Xinli Xia

The salt- and drought-inducible poplar GRAS protein SCL7 confers salt and drought tolerance in Arabidopsis thaliana

The plant-specific GRAS/SCL transcription factors play diverse roles in plant development and stress responses. In this study, a poplar SCL gene, PeSCL7, was functionally characterized in Arabidopsis thaliana, especially with regard to its role in abiotic stress resistance. Expression analysis in poplar revealed that PeSCL7 was induced by drought and high salt stresses, but was repressed by gibberellic acid (GA) treatment in leaves. Transient expression of GFP-PeSCL7 in onion epidermal cells revealed that the PeSCL7 protein was localized in the nucleus. Transgenic Arabidopsis plants overexpressing PeSCL7 showed enhanced tolerance to drought and salt treatments. The activity of two stress-responsive enzymes was increased in transgenic seedlings. Taken together, these results suggest that PeSCL7 encodes a member of the stress-responsive GRAS/SCL transcription factors that is potentially useful for engineering drought- and salt-tolerant trees.

Expression profiling and functional characterization of a DREB2-type gene from Populus euphratica.

A novel DREB (dehydration responsive element binding) gene, designated PeDREB2, was isolated from the desert-grown tree, Populus euphratica. Based on multiple sequence alignment and phylogenetic characterization, PeDREB2 was classified as an A-2 group member of the DREB family. Expression of PeDREB2 was induced by cold, drought, and high salinity, but not by abscisic acid (ABA) treatment. PeDREB2 could bind specifically to DRE elements and was targeted to the nucleus when transiently expressed in onion epidermis cells. 35S promoter-driven expression of PeDREB2 improved salt tolerance in transgenic tobacco and did not cause growth retardation. The results indicate that PeDREB2 functions as a novel transcription factor involved in the response of salt stress and might be useful in improving salt tolerance in transgenic plants.

Genome-wide identification and functional prediction of novel and drought-responsive lincRNAs in Populus trichocarpa

Summary A total of 2 542 lincRNAs were identified from Populus trichocarpa and some of them play key roles in drought stress tolerance or regulate microRNA through target mimicry patterns.

Identification of drought-responsive and novel Populus trichocarpa microRNAs by high-throughput sequencing and their targets using degradome analysis

BackgroundMicroRNAs (miRNAs) are endogenous small RNAs (sRNAs) with a wide range of regulatory functions in plant development and stress responses. Although miRNAs associated with plant drought stress tolerance have been studied, the use of high-throughput sequencing can provide a much deeper understanding of miRNAs. Drought is a common stress that limits the growth of plants. To obtain more insight into the role of miRNAs in drought stress, Illumina sequencing of Populus trichocarpa sRNAs was implemented.ResultsTwo sRNA libraries were constructed by sequencing data of control and drought stress treatments of poplar leaves. In total, 207 P. trichocarpa conserved miRNAs were detected from the two sRNA libraries. In addition, 274 potential candidate miRNAs were found; among them, 65 candidates with star sequences were chosen as novel miRNAs. The expression of nine conserved miRNA and three novel miRNAs showed notable changes in response to drought stress. This was also confirmed by quantitative real time polymerase chain reaction experiments. To confirm the targets of miRNAs experimentally, two degradome libraries from the two treatments were constructed. According to degradome sequencing results, 53 and 19 genes were identified as targets of conserved and new miRNAs, respectively. Functional analysis of these miRNA targets indicated that they are involved in important activities such as the regulation of transcription factors, the stress response, and lipid metabolism.ConclusionsWe discovered five upregulated miRNAs and seven downregulated miRNAs in response to drought stress. A total of 72 related target genes were detected by degradome sequencing. These findings reveal important information about the regulation mechanism of miRNAs in P. trichocarpa and promote the understanding of miRNA functions during the drought response.

