Haploid plants from anther cultures of poplar (Populus × beijingensis)

Homozygous genotypes are valuable for genetic and genomic studies in higher plants. However, obtaining homozygous perennial woody plants using conventional breeding techniques is currently a challenge due to a long juvenile period, high heterozygosity, and substantial inbreeding depression. In vitro androgenesis has been used to develop haploid and doubled haploid plants. In the present study, we report the regeneration of haploid lines of poplar (Populus × beijingensis) via anther culture. Anthers at the uninucleate stage were induced to produce callus using three basic media. Two auxins (naphthalene acetic acid [NAA] and 2,4-dichloro-phenoxyacetic acid [2,4-D]), and two cytokinin (kinetin [KT] and 6-benzyladenine [BA]) were tested to explore the influence of plant growth regulators on callus response. H medium (Bourgin and Nitsch 1967) supplemented with 1.0 mg/L NAA and 1.0 mg/L KT induced the highest rate of callus formation. When callus obtained from anthers were subcultured in MS medium containing 1.0 mg/L BA and 0.2 mg/L NAA, followed by transfer to half-strength MS medium supplemented with indole-3-butyric acid (0.2–0.5 mg/L), the formation of regenerated plantlets increased dramatically. Inclusion of gibberellic acid (0.02–0.2 mg/L) in addition to a combination of BA (0.6 mg/L)-NAA (0.2 mg/L) in the culture medium resulted in enhanced frequency of shoot development, as well as greater internode elongation. Ploidy analysis of 580 regenerated plants, using both flow cytometry and chromosome counting, revealed 10.3 % haploid and 1.0 % triploid plantlets. The remaining plantlets were all diploid.     

Production of alkanes (C7–C29) from different part of poplar tree via direct deoxy-liquefaction

Poplar leaves, poplar bark and poplar wood were deoxy-liquefied directly in an air-proof stainless steel reactor at different temperatures. The oils from leaves at 350 °C, from bark at 400 °C and from wood at 450 °C, at which the liquid product yields were the maximum, were analyzed by GC–MS. The oils obtained from three parts of poplar tree were quite different from each other in the relative contents of their compositions. The oil from leaves was rich in hydrocarbons (alkanes: C₇–C₂₉; aromatics) and poor in phenolics, while oil from wood was rich in phenolics and poor in hydrocarbons. The oil from bark was moderate. Relative contents of hydrocarbons in the leaves oil were as high as 60.01% but decreased to 29.71% in bark oil and 11.43% in wood oil. GC analysis of gases and FT-IR, GC–MS and elemental analysis of oils were performed in this study.     

Innate immunity: has poplar made its BED?

The perennial plant model species Populus trichocarpa has received considerable attention in the last 5 yr because of its potential use as a bioenergy crop. The completion of its genome sequence revealed extensive homologies with the herbaceous annual species Arabidopsis thaliana. This review highlights the similarities and differences at the qualitative defence response components level, notably in putative NBS-LRR protein content and downstream defence regulators. With almost a twofold NBS-LRR gene complement compared with A. thaliana, P. trichocarpa also encodes some putative R-proteins with unusual architectures and possible DNA-binding capacity. P. trichocarpa also possesses all the known main components characteristic of TIR-NB-LRR and CC-NB-LRR signalling. However, very little has been done with regard to the components involved in the poplar qualitative response to pathogens. In addition, the relationship between plant-biotroph perception/signalling and the role of salicylic acid, an important defence compound, remains uncertain. This review aims to identify the genomic components present in poplar that could potentially participate in the qualitative response and highlights where efforts should be devoted to obtain a better understanding of the poplar qualitative defence response.     

Limitations for phytoextraction management on metal-polluted soils with poplar short rotation coppice—evidence from a 6-year field trial

Poplar clones were studied for their phytoextraction capacity in the second growth cycle (6-year growth) on a site in the Belgian Campine region, which is contaminated with Cd and Zn via historic atmospheric deposition of nearby zinc smelter activities. The field trial revealed regrowth problems for some clones that could not be predicted in the first growth cycle. Four allometric relations were assessed for their capacity to predict biomass yield in the second growth cycle. A power function based on the shoot diameter best estimates the biomass production of poplar with R² values between 0.94 and 0.98. The woody biomass yield ranged from 2.1 to 4.8 ton woody Dry Mass (DM) ha^?1 y^?1. The primary goal was to reduce soil concentrations of metals caused by phytoextraction. Nevertheless, increased metal concentrations were determined in the topsoil. This increase can partially be explained by the input of metals from deeper soil layers in the top soil through litterfall. The phytoextraction option with poplar short rotation coppice in this setup did not lead to the intended soil remediation in a reasonable time span. Therefore, harvest of the leaf biomass is put forward as a crucial part of the strategy for soil remediation through Cd/Zn phytoextraction.     

