Poplar Plants Research Articles

First Report of Erysiphe adunca Causing Foliar Powdery Mildew on Poplar in China

Hybrid poplars are important economic plants and sources of biofuel all over the world. Poplars are subjected to a wide range of diseases (Cellerino 1999). In September 2019, powdery mildew was observed on the hybrid poplar Populus davidiana × P. alba var. pyramidalis, a fast-growing elite poplar cultivar widely planted in the northeast of China, in one poplar nursery room in the Northeast Forestry University in China (45.7266°N, 126.6380°E). Disease incidence during September and October was only 5 to 15% but gradually increased to 100% during November and December. The symptom of white circular powdery spots with diameters of up to 12 mm appeared on the upper surface of the mature leaves of poplar plants over 40 days old. Under nursery conditions (50 to 65% humidity, 20 to 25°C, and 16-h/8-h day/night cycles), the diameter of a mildew spot could develop from merely visible to 10 mm in 3 days. The mildew site was observed with a stereomicroscope (Leica DVM6S, Germany). A single fungal species demonstrating characteristics of the obligate pathogens Erysiphe spp. was observed. The conidiophores grew straight upward on the leaf surface until reaching 102 to 165 μm in length (n = 30). Then, even-looking, ellipse-shaped conidia were developed, forming a singular, curved string of up to tens of conidia, on top of each conidiophore. Conidia (n = 30) measured 29.4 to 35.1 × 18.1 to 21.0 μm. Appressoria were lobed or nipple-shaped and appeared in pairs or singular. The germ tubes were slightly curly unbranched tubes pointing at one direction near the long axis of the conidium. We carefully collected the mildew by scraping it off the upper surfaces of fresh diseased leaves with a scalpel and extracted the genomic DNA. The internal transcribed spacer rDNA region was amplified according to the method of Siahmard et al. (2017). The resulting nucleotide sequence (deposited in GenBank as MN998572) displayed 99.84% identity with that of Erysiphe adunca (KY653179.1). Hence, we identified the causal pathogen of this poplar powdery mildew as an E. adunca strain. Furthermore, an Mcm7 sequence (deposited as MN998573) and a Chs sequence (deposited as MN998574) were cloned according to the method of Ellingham et al. (2019). A voucher specimen and the sequence information have been deposited in the Northeast Forestry University Culture Collection (NECC) and named as NECCFP001. Pathogenicity was tested in the nursery, by merely placing healthy 40-day-old plants next to diseased plants (n = 10, with a 15 cm distance between plants), and by pressing the mildew sites of diseased leaves onto each mature leaf of the healthy plants (n = 10, also with 15 cm distance). The same symptoms appeared on heathy plants after 10 to 15 days, and 6 to 8 days, respectively. The induced mildew was confirmed morphologically to be NECCFP001 by microscopic examination. E. adunca occurs on aspen, willow, and Populus spp. throughout temperate regions of the Northern Hemisphere and forms fruiting bodies (chasmothecia) to overwinter (Horst 2001; Sinclair and Lyon 2005). However, chasmothecia were not observed in our case, suggesting no sexual morph under the nursery conditions. Incidents of powdery mildew on hybrid poplars caused by Phyllactinia populi had been reported in Jiangsu and Shandong provinces of China (Zhao et al. 2013). This is the first report of E. adunca causing poplar powdery mildew in Heilongjiang, China.

Genome-wide identification of plasma membrane aquaporin gene family in Populus and functional identification of PIP1;1 involved in osmotic stress

Aquaporins are the membrane channels that facilitate the transportation of water and small neutral molecules in plants. As such, they play an important role in stress tolerance. This function may be more pronounced in woody plants, which must efficiently absorb water and transport it over long distances. Therefore, a functional analysis of plasma membrane aquaporins (PIP) under drought stress can provide insights into the roles of these proteins in woody plants. In this work, we obtained 15 members of the PtPIP gene family in poplar. They formed two groups in a phylogenetic analysis. Gene structure, including intron phases, was found to be remarkably well conserved among members of this gene family. Five paralogous gene pairs were identified in the PtPIP family, and four of those pairs arose via whole genome duplication. A selection pressure analysis showed that PtPIP genes have undergone positive selection. The transcripts of six PtPIP genes including PtPIP1;1 were predominantly detected in the root. Transgenic poplar overexpressing PtoPIP1;1 was sensitive to osmotic stress, which phenocopied PagMYB216-overexpressing poplar plants. Transactivation and yeast one-hybrid assays showed that PagMYB216, whose homologs in other plants are induced by abiotic stresses, is able to activate PtoPIP1;1 expression through binding to the MBS motif in its promoter. Our results provide new information about PtoPIPs and their roles in response to abiotic stresses, which will be helpful for better understanding their functions in Populus.