Global identification of miRNAs and targets in Populus euphratica under salt stress

Populus euphratica, a typical hydro-halophyte, is ideal for studying salt stress responses in woody plants. MicroRNAs (miRNAs) are endogenous non-coding small RNAs that fulfilled an important post-transcriptional regulatory function. MiRNA may regulate tolerance to salt stress but this has not been widely studied in P. euphratica. In this investigation, the small RNAome, degradome and transcriptome were studied in salt stress treated P. euphratica by deep sequencing. Two hundred and eleven conserved miRNAs between Populus trichocarpa and P. euphratica have been found. In addition, 162 new miRNAs, belonging to 93 families, were identified in P. euphratica. Degradome sequencing experimentally verified 112 targets that belonged to 51 identified miRNAs, few of which were known previously in P. euphratica. Transcriptome profiling showed that expression of 15 miRNA-target pairs displayed reverse changing pattern under salt stress. Together, these results indicate that, in P. euphratica under salt stress, a large number of new miRNAs could be discovered, and both known and new miRNA were functionally cleaving to their target mRNA. Expression of miRNA and target were correspondingly induced by salt stress but that it was a complex process in P. euphratica.

De novo sequencing and transcriptome analysis of the desert shrub, Ammopiptanthus mongolicus, during cold acclimation using Illumina/Solexa

BackgroundAmmopiptanthus mongolicus (Maxim. ex Kom.) Cheng f., an evergreen broadleaf legume shrub, is distributed in Mid-Asia where the temperature can be as low as −30°C during the winter. Although A. mongolicus is an ideal model to study the plant response to cold stress, insufficient genomic resources for this species are available in public databases. To identify genes involved in cold acclimation (a phenomenon experienced by plants after low temperature stress), a high-throughput sequencing technology was applied.ResultsWe sequenced cold-treated and control (untreated) samples of A. mongolicus, and obtained 65,075,656 and 67,287,120 high quality reads, respectively. After de novo assembly and quantitative assessment, 82795 all-unigenes were finally generated with an average length of 816xa0bp. We then obtained functional annotations by aligning all-unigenes with public protein databases including NR, SwissProt, KEGG and COG. Differentially expressed genes (DEGs) were investigated using the RPKM method. Overall, 9309 up-regulated genes and 23419 down-regulated genes were identified. To increase our understanding of these DEGs, we performed GO enrichment and metabolic pathway enrichment analyses. Based on these results, a series of candidate genes involved in cold responsive pathways were selected and discussed. Moreover, we analyzed transcription factors, and found 720 of them are differentially expressed. Finally, 20 of the candidate genes that were up-regulated and known to be associated with cold stress were examined using qRT-PCR.ConclusionsIn this study, we identified a large set of cDNA unigenes from A. mongolicus. This is the first transcriptome sequencing of this non-model species under cold-acclimation using Illumina/Solexa, a next-generation sequencing technology. We sequenced cold-treated and control (untreated) samples of A. mongolicus and obtained large numbers of unigenes annotated to public databases. Studies of differentially expressed genes involved in cold-related metabolic pathways and transcription factors facilitate the discovery of cold-resistance genes.

Molecular characterization of putative vacuolar NHX-type Na+/H+ exchanger genes from the salt-resistant tree Populus euphratica

The vacuolar NHX-type Na(+)/H(+) exchangers play a key role in salt tolerance in plants. However, little is known about the Na(+)/H(+) exchangers in the salt-resistant tree, Populus euphratica. In this study, we identified six putative vacuolar Na(+)/H(+) exchanger genes from P. euphratica, designated as PeNHX1-6. Real-time polymerase chain reaction indicated that the PeNHX1/3/6 transcripts were abundant compared with the other three PeNHX genes in the three tissues (roots, stems and leaves) examined. After NaCl treatment for 6 h, the transcript levels of PeNHX1-6 were upregulated in the roots. To address the function of PeNHX1-6, complementation studies were performed with the salt-sensitive yeast mutant strain R100, which lacks activity of the endosomal Na(+)/H(+) antiporter NHX1. The results showed that PeNHX1-6 compensates, at least in part, for the function of yeast NHX1. Moreover, PeNHX3 was targeted to the tonoplast when transiently expressed in onion. Together, these results suggest that PeNHX1-6 function as vacuolar Na(+)/H(+) exchangers and that PeNHX products play an important role in the salt resistance of P. euphratica.

