Chuanping Yang

Overexpression of a GST gene (ThGSTZ1) from Tamarix hispida improves drought and salinity tolerance by enhancing the ability to scavenge reactive oxygen species

Although plant glutathione transferase (GST) genes are reported to be involved in responses to abiotic stress, few GST genes have been functionally characterized in woody halophytes. In the present study, a GST gene from Tamarix hispida, designated ThGSTZ1, was cloned and functionally characterized. Expression of ThGSTZ1 was downregulated by drought and salinity stress, and abscisic acid. Transgenic Arabidopsis thaliana plants with constitutive expression of ThGSTZ1 showed increased survival rates under drought and salinity stress. These transgenic Arabidopsis plants exhibited increased levels of GST, glutathione peroxidase, superoxide dismutase and peroxidase activity, along with decreased malondialdehyde content, electrolyte leakage rates and reactive oxygen species (ROS) levels under salt and drought stress conditions. Transgenic T. hispida that transiently overexpressed ThGSTZ1 showed increased GST and GPX activities under NaCl and mannitol treatments, as well as improved ROS scavenging ability. These results suggest that ThGSTZ1 can improve drought and salinity tolerance in plants by enhancing their ROS scavenging ability. Therefore, ThGSTZ1 represents a candidate gene with potential applications for molecular breeding to increase stress tolerance in plants.

Overexpression of ThVHAc1 and its potential upstream regulator, ThWRKY7, improved plant tolerance of Cadmium stress

As one of the most toxic heavy metals in the environment, cadmium (Cd) poses a severe threat to plant growth. We previously reported that overexpression of the Tamarix hispida V-ATPase c subunit (ThVHAc1) improved the Cd tolerance of Saccharomyces cerevisiae. In the current study, we further explored the Cd tolerance conferred by ThVHAc1 in Arabidopsis and T. hispida. ThVHAc1 transgenic Arabidopsis had higher seed germination, biomass, and chlorophyll content under CdCl2 treatment. In Cd-stressed plants, overexpression of ThVHAc1 significantly improved V-ATPase activity and affected the expression of other V-ATPase subunit-encoding genes. Intriguingly, the lower level of ROS accumulation in ThVHAc1-overexpressing lines under CdCl2 treatment demonstrated that ThVHAc1 may modulate Cd stress tolerance by regulating ROS homeostasis. Transient expression of ThVHAc1 in T. hispida further confirmed these findings. Furthermore, promoter analysis and yeast one-hybrid assay revealed that the transcription factor ThWRKY7 can specifically bind to the WRKY cis-element in the ThVHAc1 promoter. ThWRKY7 exhibited similar expression patterns as ThVHAc1 under CdCl2 treatment and improved Cd tolerance, suggesting that ThWRKY7 may be an upstream regulatory gene of ThVHAc1. Therefore, our results show that the combination of ThVHAc1 and its upstream regulator could be used to improve Cd stress tolerance in woody plants.

Expression of the MYB transcription factor gene BplMYB46 affects abiotic stress tolerance and secondary cell wall deposition in Betula platyphylla.

Summary Plant MYB transcription factors control diverse biological processes, such as differentiation, development and abiotic stress responses. In this study, we characterized BplMYB46, an MYB gene from Betula platyphylla (birch) that is involved in both abiotic stress tolerance and secondary wall biosynthesis. BplMYB46 can act as a transcriptional activator in yeast and tobacco. We generated transgenic birch plants with overexpressing or silencing of BplMYB46 and subjected them to gain‐ or loss‐of‐function analysis. The results suggest that BplMYB46 improves salt and osmotic tolerance by affecting the expression of genes including SOD,POD and P5CS to increase both reactive oxygen species scavenging and proline levels. In addition, BplMYB46 appears to be involved in controlling stomatal aperture to reduce water loss. Overexpression of BplMYB46 increases lignin deposition, secondary cell wall thickness and the expression of genes in secondary cell wall formation. Further analysis indicated that BplMYB46 binds to MYBCORE and AC‐box motifs and may directly activate the expression of genes involved in abiotic stress responses and secondary cell wall biosynthesis whose promoters contain these motifs. The transgenic BplMYB46‐overexpressing birch plants, which have improved salt and osmotic stress tolerance, higher lignin and cellulose content and lower hemicellulose content than the control, have potential applications in the forestry industry.

