Overexpression of Pennisetum purpureum CCoAOMT Contributes to Lignin Deposition and Drought Tolerance by Promoting the Accumulation of Flavonoids in Transgenic Tobacco.

Elephant grass (Pennisetum purpureum) is a fast-growing and low-nutrient demand plant that is widely used as a forage grass and potential energy crop in tropical and subtropical regions of Asia, Africa, and the United States. Transgenic tobacco with the PpCCoAOMT gene from Pennisetum purpureum produces high lignin content that is associated with drought tolerance in relation to lower accumulation of reactive oxygen species (ROS), along with higher antioxidant enzyme activities and osmotic adjustment. In this study, transgenic tobacco plants revealed no obvious cost to plant growth when expressing the PpCCoAOMT gene. Metabolomic studies demonstrated that tobacco plants tolerant to drought stress accumulated flavonoids under normal and drought conditions, which likely explains the observed tolerance phenotype in wild-type tobacco. Our results suggest that plants overexpressing PpCCoAOMT were better able to cope with water deficit than were wild-type controls; metabolic flux was redirected within primary and specialized metabolism to induce metabolites related to defense to drought stress. These results could help to develop drought-resistant plants for agriculture in the future.

Overexpression of SgGH3.1 from Fine-Stem Stylo (Stylosanthes guianensis var. intermedia) Enhances Chilling and Cold Tolerance in Arabidopsis thaliana.

Stylosanthes (stylo) species are commercially significant tropical and subtropical forage and pasture legumes that are vulnerable to chilling and frost. However, little is known about the molecular mechanisms behind stylos' responses to low temperature stress. Gretchen-Hagen 3 (GH3) proteins have been extensively investigated in many plant species for their roles in auxin homeostasis and abiotic stress responses, but none have been reported in stylos. SgGH3.1, a cold-responsive gene identified in a whole transcriptome profiling study of fine-stem stylo (S. guianensis var. intermedia) was further investigated for its involvement in cold stress tolerance. SgGH3.1 shared a high percentage of identity with 14 leguminous GH3 proteins, ranging from 79% to 93%. Phylogenetic analysis classified SgGH3.1 into Group Ⅱ of GH3 family, which have been proven to involve with auxins conjugation. Expression profiling revealed that SgGH3.1 responded rapidly to cold stress in stylo leaves. Overexpression of SgGH3.1 in Arabidopsis thaliana altered sensitivity to exogenous IAA, up-regulated transcription of AtCBF1-3 genes, activated physiological responses against cold stress, and enhanced chilling and cold tolerances. This is the first report of a GH3 gene in stylos, which not only validated its function in IAA homeostasis and cold responses, but also gave insight into breeding of cold-tolerant stylos.

SgRVE6, a LHY-CCA1-Like Transcription Factor From Fine-Stem Stylo, Upregulates NB-LRR Gene Expression and Enhances Cold Tolerance in Tobacco.

Stylosanthes species are economically important tropical and subtropical forage legumes which are generally vulnerable to chilling and frost. Fine-stem stylo (S. guianensis var. intermedia) has the most superior cold tolerance among all stylo species. A REVEILLE (RVE) gene, SgRVE6, was cloned from fine-stem stylo. Bioinformatic analysis suggests that SgRVE6 encodes a transcription factor of 292 amino acid residues, which belongs to the LATE ELONGATED HYPOCOTYL/CIRCADIAN CLOCK ASSOCIATED 1-LIKE (LCL) subgroup of RVE family and contains a SHAQKYF-class MYB domain and a LCL domain. SgRVE6 is universally expressed in root, stem and leaf tissues of fine-stem stylo and is rapidly up-regulated in all tested tissues under cold stress. Over-expressing SgRVE6 affects expression of 21 circadian clock genes, up-regulates expression of 6 nucleotide binding domain leucine-rich repeats (NB-LRR) encoding genes associated with tobacco cold tolerance, improves physiological responses to low temperature, and endows the transgenic tobaccos with higher tolerance to cold stress. This is the first time a study investigates the biological function of RVE6 in cold responses of plant species.

Vascular preferential activity of the Pennisetum purpureum cinnamyl alcohol dehydrogenase promoter in transgenic tobacco plants.

Little is known about the cross talk between the lignin biosynthesis gene promoters and the regulatory proteins that modulate molecular signaling and respond to various stresses. In this study, we characterized the promoter region of the lignin biosynthesis pathway cinnamyl alcohol dehydrogenase (CAD) gene in elephant grass, Pennisetum purpureum. Quantification of the transcript levels of the PpCAD promoter revealed it is preferentially expressed in vascular tissue, especially xylem. Histochemical and fluorometric assays confirmed the vascular-preferential expression of the PpCAD promoter, as the highest ß-glucuronidase (GUS) activity was found in the basal stem in transgenic tobacco plants expressing a 1154-bp PpCAD promoter-GUS fusion construct. Moreover, 5'-deleted PpCAD promoter analyses showed that the 1154-bp PpCAD promoter fragment had the highest transcriptional activity, whereas the 2054-bp fragment had multifarious inducible activity responding to gibberellin (GA), methyl jasmonate (MeJA), abscisic acid (ABA), and wounding. The regions from -248 to -243 bp and -1416 to -1411 bp contained W-box cis-elements, which were detected by electrophoretic mobility shift assay (EMSA). The binding effects of the GA-responsive elements (from -561 to -555 bp and -1077 to -1071 bp), MeJA-responsive element (from -1146 to -1142 bp), and the ABA-responsive cis-element (from -1879 to -1874 bp) were also validated by EMSA. Based on our results, we suggest that lignin deposition associated with PpCAD promoter activity adapts to the environment through molecular signaling involving GA, MeJA, and ABA.