PtKTI12 genes influence wobble uridine modifications and drought stress tolerance in hybrid poplar.

The multisubunit Elongator complex plays key roles in transcription by interacting with RNA polymerase II and chromatin modeling. Kti proteins have been identified as the auxiliary protein for the Elongator complex. However, our knowledge of Kti proteins in woody plants remains limited. In this study, in total 16 KTI gene homologs were identified in Populus trichocarpa. Among them, the two KTI12 candidates were named PtKTI12A and PtKTI12B. Although PtKTI12A and PtKTI12B were largely different in gene expression level and tissue specificity, both genes were induced by heat and drought stresses. PtKTI12A and PtKTI12B RNAi transgenic poplar plants showed reduced levels of modified nucleosides, in particular 5-carbamoylmethyluridine and 5-methoxycarbonylmethyl-2-thiouridine. Meanwhile, their tolerance to drought was improved when subjected to withdrawal of watering. Also, the protein products of PtKTI12A and PtKTI12B had similar subcellular localization and predicted tertiary structure. The results suggest that Kti12 proteins are involved in tRNA wobble uridine modification, stress response and drought stress tolerance in hybrid poplar.

Thaumatin-like protein(Pe-TLP)acts as a positive factor in transgenic poplars enhanced resistance to spots disease

Thaumantin-like proteins (TLPs), a class of pathogenesis–related (PR) proteins, have the potential to enhance plant resistance against fungal pathogens. They are thought to have different functions in other physiological processes. In this study, a TLP gene (PeTLP) was cloned from poplar cultivar (Populus deltoides × P. euramericana ‘Nanlin895’). According to bioinformatic analysis results about Pe-TLP (protein encoded by PeTLP gene), although typical thaumatin domain exists in its protein structure, Pe-TLP shares a close relationship with osmotin-like protein family according to KEGG metabolic pathway data. ATP assay showed that Pe-TLP had biological activity, but no direct fungicidal activity according to the result of assay of antifungal activity. Motif and structure differences may result in the poplar TLP protein not being able to effectively act on pathogens. And we grew transgenic Nanlin895 poplars over-expressed the PeTLP gene. Higher expression levels of TLP protein in transgenic poplar plants were observed using SDS-PAGE analysis. Although the Pe-TLP itself had no effect on pathogenic strain, transgenic plants with higher expression levels of PeTLP showed enhanced resistance against spots disease, and total proteins extracted from leaves of PeTLP over-expression lines showed obvious inhibitory effect on the growth of fungi under in vitro conditions. Our results suggest that the protein encoded by PeTLP functions as an elicitor of other anti-fungal proteins. Also, our findings provide a strong basis for developing a commercial poplar with the characteristics of spots disease resistance.

Identification and Functional Characterization of PtoMYB055 Involved in the Regulation of the Lignin Biosynthesis Pathway in Populus tomentosa

Wood, which is mainly composed of lignified secondary cell wall, is the most abundant biomass in woody plants. Previous studies have revealed that R2R3-type MYB transcription factors are important regulators of the formation of the secondary cell wall in vascular plants. In this study, we isolated the R2R3-type MYB transcription factor gene PtoMYB055, which is mainly expressed in xylem and phloem tissue, from Populus tomentosa and demonstrate that PtoMYB055 is a key regulator of lignin biosynthesis. PtoMYB055 as a transcriptional activator is localized to the nucleus. Overexpression of PtoMYB055 upregulates expression of lignin biosynthetic genes in transgenic poplar plants, resulting in ectopic deposition of lignin in phloem tissue and an increase in thickness of the secondary cell wall. In sum, PtoMYB055 is a transcriptional activator that is involved in regulating lignin biosynthesis during the formation of the secondary cell wall in poplar.

