Tobacco Overexpressing Research Articles

EgPHI-1, a PHOSPHATE-INDUCED-1 gene from Eucalyptus globulus, is involved in shoot growth, xylem fiber length and secondary cell wall properties.

EgPHI-1 is a member of PHI-1/EXO/EXL protein family. Its overexpression in tobacco resulted in changes in biomass partitioning, xylem fiber length, secondary cell wall thickening and composition, and lignification. Here, we report the functional characterization of a PHOSPHATE-INDUCED PROTEIN 1 homologue showing differential expression in xylem cells from Eucalyptus species of contrasting phenotypes for wood quality and growth traits. Our results indicated that this gene is a member of the PHI-1/EXO/EXL family. Analysis of the promoter cis-acting regulatory elements and expression responses to different treatments revealed that the Eucalyptus globulus PHI-1 (EgPHI-1) is transcriptionally regulated by auxin, cytokinin, wounding and drought. EgPHI-1 overexpression in transgenic tobacco changed the partitioning of biomass, favoring its allocation to shoots in detriment of roots. The stem of the transgenic plants showed longer xylem fibers and reduced cellulose content, while the leaf xylem had enhanced secondary cell wall thickness. UV microspectrophotometry of individual cell wall layers of fibers and vessels has shown that the transgenic plants exhibit differences in the lignification of S2 layer in both cell types. Taken together, the results suggest that EgPHI-1 mediates the elongation of secondary xylem fibers, secondary cell wall thickening and composition, and lignification, making it an attractive target for biotechnological applications in forestry and biofuel crops.

Characterization and overexpression of sterol Δ22-desaturase, a key enzyme modulates the biosyntheses of stigmasterol and withanolides in Withania somnifera (L.) Dunal

Withanolides constitute an extensive and vital class of metabolites displaying wide array of structural and therapeutic properties with unique side-chain modifications. These show diversified scaffolds and are promising pharmaceutical molecules with well documented anti-inflammatory and anti-cancer properties. Sterols are dynamic class of compounds and essential molecules having structural and functional significance. These contribute to the synthesis of withanolides by providing structural precursors. In this context, we have characterized sterol Δ22-desaturase from Withania somnifera and also functionally validating it by confirming its desaturase nature in conjunction with quantitative real-time expression profiling and metabolite evaluation. Further, transgenic hairy roots of W. somnifera displayed a higher accumulation of stigmasterol and withanolides. The increase in chemical constituents was concomitant with an increased gene copy number predicted via Southern blotting. Additionally, transgenic lines of tobacco over-expressing WsCYP710A11 displayed a substantial increase in its expression, corroborating well with enhanced stigmasterol content. Characterization of CYP710A11 from W. somnifera and its homologous transgenic expression has demonstrated its role in the regulation of withanolides biosynthesis. It also exhibited a differential transcriptional profile in response to exogenous elicitations. These empirical findings suggest the crucial role of CYP710A11 in stigmasterol biosynthesis. This in turn has implications for the overproduction of withanolides via pathway channelling.

Transcriptome and Flavonoids Metabolomic Analysis Identifies Regulatory Networks and Hub Genes in Black and White Fruits of Lycium ruthenicum Murray.

Lycium ruthenicum Murry. is a highly nutritional cash crop due to its fruit abundant anthocyanins. To understand the complex metabolic networks underlying the color formation in black and white fruits of L. ruthenicum, we conducted transcriptome and flavonoid metabolic profiling to identify the candidate genes possibly involved in flavonoid biosynthesis. As a result, 147 flavonoids were identified and there was almost no anthocyanin in white fruits, while luteolin, kaempferol, and quercetin derivatives showed markedly higher abundance. Furthermore, applying weighted gene co-expression network analyses, 3 MYB, 2 bHLH, 1WRKY and 1 NAC transcription factor, associated with anthocyanin biosynthesis were identified. A bHLH transcription factor, LrAN1b showed the greatest correlations with anthocyanin accumulation with no expression in white fruits. In addition, gene function analysis and qRT-PCR experiments identified a new activated anthocyanin MYB transcription factor designed as LrAN2-like. Yeast two-hybrid and transient tobacco overexpression experiments showed that LrAN1b could interact with LrAN2-like and LrAN11 to form MBW complex to activate the anthocyanin pathway. The yeast one-hybrid experiment indicated that LrAN2-like bonded anthocyanin structural gene LrDFR and LrANS promoters. Heterologous expression of LrAN1b in tobacco can significantly increase the anthocyanin content of tobacco florals and capsules, and activate anthocyanin synthesis related genes. Taken together, an anthocyanin regulatory network model in L. ruthenicum fruit was proposed firstly and we speculate that the white fruit phenotype was due to abnormal expression of LrAN1b. The findings provide new insight into the underlying mechanism of flavonoids, laying the foundation for future functional and molecular biological research in L. ruthenicum.

