Genome-Wide Characterization of DNase I-Hypersensitive Sites and Cold Response Regulatory Landscapes in Grasses.

Deep sequencing of DNase-I treated chromatin (DNase-seq) can be used to identify DNase I-hypersensitive sites (DHSs) and facilitates genome-scale mining of de novo cis-regulatory DNA elements. Here, we adapted DNase-seq to generate genome-wide maps of DHSs using control and cold-treated leaf, stem, and root tissues of three widely studied grass species: Brachypodium distachyon, foxtail millet (Setaria italica), and sorghum (Sorghum bicolor). Functional validation demonstrated that 12 of 15 DHSs drove reporter gene expression in transiently transgenic B. distachyon protoplasts. DHSs under both normal and cold treatment substantially differed among tissues and species. Intriguingly, the putative DHS-derived transcription factors (TFs) are largely colocated among tissues and species and include 17 ubiquitous motifs covering all grass taxa and all tissues examined in this study. This feature allowed us to reconstruct a regulatory network that responds to cold stress. Ethylene-responsive TFs SHINE3, ERF2, and ERF9 occurred frequently in cold feedback loops in the tissues examined, pointing to their possible roles in the regulatory network. Overall, we provide experimental annotation of 322,713 DHSs and 93 derived cold-response TF binding motifs in multiple grasses, which could serve as a valuable resource for elucidating the transcriptional networks that function in the cold-stress response and other physiological processes.

Arabidopsis thaliana trehalose-6-phosphate phosphatase gene TPPI enhances drought tolerance by regulating stomatal apertures.

Transpiration occurs through stomata. The alteration of stomatal apertures in response to drought stress is an important process associated with water use efficiency (WUE). Trehalose-6-phosphate phosphatase (TPP) family genes have been reported to participate in adjustment of stomatal aperture. However, there have been no reports of the trehalose metabolism pathway genes improving WUE, and the upstream signalling pathway modulating these genes is not clear. Here, we demonstrate that a member of the TPP gene family, AtTPPI, confers drought resistance and improves WUE by decreasing stomatal apertures and improving root architecture. The reduced expression of AtTPPI caused a drought-sensitive phenotype, while its overexpression significantly increased drought tolerance. Abscisic acid (ABA)-induced stomatal closure experiments confirmed that AtTPPI mutation increased the stomatal aperture compared with that of wild-type plants; in contrast, overexpression plants had smaller stomatal apertures than those of wild-type plants. Moreover, AtTPPI mutation also caused stunted primary root length and compromised auxin transport, while overexpression plants had longer primary root lengths. Yeast one-hybrid assays showed that ABA-responsive element-binding factor1 (ABF1), ABF2, and ABF4 directly regulated AtTPPI expression. In summary, the way in which AtTPPI responds to drought stress suggests that AtTPPI-mediated stomatal regulation is an important mechanism to cope with drought stress and improve WUE.

Overexpression of the trehalose-6-phosphate phosphatase family gene AtTPPF improves the drought tolerance of Arabidopsis thaliana.

BACKGROUND: Trehalose-6-phosphate phosphatases (TPPs), which are encoded by members of the TPP gene family, can improve the drought tolerance of plants. However, the molecular mechanisms underlying the dynamic regulation of TPP genes during drought stress remain unclear. In this study, we explored the function of an Arabidopsis TPP gene by conducting comparative analyses of a loss-of-function mutant and overexpression lines. RESULTS: The loss-of-function mutation of Arabidopsis thaliana TPPF, a member of the TPP gene family, resulted in a drought-sensitive phenotype, while a line overexpressing TPPF showed significantly increased drought tolerance and trehalose accumulation. Compared with wild-type plants, tppf1 mutants accumulated more H2O2 under drought, while AtTPPF-overexpressing plants accumulated less H2O2 under drought. Overexpression of AtTPPF led to increased contents of trehalose, sucrose, and total soluble sugars under drought conditions; these compounds may play a role in scavenging reactive oxygen species. Yeast one-hybrid and luciferase activity assays revealed that DREB1A could bind to the DRE/CRT element within the AtTPPF promoter and activate the expression of AtTPPF. A transcriptome analysis of the TPPF-overexpressing plants revealed that the expression levels of drought-repressed genes involved in electron transport activity and cell wall modification were upregulated, while those of stress-related transcription factors related to water deprivation were downregulated. These results indicate that, as well as its involvement in regulating trehalose and soluble sugars, AtTPPF is involved in regulating the transcription of stress-responsive genes. CONCLUSION: AtTPPF functions in regulating levels of trehalose, reactive oxygen species, and sucrose levels during drought stress, and the expression of AtTPPF is activated by DREB1A in Arabidopsis. These findings shed light on the molecular mechanism by which AtTPPF regulates the response to drought stress.