Transcription factor LkWOX4 is involved in adventitious root development in Larix kaempferi.

WUSCHEL-related homeobox4 (WOX4) plays important roles in vascular formation and adventitious root (AR) development. Here, we cloned the WOX4 from the AR of Larix kaempferi, whose cDNA is 1452bp in length and encodes 483 amino acids. LkWOX4 is mainly expressed in the layer formation area of the stem at 10days after cutting and its expression levels in the middles and ends of the ARs were higher than that in the AR tips. The fused protein LkWOX4-GFP localized in the nucleus. The heterologous overexpression of LkWOX4 in 84K poplar significantly increased AR numbers and decreased AR lengths. In LkWOX4 plants, the endogenous jasmonic acid and abscisic acid contents significantly decreased in stems, while the auxin, jasmonic acid and abscisic acid contents significantly increased in ARs. RNA-Seq of those LkWOX4 overexpression poplar plants showed that the expression of plant hormone signaling genes (ARF2, ARF3, ARF7 and ARF18), rooting-related transcription factors (WOX5, LBD29 and SCR) and root development-related genes (CYCD3, GRF1 and TAA1) were affected. Moreover, we found that LkWOX4 interacts with LkPAT18, LkACBP6, and LkCIP7 using yeast two hybrid screening. Thus, we found LkWOX4 involves in the AR initiation and development, which might be regulated through the IAA, JA and ABA signaling pathways.

Study on the flexural behavior of poplar beams externally strengthened by BFRP strips

Poplar plants with peculiar characteristics grow extensively and naturally in the Northern Hemisphere, and such poplar is the main timber material in Xinjiang, China. However, poplar has defects that affect its ultimate bearing capacity. This study investigates the flexural behavior of the timber members from poplar. Three lengths of basalt fiber-reinforced polymer (BFRP) strips were used to strengthen timber beams from poplar with rectangular cross-sections. From the experimental data, the timber beams strengthened by 1900-mm-long and 50-mm-wide BFRP strips showed the best ultimate bearing capacity and maximum deflection of mid-span compared with the other two strengthened beams. The increments were 77.8% and 110.1% compared with the control beam. The assumption “plane section remains plane” remains practicable in the poplar beams bonded by BFRP strips, and the neutral axis of the strengthened beams had a tendency to move toward the tensile zone. Furthermore, the processes of the flexural behavior of the specimens were simulated by finite element analysis modeling, but all the analyzed values of ultimate bearing capacity and deflection of the beams are higher than those of the experiment. These values can roughly be used as reference for experimental results. However, accurate modeling should be developed and modified in subsequent research on timber beams from Xinjiang poplar.

PtiCYP85A3, a BR C-6 Oxidase Gene, Plays a Critical Role in Brassinosteroid-Mediated Tension Wood Formation in Poplar.

In angiosperm trees, the gelatinous layer (G-layer) takes a great part of the fiber cell wall in the tension wood (TW). However, the mechanism underlying G-layer formation in poplar is largely unknown. In this work, we demonstrate that G-layer formation in poplar TW cells is regulated by brassinosteroid (BR) and its signaling. PtiCYP85A3, a key BR biosynthesis gene, was predominantly expressed in the xylem of TW, accompanied with a relatively higher castasterone (CS) accumulation, than in the xylem of opposite wood (OW). A wider expression zone of BZR1, a key transcriptional factor in BR singling pathway, was also observed in G-fiber cells on TW side than in wood fiber cells on the OW side, as indicated by immunohistochemistry assays. Transgenic poplar plants overexpressing PtiCYP85A3 produced thicker G-layer with higher cellulose proportion, and accumulated more BZR1 protein in the xylem of TW than did the wild type (WT) plants. Expression of most TW-associated CesAs, which were induced by 2, 4-epibrassinolide, an active BR, and inhibited by brassinazole, a BR biosynthesis inhibitor, were also up-regulated in the xylem of TW in transgenic plants compared to that in WT plants. Further studies with dual-luciferase assays demonstrated that the promoters of PtiCesAs were activated by PtiMYB128, a TW specific transcription factor, which was then regulated by BZR1. All these results indicate that BR plays a crucial role in the G-layer formation of TW fiber cells by regulating the expression of BZR1, PtiMYB128, and PtiCesAs in poplar.