Comprehensive analysis of trihelix genes and their expression under biotic and abiotic stresses in Populus trichocarpa.

Trihelix genes play important roles in plant growth and development and responses to biotic and abiotic stresses. Here, we identified 56 full-length trihelix genes in Populus trichocarpa and classified them into five groups. Most genes within a given group had similar gene structures and conserved motifs. The trihelix genes were unequally distributed across 19 different linkage groups. Fifteen paralogous pairs were identified, 14 of which have undergone segmental duplication events. Promoter cis-element analysis indicated that most trihelix genes contain stress- or phytohormone-related cis-elements. The expression profiles of the trihelix genes suggest that they are primarily expressed in leaves and roots. Quantitative real-time reverse transcription polymerase chain reaction analysis indicated that members of the trihelix gene family are significantly induced in response to osmotic, abscisic acid, salicylic acid, methyl jasmonate and pathogen infection. PtrGT10 was identified as a target gene of miR172d, which is involved in the osmotic response. Repression of PtrGT10 could increase reactive oxygen species scavenging ability and decrease cell death. This study provides novel insights into the phylogenetic relationships and functions of the P. trichocarpa trihelix genes, which will aid future functional studies investigating the divergent roles of trihelix genes belonging to other species.

Overexpression of ThGSTZ1 from Tamarix hispida improves tolerance to exogenous ABA and methyl viologen

Key messageMolecular analysis of a zeta subfamilyGSTgene fromT. hispidainvolved in ABA and methyl viologen tolerance in transgenic Arabidopsis andTamarix.AbstractGlutathione S-transferase (GST) genes are important for the improvement of plant abiotic stress tolerance, and our previous study demonstrated that the ThGSTZ1 gene from Tamarix hispida improves plant salt and drought tolerance. To further understand the role of ThGSTZ1 in the response of plants to abscisic acid (ABA) and oxidative stress, three ThGSTZ1-overexpressing transgenic Arabidopsis thaliana lines were analyzed in the current study. The results showed that the transgenic lines exhibited higher biomass accumulation, higher activities of GST and other protective enzymes, and less reactive oxygen species (ROS) and cell damage than wild-type (WT) plants under ABA and methyl viologen (MV) stress. In addition, the analysis of a transgenic T. hispida line transiently expressing ThGSTZ1 confirmed these results. The activities of GST, glutathione peroxidase, and superoxide dismutase were markedly higher in the ThGSTZ1-overexpressing lines compared with the control lines under both ABA and MV treatments, and the transgenic lines also exhibited a lower degree of electrolyte leakage (EL) and a decreased H2O2 content. All these results suggested that ThGSTZ1 can also improve plant ABA and oxidation tolerance by regulating ROS metabolism and that ThGSTZ1 represents an excellent candidate gene for molecular breeding to increase plant stress tolerance.

Characterization and expression analysis of histone deacetylases family RPD3/HDA1 in Populus trichocarpa

Histone deacetylases (HDACs) catalyze histone deacetylation and play an important role in suppression of gene transcription in multiple cellular processes. HDACs are widely distributed in eukaryotes, however, detailed characterrization of HDACs in woody plants is not available. In this study, the sequences of reduced potassium dependency 3/histone deacetylase 1 (RPD3/HDA1) family proteins in black cottonwood (Populus trichocarpa Torr. & A. Gray) were characterized and their expression patterns in response to cold and salt stresses were determined. The RPD3/HDA1 proteins had conserved HDAC domains and can be divided into three classes based on sequence similarity and phylogenetic analysis. The transcripts of the HDAC genes were detected in different amounts in leaves, stems, and roots. The expressions of HDAC genes HDA902, HDA903, HDA904, HDA909, and HDA912 were up-regulated in a cold stress. Interestingly, in a salt stress, most of the HDAC genes were down-regulated. These results indicate that the poplar HDAC genes were regulated by the cold and salt stresses, and the members of the RPD3/HDA1 family play a role in stress responses.