SMRT and Illumina RNA sequencing reveal novel insights into the heat stress response and crosstalk with leaf senescence in tall fescue

BackgroundAs a cool-season grass species, tall fescue (Festuca arundinacea) is challenged by increasing temperatures. Heat acclimation or activation of leaf senescence, are two main strategies when tall fescue is exposed to heat stress (HS). However, lacking a genome sequence, the complexity of hexaploidy nature, and the short read of second-generation sequencing hinder a comprehensive understanding of the mechanism. This study aims to characterize the molecular mechanism of heat adaptation and heat-induced senescence at transcriptional and post-transcriptional levels.ResultsTranscriptome of heat-treated (1 h and 72 h) and senescent leaves of tall fescue were generated by combining single-molecular real-time and Illumina sequencing. In total, 4076; 6917, and 11,918 differentially expressed genes (DEGs) were induced by short- and long-term heat stress (HS), and senescence, respectively. Venn and bioinformatics analyses of DEGs showed that short-term HS strongly activated heat shock proteins (Hsps) and heat shock factors (Hsfs), as well as specifically activated FK506-binding proteins (FKBPs), calcium signaling genes, glutathione S-transferase genes, photosynthesis-related genes, and phytohormone signaling genes. By contrast, long-term HS shared most of DEGs with senescence, including the up-regulated chlorophyll catabolic genes, phytohormone synthesis/degradation genes, stress-related genes, and NACs, and the down-regulated photosynthesis-related genes, FKBPs, and catalases. Subsequently, transient overexpression in tobacco showed that FaHsfA2a (up-regulated specifically by short-term HS) reduced cell membrane damages caused by HS, but FaNAC029 and FaNAM-B1 (up-regulated by long-term HS and senescence) increased the damages. Besides, alternative splicing was widely observed in HS and senescence responsive genes, including Hsps, Hsfs, and phytohormone signaling/synthesis genes.ConclusionsThe short-term HS can stimulate gene responses and improve thermotolerance, but long-term HS is a damage and may accelerate leaf senescence. These results contribute to our understanding of the molecular mechanism underlying heat adaptation and heat-induced senescence.

Expression of Glutamate Dehydrogenase Genes of Mulberry in Morus alba L. and Transgenic Tobacco in Relation to Biotic and Abiotic Stresses

Mulberry (Morus alba L.) is an economically and ecologically important plant with strong stress resistance. Glutamate dehydrogenase (GDH) can catalyze the reversible reaction of α-ketoglutarate and $${\text{NH}}_{4}^{ + }$$ to glutamate, and plays an important role in resistance to adverse conditions. In this study, two MaGDH genes were cloned, and their biological characteristics studied. The results showed that the MaGDH1 and MaGDH2 protein were localized in the cell membrane, cytoplasm, chloroplasts, and MaGDH1 also localized in mitochondria. These proteins were expressed mainly in flowers. During seed germination, MaGDH1 and MaGDH2 expression levels were the highest when the radicle penetrated the seed coat. Stress tests suggested that MaGDH genes participate in responses to abiotic stress, and that the MaGDH2 protein had an inhibitory effect on the fungal pathogen Ciboria shiraiana, extracted from mulberry fruit. MaGDH overexpression in transgenic tobacco increased the solubility of proteins and chlorophyll content of transgenic tobacco, improving tolerance to salt and drought stresses.