Role of the proteome in providing phenotypic stability in control and ectomycorrhizal poplar plants exposed to chronic mild Pb stress

Lead is a dangerous pollutant that accumulates in plant tissues and causes serious damage to plant cell macromolecules. However, plants have evolved numerous tolerance mechanisms, including ectomycorrhizae, to maintain cellular Pb2+ at the lowest possible level. When those mechanisms are successful, Pb-exposed plants should exhibit no negative phenotypic changes. However, actual molecular-level plant adjustments at Pb concentrations below the toxicity threshold are largely unknown, similar to the molecular effects of protective ectomycorrhizal root colonization. In this study, we (1) determined the molecular adjustments in plants exposed to Pb but without visible Pb stress symptoms and (2) examined ectomycorrhizal root colonization (the role of fungal biofilters) with respect to molecular-level Pb perception by plant root cells. Biochemical, microscopic, proteomic and metabolomic studies were performed to determine the molecular status of Populus × canescens microcuttings grown in agar medium enriched with 0.75 mM Pb(NO3)2. Noninoculated and inoculated with Paxillus involutus poplars were analyzed in two independent comparisons of the corresponding control and Pb treatments. After six weeks of growth, Pb caused no negative phenotypic effects. No Pb-exposed poplar showed impaired growth or decreased leaf pigmentation. Proteomic signals of intensified Pb sequestration in the plant cell wall and vacuoles, cytoskeleton modifications, H+-ATPase-14-3-3 interactions, and stabilization of protein turnover in chronically Pb-exposed plants co-occurred with high metabolomic stability. There were no differentially abundant root primary metabolites; only a few differentially abundant root secondary metabolites and no Pb-triggered ROS burst were observed. Our results strongly suggest that proteome adjustments targeting Pb sequestration and ROS scavenging, which are considerably similar but less intensive in ectomycorrhizal poplars than in control poplars due to the P. involutus biofilter (as confirmed in a mineral study), were responsible for the metabolomic and phenotypic stability of poplars exposed to chronic mild Pb stress.

Energy efficiency of protection of black poplar plantings from weeds

Purpose. Determine the energy efficiency of growing black poplar plantings.Methods. Field, laboratory.Results. It is established that from the energy point of view, the use of mulching allowed to obtain more energy with the harvest than spent on the technology of planting and caring for black poplar plants. The best mulching options were the use of straw layer 5 cm with Kee of 4.78. In addition, with the use of wood sawdust Kee values were at the level of the control weed version of the experiment. Application of the herbicide Stomp 330 EC, 5 l/ha contributed to obtaining Kee at the level of 4.21, and for the application of herbicide Zenkor 70, WG, 1.5 kg/ha, Kee was 4.77, and for the application of herbicide Starga EC, 2.0 respectively 4.24 and with the application of Stefam New, EC, 1.0 l/ha,4.89. The highest energy efficiency ratio was obtained with the combined application of Starga (1.5 l/ha) + Stefam New (0.5 l/ha), 5.27. In black poplar plantations, where in the first year of vegetation three consecutive inter-row cultivations were carried out, a total of 97.9 GJ/ha of energy was formed in three years at an energy efficiency coefficient of 4.77. The use of three hand weeding required additional human labor costs, but in total this had a negligible effect on the energy efficiency ratio of 4.94. However, the use of manual labor on an industrial scale is impossible due to the actual lack of sufficient labor reserves and the need for decent pay for employees, which cannot be justified even by high values of energy efficiency.Conclusions. It was established that in black poplar plantations without the presence of weeds the total energy yield for three years was 107.6 GJ/ha, whereas plantings without weed control provided the formation of only 46.1 GJ/ha, and the energy efficiency of the herbicide protection of black poplar provided the formation of 87.1 to 106.4 GJ/ha.

EARLY BUD BREAK 1 triggers bud break in peach trees by regulating hormone metabolism, the cell cycle, and cell wall modifications.