Identification of novel cis-elements bound by BplMYB46 involved in abiotic stress responses and secondary wall deposition: Identification of novel cis-elements bound by BplMYB46

Transcription factors (TFs) play vital roles in various biological processes by binding to cis-acting elements to control expressions of their target genes. The MYB TF BplMYB46, from Betula platyphylla, is involved in abiotic stress responses and secondary wall deposition. In the present study, we used a TF-centered yeast one-hybrid technology (TF-centered Y1H) to identify the cis-acting elements bound by BplMYB46. We screened a short-insert random library and identified three cis-elements bound by BplMYB46: an E-box (CA(A/T/C)(A/G/C)TG) and two novel motifs, a TC-box (T(G/A)TCG(C/G)) and a GT-box (A(G/T)T(A/C)GT(T/G)C). Chromatin immunoprecipitation (ChIP) and effector-reporter coexpression assays in Nicotiana tabacum confirmed binding of BplMYB46 to the TC-box, GT-box, and E-box motifs in the promoters of the phenylalanine ammonia lyase (PAL), peroxidase (POD), and superoxide dismutase (SOD) genes, which function in abiotic stress tolerance and secondary wall biosynthesis. This finding improves our understanding of potential regulatory mechanisms in the response to abiotic stress and secondary wall deposition of BplMYB46 in B. platyphylla.

Identification, phylogeny, and transcript profiling of aquaporin genes in response to abiotic stress in Tamarix hispida

Aquaporins belong to the highly conserved major intrinsic protein family and are involved in the transcellular membrane transport of water and other small solutes. However, there has been little work on cloning aquaporin (AQP) family genes and characterizing their functions in plants under various environmental stimuli. In this study, a total of 18 full-length AQP genes were identified in Tamarix hispida, a woody halophyte. Sequence analysis showed that most of these AQP proteins have six transmembrane domains connected by five loops. Phylogenetic analysis revealed that the members of the AQP family can be divided into four groups based on their structural characteristics, including plasma membrane intrinsic proteins (PIPs), tonoplast intrinsic proteins (TIPs), NOD26-like intrinsic proteins (NIPs), and small basic intrinsic proteins (SIPs). Furthermore, the expression profiles of AQP genes were analyzed in the roots, stems, and leaves under salinity, drought, heavy metal, and abscisic acid (ABA) treatments using real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) assays. The results demonstrated that the AQP were involved in abiotic stress responses, indicating that they play important roles in response to abiotic stress and are involved in ABA-dependent stress-signaling pathways. These data will be useful to elucidate the complexity of the AQP gene family and the molecular mechanisms of abiotic stress tolerance.

Gene expression profiles in different stem internodes reveal the genetic regulation of primary and secondary stem development in Betula platyphylla

Secondary growth of stems is an important process for the radial increase of trees. To gain an insight into the molecular mechanisms underlying stem development from primary to secondary growth and to provide information for molecular research and breeding in Betula platyphylla (birch), the gene expression profiles of material from the first, third, and fifth internodes (IN) of 3-month-old seedlings were analyzed. Compared with the first IN, 177 genes were up-regulated and 157 genes down-regulated in the third IN; in the fifth IN, 180 genes were up-regulated and 275 genes were down-regulated. The expressions of 24 genes were up-regulated and 6 genes were down-regulated in the fifth IN relative to the third IN. The differentially expressed genes were annotated as having roles in cambium, xylem, and phloem development and formation; including cell wall expansion, cellulose biosynthesis, lignin biosynthesis and deposition, xylem extension, cell wall modification, and growth hormone responses. The expressions of genes related to cell wall expansion and cellulose biosynthesis in the primary cell wall were down-regulated in the third and fifth IN relative to the first IN. Genes involved in lignin biosynthesis, xylem extension, and cellulose synthesis in the secondary cell wall were up-regulated in the third and fifth IN relative to the first IN. These results described the patterns of gene expression during stem development in birch and provided candidate genes for further functional characterization.