Populus euphratica: the transcriptomic response to drought stress

Populus euphratica Olivier is widely established in arid and semiarid regions but lags in the availability of transcriptomic resources in response to water deficiency. To investigate the mechanisms that allow P. euphratica to maintain growth in arid regions, the responses of the plant to soil water deficit were analyzed at a systems level using physiological and pyrosequencing approaches. We generated 218,601 and 287,120 reads from non-stressed control and drought-stressed P. euphratica leaves respectively, totaling over 200xa0million base pairs. After assembly, 24,013 transcripts were yielded with an average length of 1,128xa0bp. We determined 2,279 simple sequence repeats, which may have possible information for understanding drought adaption of woody plants. Stomatal closure was inhibited under moderate drought to maintain a relatively high rate of CO2 assimilation and water transportation, which was supposed to be important for P. euphratica to maintain normal growth and develop vigorous root systems in an adverse environment. This was accompanied by strong transcriptional remodeling of stress-perception, signaling and transcription regulation, photoprotective system, oxidative stress detoxification, and other stress responsive genes. In addition, genes involved in stomatal closure inhibition, ascorbate–glutathione pathway and ubiquitin–proteasome system that may specially modulate the drought stress responses of P. euphratica are highlighted. Our analysis provides a comprehensive picture of how P. euphratica responds to drought stress at physiological and transcriptome levels which may help to understand molecular mechanisms associated with drought response and could be useful for genetic engineering of woody plants.

Overexpression of the poplar NF-YB7 transcription factor confers drought tolerance and improves water-use efficiency in Arabidopsis

Water deficit is a serious environmental factor limiting the growth and productivity of plants worldwide. Improvement of drought tolerance and efficient water use are significant strategies to overcome this dilemma. In this study, a drought-responsive transcription factor, NUCLEAR FACTOR Y subunit B 7 (PdNF-YB7), induced by osmotic stress (PEG6000) and abscisic acid, was isolated from fast-growing poplar clone NE-19 [Populus nigra × (Populus deltoides × Populus nigra)]. Ectopic overexpression of PdNF-YB7 (oxPdB7) in Arabidopsis enhanced drought tolerance and whole-plant and instantaneous leaf water-use efficiency (WUE, the ratio of biomass produced to water consumed). Overexpressing lines had an increase in germination rate and root length and decrease in water loss and displayed higher photosynthetic rate, instantaneous leaf WUE, and leaf water potential to exhibit enhanced drought tolerance under water scarcity. Additionally, overexpression of PdNF-YB7 in Arabidopsis improved whole-plant WUE by increasing carbon assimilation and reducing transpiration with water abundance. These drought-tolerant, higher WUE transgenic Arabidopsis had earlier seedling establishment and higher biomass than controls under normal and drought conditions. In contrast, Arabidopsis mutant nf-yb3 was more sensitive to drought stress with lower WUE. However, complementation analysis indicated that complementary lines (nf-yb3/PdB7) had almost the same drought response and WUE as wild-type Col-0. Taken together, these results suggest that PdNF-YB7 positively confers drought tolerance and improves WUE in Arabidopsis; thus it could potentially be used in breeding drought-tolerant plants with increased production even under water deficiency.

Expression profiles of precursor and mature microRNAs under dehydration and high salinity shock in Populus euphratica.

MicroRNAs (miRNAs) are small non-coding RNAs that play vital roles in plant abiotic stress responses via cleavage or translational inhibition of their target mRNAs. Populus euphratica is a typical stress-resistant sessile organism that grows in desert areas. Here, we identified sequences of 12 miRNA precursors from 11 families and 13 mature miRNAs from 12 families by PCR amplification in P. euphratica. To detect expression differences in mature miRNAs and their precursors under dehydration and high salinity shock in P. euphratica, we examined 14 miRNA precursors from 13 miRNA families and 17 mature miRNAs from 17 miRNA families using the SYBR Green RT–PCR assay. This is the first report of expression profiles for both precursor and mature miRNAs in P. euphratica. By profiling both the mature miRNAs and the precursors under abiotic stress shock, it was possible to identify miRNA whose processing is regulated during stress shock environments. A majority of the genes predicted to be targets for plant miRNAs are involved in development, stress resistance and metabolic processes. We have cloned and experimentally identified in vivo five of the predicted target genes and quantified the five target mRNAs from the same RNA sample simultaneously. Based on this study, we propose some regulatory pathways that illustrate the important role that miRNAs play in response to abiotic stress shock in P. euphratica.