In a previous study we identified EARLY BUD BREAK 1 (EBB1), an ERF transcription factor, in peach (Prunus persica var. nectarina cultivar Zhongyou 4); however, little is known of how PpEBB1 may regulate bud break. To verify the function of PpEBB1 in bud break, PpEBB1 was transiently transformed into peach buds, resulting in early bud break. Bud break occurred earlier in PpEBB1-oe poplar (Populus trichocarpa) obtained by heterologous transformation than in wild type (WT), consistent with the peach bud results, indicating that PpEBB1 can promote bud break. To explore how PpEBB1 affects bud break, differentially expressed genes (DEGs) between WT and PpEBB1-oe poplar plants were identified by RNA-sequencing. The expression of DEGs associated with hormone metabolism, cell cycle, and cell wall modifications changed substantially according to qRT-PCR. Auxin, ABA, and total trans-zeatin-type cytokinin levels were higher in the PpEBB1-oe plants than in WT plants, while the total N6-(Δ 2-isopentenyl)-adenine-type cytokinins was lower. Yeast two-hybrid and bimolecular fluorescence complementation assays verified that a cell wall modification-related protein (PpEXBL1) interacted with PpEBB1 suggesting that PpEBB1 could interact with these cell wall modification proteins directly. Overall, our study proposed a multifaceted explanation for how PpEBB1 regulates bud break and showed that PpEBB1 promotes bud break by regulating hormone metabolism, the cell cycle, and cell wall modifications.

AUXIN RESPONSE FACTOR 1 Acts as a Positive Regulator in the Response of Poplar to Trichoderma asperellum Inoculation in Overexpressing Plants.

Numerous Trichoderma strains have been reported to be optimal biofertilizers and biocontrol agents with low production costs and environmentally friendly properties. Trichoderma spp. promote the growth and immunity of plants by multiple means. Interfering with the hormonal homeostasis in plants is the most critical strategy. However, the mechanisms underlying plants’ responses to Trichoderma remain to be further elucidated. Auxin is the most important phytohormone that regulates almost every aspect of a plant’s life, especially the trade-off between growth and defense. The AUXIN RESPONSE FACTOR (ARF) family proteins are key players in auxin signaling. We studied the responses and functions of the PdPapARF1 gene in a hybrid poplar during its interaction with beneficial T. asperellum strains using transformed poplar plants with PdPapARF1 overexpression (on transcription level in this study). We report that PdPapARF1 is a positive regulator for promoting poplar growth and defense responses, as does T. asperellum inoculation. PdPapARF1 also turned out to be a positive stimulator of adventitious root formation. Particularly, the overexpression of PdPapARF1 induced a 32.3% increase in the height of 40-day-old poplar plants and a 258% increase in the amount of adventitious root of 3-week-old subcultured plant clones. Overexpressed PdPapARF1 exerted its beneficial functions through modulating the hormone levels of indole acetic acid (IAA), jasmonic acid (JA), and salicylic acid (SA) in plants and activating their signaling pathways, creating similar results as inoculated with T. asperellum. Particularly, in the overexpressing poplar plants, the IAA level increased by approximately twice of the wild-type plants; and the signaling pathways of IAA, JA, and SA were drastically activated than the wild-type plants under pathogen attacks. Our report presents the potential of ARFs as the crucial and positive responders in plants to Trichoderma inducing.

Effect of Microbial Consortium (Bio-Fertilizer) on Growth of Poplar Clones in Nursery

The present study was conducted at Department of Forestry and Natural Resources, PAU, Ludhiana during the year 2015-16. The influence of different doses of microbial consortium (bio-fertilizer) addition to nine different poplar clones in nursery on growth was studied. Different doses of bio-fertilizer showed the significant results for collar diameter, leaf area, and biomass accumulation except plant height and also reveal that different clones showed the significant results for collar diameter, leaf area, plant height, biomass accumulation. Study also revealed that the interaction between different doses of bio-fertilizers and clones showed the significant results for collar diameter, leaf area, and plant height except in plant biomass. The application of microbial consortium (bio-fertilizers) increased the growth parameters in poplar plants. Application of microbial consortium resulted in enhanced growth for most of the characters and can be used as alternative to chemical fertilizer