Enhancement of methyl salicylate accumulation promotes early flowering in transgenic tobacco plants by overexpressing a carboxymethyl transferase (SAMT) gene from Lycium chinense

Flowering time is an important agronomic trait, which is of great significance to the plant growth process. Salicylic acid (SA) is a key hormone that regulates plant growth and development. It can be converted into methyl salicylate (MeSA) catalyzed by SA carboxymethyl transferase (SAMT). Previous studies showed that SA was related to the regulation of plant flowering; however, the specific mechanism associated with this process remained to be further elucidated. In this study, the transgenic tobacco overexpressing LcSAMT gene and WT tobacco were cultivated to observe the flowering status of plant. The biomass and growth of tobaccos were recorded at different stages of plant growth. Endogenous MeSA content, phenylalanine ammonia-lyase (PAL) activity, and anthocyanin content were determined in transgenic tobacco plants during flowering stage. It was observed that the flowering time of transgenic tobacco was usually 6–7 days earlier than WT tobacco, and a higher endogenous MeSA content, PAL activity, and anthocyanin content were found in transgenic tobacco plants during flowering stage. Quantitative real-time PCR (qPCR) and transcriptome analysis were also performed in this study. There were 3924 differentially expressed genes (DEGs) identified between WT and transgenic tobacco plants by transcriptome analysis. These DEGs were primarily associated with plant hormone signal transduction pathways, plant pathogen interaction pathways, and mitogen-activated protein kinase (MAPK) pathways. QPCR analysis of 15 DEGs revealed that these genes were markedly upregulated in transgenic tobacco which was in accordance with transcriptome sequencing results. This study indicated that the SAMT gene-mediated SA homeostasis played important roles in the regulation of tobacco flowering.

Contrasting Responses to Stress Displayed by Tobacco Overexpressing an Algal Plastid Terminal Oxidase in the Chloroplast.

The plastid terminal oxidase (PTOX) – an interfacial diiron carboxylate protein found in the thylakoid membranes of chloroplasts – oxidizes plastoquinol and reduces molecular oxygen to water. It is believed to play a physiologically important role in the response of some plant species to light and salt (NaCl) stress by diverting excess electrons to oxygen thereby protecting photosystem II (PSII) from photodamage. PTOX is therefore a candidate for engineering stress tolerance in crop plants. Previously, we used chloroplast transformation technology to over express PTOX1 from the green alga Chlamydomonas reinhardtii in tobacco (generating line Nt-PTOX-OE). Contrary to expectation, growth of Nt-PTOX-OE plants was more sensitive to light stress. Here we have examined in detail the effects of PTOX1 on photosynthesis in Nt-PTOX-OE tobacco plants grown at two different light intensities. Under ‘low light’ (50 μmol photons m–2 s–1) conditions, Nt-PTOX-OE and WT plants showed similar photosynthetic activities. In contrast, under ‘high light’ (125 μmol photons m–2 s–1) conditions, Nt-PTOX-OE showed less PSII activity than WT while photosystem I (PSI) activity was unaffected. Nt-PTOX-OE grown under high light also failed to increase the chlorophyll a/b ratio and the maximum rate of CO2 assimilation compared to low-light grown plants, suggesting a defect in acclimation. In contrast, Nt-PTOX-OE plants showed much better germination, root length, and shoot biomass accumulation than WT when exposed to high levels of NaCl and showed better recovery and less chlorophyll bleaching after NaCl stress when grown hydroponically. Overall, our results strengthen the link between PTOX and the resistance of plants to salt stress.

UGT74AF3 enzymes specifically catalyze the glucosylation of 4-hydroxy-2,5-dimethylfuran-3(2H)-one, an important volatile compound in Camellia sinensis

4-Hydroxy-2,5-dimethylfuran-3(2H)-one (HDMF) is an important odorant in some fruits, and is proposed to play a crucial role in the caramel-like notes of some teas. However, its biosynthesis and metabolism in tea plants are still unknown. Here, HDMF glucoside was unambiguously identified as a native metabolite in tea plants. A novel glucosyltransferase UGT74AF3a and its allelic protein UGT74AF3b specifically catalyzed the glucosylation of HDMF and the commercially important structural homologues 2 (or 5)-ethyl-4-hydroxy-5 (or 2)-methylfuran-3(2H)-one (EHMF) and 4-hydroxy-5-methylfuran-3(2H)-one (HMF) to their corresponding β-D-glucosides. Site-directed mutagenesis of UGT74AF3b to introduce a single A456V mutation resulted in improved HDMF and EHMF glucosylation activity and affected the sugar donor preference compared with that of the wild-type control enzyme. The accumulation of HDMF glucoside was consistent with the transcript levels of UGT74AF3 in different tea cultivars. In addition, transient UGT74AF3a overexpression in tobacco significantly increased the HDMF glucoside contents, and downregulation of UGT74AF3 transcripts in tea leaves significantly reduced the concentration of HDMF glucoside compared with the levels in the controls. The identification of HDMF glucoside in the tea plant and the discovery of a novel-specific UDP-glucose:HDMF glucosyltransferase in tea plants provide the foundation for improvement of tea flavor and the biotechnological production of HDMF glucoside.

