 enhances salt tolerance may via trans-activating SOS1 / SOS3 in Arabidopsis. Knowledge gained in this study will facilitate further functional analysis of LpARRs for molecular breeding of ryegrass and other related grass species. • Overexpressing LpARR10 resulted in a reduction in Na+ content compared to WT plants. • The LpARR10 -OE lines exhibited significantly higher expression levels of SOS1 and SOS3. • LpARR10 acts as a transcriptional activator and positively regulates salt tolerance by up-regulating the expressions of SOS1 and SOS3. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]