Brassinosteroid overproduction improves lignocellulose quantity and quality to maximize bioethanol yield under green-like biomass process in transgenic poplar

BackgroundAs a leading biomass feedstock, poplar plants provide enormous lignocellulose resource convertible for biofuels and bio-chemicals. However, lignocellulose recalcitrance particularly in wood plants, basically causes a costly bioethanol production unacceptable for commercial marketing with potential secondary pollution to the environment. Therefore, it becomes important to reduce lignocellulose recalcitrance by genetic modification of plant cell walls, and meanwhile to establish advanced biomass process technology in woody plants. Brassinosteroids, plant-specific steroid hormones, are considered to participate in plant growth and development for biomass production, but little has been reported about brassinosteroids roles in plant cell wall assembly and modification. In this study, we generated transgenic poplar plant that overexpressed DEETIOLATED2 gene for brassinosteroids overproduction. We then detected cell wall feature alteration and examined biomass enzymatic saccharification for bioethanol production under various chemical pretreatments.ResultsCompared with wild type, the PtoDET2 overexpressed transgenic plants contained much higher brassinosteroids levels. The transgenic poplar also exhibited significantly enhanced plant growth rate and biomass yield by increasing xylem development and cell wall polymer deposition. Meanwhile, the transgenic plants showed significantly improved lignocellulose features such as reduced cellulose crystalline index and degree of polymerization values and decreased hemicellulose xylose/arabinose ratio for raised biomass porosity and accessibility, which led to integrated enhancement on biomass enzymatic saccharification and bioethanol yield under various chemical pretreatments. In contrast, the CRISPR/Cas9-generated mutation of PtoDET2 showed significantly lower brassinosteroids level for reduced biomass saccharification and bioethanol yield, compared to the wild type. Notably, the optimal green-like pretreatment could even achieve the highest bioethanol yield by effective lignin extraction in the transgenic plant. Hence, this study proposed a mechanistic model elucidating how brassinosteroid regulates cell wall modification for reduced lignocellulose recalcitrance and increased biomass porosity and accessibility for high bioethanol production.ConclusionsThis study has demonstrated a powerful strategy to enhance cellulosic bioethanol production by regulating brassinosteroid biosynthesis for reducing lignocellulose recalcitrance in the transgenic poplar plants. It has also provided a green-like process for biomass pretreatment and enzymatic saccharification in poplar and beyond.

Overexpression of PtAnnexin1 from Populus trichocarpa enhances salt and drought tolerance in transgenic poplars

Plant annexins are phospholipid- and membrane-binding proteins that are associated with plant development and response to various abiotic stresses. Although previous studies have focused on annexin gene characterization and function from numerous plants, no studies to date have explored annexin1 genes from the poplar, Populus trichocarpa. PtAnnexin1 from P. trichocarpa encodes a putative annexin protein. It is expressed at varying levels in different tissues and is strongly induced by different abiotic stresses. In this study, in vitro enzyme activity assays showed that PtAnnexin1 protein had GTPase/ATPase activity, but no peroxidase activity. Phenotypic analyses demonstrated that transgenic poplar plants overexpressing PtAnnexin1 showed significant growth dominance compared to wild-type (WT) plants in response to drought and salt stresses. In addition, poplars overexpressing PtAnnexin1 had higher total chlorophyll content; increased POD and SOD activities; and higher contents of proline and MDA, all of which contributed to an increase in salt and drought stress tolerance. These findings demonstrate that PtAnnexin1 plays an important role in the abiotic stress response of poplar plants, and that overexpression of PtAnnexin1 in transgenic poplar improves salt and drought tolerance.