The JcWRKY tobacco transgenics showed improved photosynthetic efficiency and wax accumulation during salinity

Salinity is one of the major factors negatively affecting crop productivity. WRKY transcription factors (TFs) are involved in salicylic acid (SA) mediated cellular reactive oxygen species homeostasis in response to different stresses, including salinity. Therefore, the effect of NaCl, NaCl + SA and SA treatments on different photosynthesis-related parameters and wax metabolites were studied in the Jatropha curcas WRKY (JcWRKY) overexpressing tobacco lines. JcWRKY transgenics showed improved photosynthesis rate, stomatal conductance, intercellular CO2 concentration/ambient CO2 concentration ratio (Ci/Ca ratio), electron transport rate (ETR), photosynthesis efficiency (Fv/Fm), photochemical quenching (qP), non-photochemical quenching (NPQ) and quantum yield of PSII electron transport (ΦPSII) in response to salinity stress, while exogenous SA application had subtle effect on these parameters. Alkane, the major constituent of wax showed maximum accumulation in transgenics exposed to NaCl. Other wax components like fatty alcohol, carboxylic acid and fatty acid were also higher in transgenics with NaCl + SA and SA treatments. Interestingly, the transgenics showed a higher number of open stomata in treated plants as compared to wild type (WT), indicating less perception of stress by the transgenics. Improved salinity tolerance in JcWRKY overexpressing tobacco transgenics is associated with photosynthetic efficiency and wax accumulation, mediated by efficient SA signalling. The transgenics showed differential regulation of genes related to photosynthesis (NtCab40, NtLhcb5 and NtRca1), wax accumulation (NtWIN1) and stomatal regulation (NtMUTE, NtMYB-like, NtNCED3-2 and NtPIF3). The present study indicates that JcWRKY is a potential TF facilitating improved photosynthesis with the wax metabolic co-ordination in transgenics during stress.

PpNAC187 Enhances Lignin Synthesis in 'Whangkeumbae' Pear (Pyrus pyrifolia) 'Hard-End' Fruit.

A disorder in pears that is known as ‘hard-end’ fruit affects the appearance, edible quality, and market value of pear fruit. RNA-Seq was carried out on the calyx end of ‘Whangkeumbae’ pear fruit with and without the hard-end symptom to explore the mechanism underlying the formation of hard-end. The results indicated that the genes in the phenylpropanoid pathway affecting lignification were up-regulated in hard-end fruit. An analysis of differentially expressed genes (DEGs) identified three NAC transcription factors, and RT-qPCR analysis of PpNAC138, PpNAC186, and PpNAC187 confirmed that PpNAC187 gene expression was correlated with the hard-end disorder in pear fruit. A transient increase in PpNAC187 was observed in the calyx end of ‘Whangkeumbae’ fruit when they began to exhibit hard-end symptom. Concomitantly, the higher level of PpCCR and PpCOMT transcripts was observed, which are the key genes in lignin biosynthesis. Notably, lignin content in the stem and leaf tissues of transgenic tobacco overexpressing PpNAC187 was significantly higher than in the control plants that were transformed with an empty vector. Furthermore, transgenic tobacco overexpressing PpNAC187 had a larger number of xylem vessel elements. The results of this study confirmed that PpNAC187 functions in inducing lignification in pear fruit during the development of the hard-end disorder.