ВПЛИВ СОЛЬОВОГО СТРЕСУ НА РОСЛИНИ ТОПОЛІ КЛОНУ 'INRA 353-38' ТА ВЕРБИ КЛОНУ 'ЖИТОМИРСЬКА-1' В УМОВАХ КУЛЬТУРИ IN VITRO

Soil salinization is an important abiotic factor negatively affecting plant growth, development and productivity. Fast-growing poplar and willow trees are important plants for bioenergy production demonstrating varying degrees of adaptation to different habitats. The study of salt resistance in different clones of poplars and willows will reveal genotypes that can be planted in saline soils for producing biomass for the bioenergy industry. Therefore, the aim of the study was to investigate the effects of salt stress on poplar plants of clone 'INRA 353-38' (Populus tremula × P. tremuloides) and willow clone 'Zhytomyrska – 1' (Salix sp.) under in vitro culture. For this purpose the plants were cultivated on MS nutrient medium with the addition of sodium chloride in concentrations 25 mM, 50 mM and 100 mM. The control plants were grown on the sodium chloridefree medium. The plant status (with a 4-score scale), the intensity of their growth (by shoot length) and rooting capacity (by the number of roots) were assessed on the 10th and the 30th day of cultivation. The results obtained indicate a high level of sensitivity to sodium chloride of both studied clones under in vitro cultivation. But the willow 'Zhytomyrska – 1' had a higher sensitivity to salt stress comparing to hybrid polar 'ІNRA 353-38' since growth parameters of willow were significantly decreased even under the concentration of sodium chloride 50 mM, and in the case of short term influence (10 days) of the highest concentration of sodium chloride (100 mM) all willow plants terminated their growth and quickly died. The growth parameters of hybrid poplar were declined within a month, mainly under the highest concentration of sodium chloride, but even under such conditions some part of the shoots were able to survive.

Changes in Proteome and Protein Phosphorylation Reveal the Protective Roles of Exogenous Nitrogen in Alleviating Cadmium Toxicity in Poplar Plants.

Phytoremediation soil polluted by cadmium has drawn worldwide attention. However, how to improve the efficiency of plant remediation of cadmium contaminated soil remains unknown. Previous studies showed that nitrogen (N) significantly enhances cadmium uptake and accumulation in poplar plants. In order to explore the important role of nitrogen in plants’ responses to cadmium stress, this study investigates the poplar proteome and phosphoproteome difference between Cd stress and Cd + N treatment. In total, 6573 proteins were identified, and 5838 of them were quantified. With a fold-change threshold of > 1.3, and a p-value < 0.05, 375 and 108 proteins were up- and down-regulated by Cd stress when compared to the control, respectively. Compared to the Cd stress group, 42 and 89 proteins were up- and down-regulated by Cd + N treatment, respectively. Moreover, 522 and 127 proteins were up- and down-regulated by Cd + N treatment compared to the CK group. In addition, 1471 phosphosites in 721 proteins were identified. Based on a fold-change threshold of > 1.2, and a p-value < 0.05, the Cd stress up-regulated eight proteins containing eight phosphosites, and down-regulated 58 proteins containing 69 phosphosites, whereas N + Cd treatment up-regulated 86 proteins containing 95 phosphosites, and down-regulated 17 proteins containing 17 phosphosites, when compared to Cd stress alone. N + Cd treatment up-regulated 60 proteins containing 74 phosphosites and down-regulated 37 proteins containing 42 phosphosites, when compared to the control. Several putative responses to stress proteins, as well as transcriptional and translational regulation factors, were up-regulated by the addition of exogenous nitrogen following Cd stress. Especially, heat shock protein 70 (HSP70), 14-3-3 protein, peroxidase (POD), zinc finger protein (ZFP), ABC transporter protein, eukaryotic translation initiation factor (elF) and splicing factor 3 B subunit 1-like (SF3BI) were up-regulated by Cd + N treatment at both the proteome and the phosphoproteome levels. Combing the proteomic data and phosphoproteomics data, the mechanism by which exogenous nitrogen can alleviate cadmium toxicity in poplar plants was explained at the molecular level. The results of this study will establish the solid molecular foundation of the phytoremediation method to improve cadmium-contaminated soil.

Variability in flooding tolerance, growth and leaf traits in a Populus deltoides intraspecific progeny.