Identification of miRNA Targets by AtFT Overexpression in Tobacco

The onset of flowering is regulated by complex gene networks that integrate multiple genetic cues to floral transition in plants. The highly conserved florigen FLOWERING LOCUS T (FT) functions at the core of this process. Here, we demonstrate that overexpression of Arabidopsis FT in tobacco (oxAtFT) leads to precocious flowering in a photoperiod-independent manner. Strikingly altered phenotypes such as stunted growth, increased number of axillary branches, change in leaf morphology, and elongated style were observed in oxAtFT transgenic tobacco plants. In addition, the generation time of oxAtFT tobacco was drastically reduced by 8–10 weeks. We observed defects in lateral root formation in both oxAtFT tobacco and oxFT Arabidopsis lines. Interestingly, oxAtFT tobacco displayed significant heat and drought tolerance as compared to the wild type. Through small RNA profiling, we observed the upregulation of a conserved microRNA (nta-miR393a-5p) of miR393 family that targets the TRANSPORT INHIBITOR RESPONSE 1-like (TIR1) gene. Overall, the heterologous expression of AtFT in tobacco may have potential in imparting abiotic stress tolerance in other plant species through plant biotechnology–based approaches.

Genome-Wide Identification and Characterization of Cucumber BPC Transcription Factors and Their Responses to Abiotic Stresses and Exogenous Phytohormones.

BASIC PENTACYSTEINE (BPC) is a small transcription factor family that functions in diverse growth and development processes in plants. However, the roles of BPCs in plants, especially cucumber (Cucumis sativus L.), in response to abiotic stress and exogenous phytohormones are still unclear. Here, we identified four BPC genes in the cucumber genome, and classified them into two groups according to phylogenetic analysis. We also investigated the gene structures and detected five conserved motifs in these CsBPCs. Tissue expression pattern analysis revealed that the four CsBPCs were expressed ubiquitously in both vegetative and reproductive organs. Additionally, the transcriptional levels of the four CsBPCs were induced by various abiotic stress and hormone treatments. Overexpression of CsBPC2 in tobacco (Nicotiana tabacum) inhibited seed germination under saline, polyethylene glycol, and abscisic acid (ABA) conditions. The results suggest that the CsBPC genes may play crucial roles in cucumber growth and development, as well as responses to abiotic stresses and plant hormones. CsBPC2 overexpression in tobacco negatively affected seed germination under hyperosmotic conditions. Additionally, CsBPC2 functioned in ABA-inhibited seed germination and hypersensitivity to ABA-mediated responses. Our results provide fundamental information for further research on the biological functions of BPCs in development and abiotic stress responses in cucumber and other plant species.

Transcription factor TERF1 regulates nuclear genes expression through miRNAs in tobacco under drought stress condition

Ethylene is an important phytohormone that regulates plant response to drought stress. ETHYLENE RESPONSE FACTOR 1 (ERF1), a transcription factor of ERF/AP2 family, plays an important role in activating ethylene signaling pathway through binding the GCC box in the promoters of ethylene responsive genes. Although we know some protein-coding genes regulated by ERF1, we know nothing about how ERF1 regulates the expression of miRNAs. We utilized the tobacco overexpressing TOMATO ETHYLENE RESPONSE FACTOR1 (TERF1), an ERF1 transcription factor isolated from tomato, to investigate the miRNAs expression profile under natural dehydration condition by method of qRT-PCR. Results show that 25 miRNAs are significantly induced and only 10 miRNAs are significantly repressed by TERF1. Binding sites for ERF transcription factors are observed in six upregulated miRNAs and the core genes involved in the processing of pre-miRNA are also significantly induced by TERF1. We predicted the target genes regulated by the differentially expressed miRNAs by the on-line programme of psRNATarget. Gene ontology (GO) analysis shows that the significantly enriched biological processes for the target genes regulated by the downregulated miRNAs are located in chloroplast. We also predicted the important regulatory genes regulated by the differentially expressed miRNAs, including transcription factors, kinases and phosphatases. Our research provides novel mechanism for regulation of nuclear genes expression by TERF1 at posttranscriptional level under drought stress condition.

SpPKE1, a Multiple Stress-Responsive Gene Confers Salt Tolerance in Tomato and Tobacco.

Understanding the mechanism of abiotic-tolerance and producing germplasm of abiotic tolerance are important in plant research. Wild species often show more tolerance of environmental stress factors than their cultivated counterparts. Genes from wild species show potential abilities to improve abiotic resistance in cultivated species. Here, a tomato proline-, lysine-, and glutamic-rich type gene SpPKE1 was isolated from abiotic-resistant species (Solanum pennellii LA0716) for over-expression in tomato and tobacco for salt tolerance. The protein encoded by SpPKE1 was predominantly localized in the cytoplasm in tobacco. SpPKE1 and SlPKE1 (from cultivated species S. lycopersicum cv. M82) shared 89.7% similarity in amino acid sequences and their transcripts abundance in flowers and fruits was reduced by the imposition of drought or oxidative stress and the exogenous supply of abscisic acid. The DNA of the PKE1 promoter was highly methylated in fruit and leaf, and the methylation of the coding sequence in leaf was significantly higher than that in fruit at different development stages. The over-expression of SpPKE1 under the control of a CaMV (Cauliflower Mosaic Virus) 35S promoter in transgenic tomato and tobacco plants enhanced their tolerance to salt stress. PKE1 was downregulated by abiotic stresses but enhanced the plant’s salt stress tolerance. Therefore, this gene may be involved in post-transcriptional regulation and may be an important candidate for molecular breeding of salt-tolerant plants.

In silico characterization of SAMdC from Pokkali rice and its overexpression in transgenic tobacco

Polyamines help to mitigate salt stress in plants. SAMdC or S-adenosyl methionine decarboxylase is a crucial enzyme in the polyamine biosynthetic pathway. Overexpressing the samdc gene of Pokkali (a salt-tolerant Indica rice landrace) in the model plant tobacco through Agrobacterium-mediated transformation resulted in tobacco plants capable of withstanding high salinity (200 mM) stress. The transgenic tobacco plants maintained a steady level of the higher polyamines and resisted ionic imbalance under salt stress. Since appropriate binding of the substrate with the enzyme is essential for an enzymatic reaction, we performed molecular docking experiment of SAMdC enzyme of Pokkali rice and tobacco to get an idea about its commonality in the two plants in the backdrop of Japonica rice and Arabidopsis. In silico characterization of SAMdC revealed that the enzyme used the same substrate in Pokkali rice and tobacco, from where the gene is introgressed and where it is overexpressed.

Overexpression of a pathogenesis-related gene NbHIN1 confers resistance to Tobacco Mosaic Virus in Nicotiana benthamiana by potentially activating the jasmonic acid signaling pathway

Harpin proteins secreted by plant-pathogenic gram-negative bacteria induce diverse plant defenses against different pathogens. Harpin-induced 1 (HIN1) gene highly induced in tobacco after application of Harpin protein is involved in a common plant defense pathway. However, the role of HIN1 against Tobacco mosaic virus (TMV) remains unknown. In this study, we functionally characterized the Nicotiana benthamiana HIN1 (NbHIN1) gene and generated the transgenic tobacco overexpressing the NbHIN1 gene. In a subcellular localization experiment, we found that NbHIN1 localized in the plasma membrane and cytosol. Overexpression of NbHIN1 did not lead to observed phenotype compared to wild type tobacco plant. However, the NbHIN1 overexpressing tobacco plant exhibited significantly enhanced resistance to TMV infection. Moreover, RNA-sequencing revealed the transcriptomic profiling of NbHIN1 overexpression and highlighted the primary effects on the genes in the processes related to biosynthesis of amino acids, plant-pathogen interaction and RNA transport. We also found that overexpression of NbHIN1 highly induced the expression of NbRAB11, suggesting that jasmonic acid signaling pathway might be involved in TMV resistance. Taken together, for the first time we demonstrated that overexpressing a pathogenesis-related gene NbHIN1 in N. benthamiana significantly enhances the TMV resistance, providing a potential mechanism that will enable us to engineer tobacco with improved TMV resistance in the future.

DNA Repair Gene ZmRAD51A Improves Rice and Arabidopsis Resistance to Disease.