Climate change will increase the risk of flooding in several areas of the world where Populus deltoides Marshall (eastern cottonwood) is planted, so it would be desirable for this species to select for flooding tolerance. The aims of this work were to explore the variability in growth, leaf traits and flooding tolerance in an F1 full-sib intraspecific progeny of P. deltoides, to analyze the correlations of leaf and growth traits with flooding tolerance and to assess their suitability for use in breeding programs. Two-month-old parental clones and their progeny of 30 full-sib F1 genotypes were grown in pots and subjected to two treatments: (i) plants watered to field capacity (control) and (ii) plants flooded up to 10 cm above soil level for 35days. Growth (height, diameter and biomass partition) and leaf traits (leaf size and number, specific leaf area, leaf senescence, abscission, stomatal conductance, carbon isotope discrimination, stomatal index) were measured. Flooding tolerance for each genotype was estimated as the ratio of the biomass of stressed plants to the biomass of control plants. Results showed segregation in terms of flooding tolerance in the F1 progeny. A significant genotype effect was found for leaf size and number, carbon isotopic discrimination and stomatal conductance, but it did not correlate with flooding tolerance. Height, diameter and root-to-shoot ratio had a positive phenotypic correlation with flooding tolerance, and there was a positive genetic correlation of height and diameter with biomass on both treatments. The narrow sense heritability values for the traits analyzed ranged from 0 to 0.56. We conclude that growth traits are more adequate than leaf traits for selection to increase flooding tolerance. A vigorous initial growth would increase flooding tolerance in young poplar plants.

Overexpression of PtDefensin enhances resistance to Septotis populiperda in transgenic poplar

Plant defensins have been implicated in the plant defense system, but their role in poplar immunity is still unclear. In the present study, we present evidence that PtDefensin, a putative plant defensin, participates in the defense of poplar plants against Septotis populiperda infection. After the construction of recombinant plasmid PET-32a-PtDefensin, PtDefensin protein was expressed in Escherichia coli strain BL21 (DE3) and purified through Ni-IDA resin affinity chromatography. The Trx-PtDefensin fusion protein displayed no cytotoxic activity against RAW264.7 cells but had cytotoxic activity against E. coli K12D31 cells. Analyses of PtDefensin transcript abundance showed that the expression levels of PtDefensin responded to abiotic and biotic stresses. Overexpression of PtDefensin in ‘Nanlin 895’ poplars (Populus × euramericana cv ‘Nanlin895’) increased resistance to Septotis populiperda, coupled with upregulation of MYC2 (basic helix-loop-helix (bHLH) transcription factor) related to jasmonic acid (JA) signal transduction pathways and downregulation of Jasmonate-zim domain (JAZ), an inhibitor in the JA signal transduction pathway. We speculate that systemic acquired resistance (SAR) was activated in non-transgenic poplars after S. populiperda incubation, and that induced systemic resistance (ISR) was activated more obviously in transgenic poplars after S. populiperda incubation. Hence, overexpression of PtDefensin may improve the resistance of poplar plants to pathogens.

Plastid-expressed Bacillus thuringiensis (Bt) cry3Bb confers high mortality to a leaf eating beetle in poplar.

The Bacillus thuringiensis (Bt) cry3Bb gene was successfully introduced into poplar plastid genome, leading to transplastomic poplar with high mortality to Plagiodera versicolora. Poplar (Populus L.) is one of the main resource of woody industry, but being damaged by insect pests. The feasibility and efficiency of plastid transformation technology for controlling two lepidopteran caterpillars have been demonstrated previously. Here, we introduced B. thuringiensis (Bt) cry3Bb into poplar plastid genome by biolistic bombardment for controlling P. versicolora, a widely distributed forest pest. Chimeric cry3Bb gene is controlled by the tobacco plastid rRNA operon promoter combined with the 5'UTR from gene10 of bacteriophage T7 (NtPrrn:T7g10) and the 3'UTR from the E. coli ribosomal RNA operon rrnB (TrrnB). The integration of transgene and homoplasmy of transplastomic poplar plants was confirmed by Southern blot analysis. Northern blot analysis indicated that cry3Bb was transcribed to both read through and shorter length transcripts in plastid. The transplastomic poplar expressing Cry3Bb insecticidal protein showed the highest accumulation level in young leaves, which reach up to 16.8μg/g fresh weight, and comparatively low levels in mature and old leaves. Feeding the young leaves from Bt-Cry3Bb plastid lines to P. versicolora caused 100% mortality in the first-instar larvae after only 1day, in the second-instar larvae after 2days, and in the third-instar larvae for 3days. Thus, we report a successful extension of plastid engineering poplar against the chrysomelid beetle.