RAD51 (DNA repair gene) family genes play ubiquitous roles in immune response among species from plants to mammals. In this study, we cloned the ZmRAD51A gene (a member of RAD51) in maize and generated ZmRAD51A overexpression (ZmRAD51A-OE) in rice, tobacco, and Arabidopsis. The expression level of ZmRAD51A was remarkably induced by salicylic acid (SA) application in maize, and the transient overexpression of ZmRAD51A in tobacco induced a hypersensitive response. The disease resistance was significantly enhanced in ZmRAD51A- OE (overexpressing) plants, triggering an increased expression of defense-related genes. High-performance liquid chromatography (HPLC) analysis showed that, compared to control lines, ZmRAD51A-OE in rice plants resulted in higher SA levels, and conferred rice plants resistance to Magnaporthe oryzae. Moreover, the ZmRAD51A-OE Arabidopsis plants displayed increased resistance to Pseudomonas syringae pv. tomato DC3000 when compared to wild types. Together, our results provide the evidence that, for the first time, the maize DNA repair gene ZmRAD51A plays an important role in in disease resistance.

The maize late embryogenesis abundant protein ZmDHN13 positively regulates copper tolerance in transgenic yeast and tobacco

Late embryogenesis abundant (LEA) proteins accumulate in the late stage of plant seed development, and are upregulated in most plants during drought, cold, heat, or salinity stress. LEA proteins can be classified by amino-acid sequence into seven groups. Dehydrins belong to LEA protein group II. In previous studies, the maize KS type dehydrin ZmDHN13 increased the tolerance of transgenic tobacco to oxidative stress. In the present study, ZmDHN13 was identified under copper stress conditions, and the protein was then characterized using transgenic yeast and tobacco plants to investigate its functions. ZmDHN13 bound Cu2+. Its overexpression in transgenic tobacco conferred tolerance to copper stress by binding metals and reducing the accumulation of reactive oxygen species (ROS). Three conserved domains displayed a cooperative effect under copper stress conditions.

Transcriptome analysis of 1- and 3-year-old Panax notoginseng roots and functional characterization of saponin biosynthetic genes DS and CYP716A47-like.

Transcriptome analysis revealed high expression of saponin biosynthetic genes may account for highly accumulated saponins in 3-year-old Panax notoginseng roots and DS and CYP716A47 - like were functionally verified by transgenic tobacco. Panax notoginseng is a well-known traditional medical herb that contains bioactive compounds known as saponins. Three major dammarene-type triterpene saponins including R1, Rb1, and Rg1 were found to be highly accumulated in the roots of 3-year-old plants when compared to those of 1-year-old plants. However, the underlying cellular mechanism is poorly understood. In this study, transcriptome analysis revealed that most genes involved in saponin biosynthesis in P. notoginseng roots augmented during their growth periods. The analysis of the KEGG pathway indicated that the primary metabolism, cell growth, and differentiation were less active in the roots of 3-year-old plant; however, secondary metabolisms were enhanced, thus providing molecular evidence for the harvesting of P. notoginseng roots in the 3rd year of growth. Furthermore, the functional role of DS and CYP716A47-like, two of the candidate genes involved in saponin biosynthesis isolated from P. notoginseng, were verified via overexpression in cultivated tobacco. Approximately, 0.325µgg-1 of dammarenediol-II and 0.320µgg-1 of protopanaxadiol were recorded in the dry leaves of transgenic tobacco overexpressed with DS and both DS and CYP716A47-like, respectively. This study provides insights into the molecular mechanisms for saponin accumulation in P. notoginseng roots during its growth period and paves a promising way to produce dammarenediol-II and protopanaxadiol via transgenic techniques.

TERF1 Regulates Nuclear Gene Expression Through Chloroplast Retrograde Signals

Ethylene is an important phytohormone that regulates many important biological processes in plant. ERF (ethylene response factor) proteins are key transcription factors that activate the ethylene signaling pathway. However, our knowledge about the mechanism of the ERF transcription factors in regulating nuclear genes expression is limited. Retrograde signaling pathway in chloroplast is a novel kind of mechanism that regulates nuclear gene expression by different signals in plastid. Based on our former research we analyzed the components related with retrograde signaling from plastid in the transgenic tobacco overexpressing TERF1, a member of ERF family transcription factors, to elucidate the interaction between ethylene signaling pathway and different retrograde signaling pathway in plastid under normal growth condition. Results show that TERF1 regulates different retrograde signals in plastid and thus regulates different nuclear genes expression under normal growth condition. We propose a new mechanism that links ethylene signaling pathway and retrograde signaling pathway as well as new potential of TERF1 in regulating nuclear genes expression at post-transcriptional level.