Seasonal changes in the efflux of sugars from parenchyma cells into the apoplast in poplar stems (Populus × canademis “robusta”)

Summary The efflux of sugars from parenchyma cells into the apoplast has been studied in the wood of Populus × canadensis robusta in relation to the sugar level of the tissue and the sugar content of the tracheal sap during all physiological stages of the year. In poplar, the spring maximum in sugar content of the tracheal sap is clearly not the result of continuous exosmosis during winter but is reached within a short period in spring. The seasonal patterns of starch and sugar content of the wood and of the sugar content of the tracheal sap are described. The efflux of sugars from tissue sections changes drastically over the year, i.e., from 0.3 to 8.3 g mg^–1DWT day^–1. In general, it is high in fall and winter, and low during late spring and summer. However, high tissue sugar levels proved not always to be correlated with high efflux rates or with high sugar levels in the tracheal sap, indicating that the intracellular compartmentalization of sugars, their passive and catalysed release into, and their re-uptake from the apoplast are all essentially involved in determining the actual sugar content of the sap. Sucrose, which is the dominant sugar in the tracheal sap during winter (pH 7.0–7.5) and in the efflux experiments at pH 7.5, in contrast to the hexoses which prevail in the spring sap (pH around 5.5) and also in the efflux experiments at pH 5.6, is considered to be preferentially released in poplar and to become extraplasmatically hydrolysed. The reasons for tree-specific differences are discussed.     

Chromosome-level genome assembly of a triploid poplar Populus alba ‘Berolinensis’

Many recent studies have provided significant insights into polyploid breeding, but limited research has been carried out on trees. The genomic information needed to understand growth and response to abiotic stress in polyploidy trees is largely unknown, but has become critical due to the threats to forests imposed by climate change. Populus alba ‘Berolinensis,’ also known “Yinzhong poplar,” is a triploid poplar from northeast China. This hybrid triploid poplar is widely used as a landscape ornamental and in urban forestry owing to its adaptation to adverse environments and faster growth than its parental diploid. It is an artificially synthesized male allotriploid hybrid, with three haploid genomes of P. alba ‘Berolinensis’ originating from different poplar species, so it is attractive for studying polyploidy genomic mechanisms in heterosis. In this study, we focused on the allelic genomic interactions in P. alba ‘Berolinensis,’ and generated a high-quality chromosome-level genome assembly consisting of 19 allelic chromosomes. Its three haploid chromosome sets are polymorphic with an average of 25.73 nucleotide polymorphism sites per kilobase. We found that some stress-related genes such as RD22 and LEA7 exhibited sequence differences between different haploid genomes. The genome assembly has been deposited in our polyploid genome online analysis website TreeGenomes ( https://www.treegenomes.com ). These polyploid genome-related resources will provide a critical foundation for the molecular breeding of P. alba ‘Berolinensis’ and help us uncover the allopolyploidization effects of heterosis and abiotic stress resistance and traits of polyploidy species in the future.     

How important is woody tissue photosynthesis in poplar during drought stress?

Key message

Woody tissue photosynthesis might play a key role in maintaining plant carbon economy and hydraulic function under unfavourable conditions such as drought stress.

Abstract

Within trees, a portion of respired CO₂ is assimilated by bark and woody tissue photosynthesis, but its physiological role remains unclear, in particular under unfavour able conditions like drought stress. We hypothesised that woody tissue photosynthesis will contribute to overall tree carbon gain both under sufficient water supply and during drought, and plays a role in maintaining the hydraulic function. We subjected half of the trees to a stem and branch light-exclusion treatment to prevent bark and woody tissue photosynthesis. Then, we measured leaf gas exchange and stem growth in Populus deltoides x nigra ‘Monviso’ trees both under well-watered and dry conditions. We additionally measured cavitation using acoustic emission in detached control and light-excluded branches to illustrate the role of woody tissue photosynthesis in xylem embolism repair. Under well-watered conditions, light exclusion resulted in reduced stem growth relative to control trees by 30 %. In response to drought, stem shrinkage of light-excluded trees was more pronounced as compared to control trees. During drought stress also maximum photosynthesis and transpiration rate tended to decrease more rapidly in light-excluded trees compared to control trees. Leaf fall in light-excluded branches together with the larger number of acoustic emissions in control branches indicates that in the latter more xylem vessels were still hydraulically functional under drought. Therefore, our study highlights that photosynthesis at branch and stem level might be a key factor in the resilience of trees to drought stress by maintaining both the plant carbon economy and hydraulic function.

     

Integrated drought responses of black poplar: how important is phenotypic plasticity?

Climate change is expected to increase drought frequency and intensity which will threaten plant growth and survival. In such fluctuating environments, perennial plants respond with hydraulic and biomass adjustments, resulting in either tolerant or avoidant strategies. Plants' response to stress relies on their phenotypic plasticity. The goal of this study was to explore physiology of young Populus nigra in the context of a time‐limited and progressive water deficit in regard to their growth and stress response strategies. Fourteen French 1‐year‐old black poplar genotypes, geographically contrasted, were subjected to withholding water during 8 days until severe water stress. Water fluxes (i.e. leaf water potentials and stomatal conductance) were analyzed together with growth (i.e. radial and longitudinal branch growth, leaf senescence and leaf production). Phenotypic plasticity was calculated for each trait and response strategies to drought were deciphered for each genotype. Black poplar genotypes permanently were dealing with a continuum of adjusted water fluxes and growth between two extreme strategies, tolerance and avoidance. Branch growth, leaf number and leaf hydraulic potential traits had contrasted plasticities, allowing genotype characterization. The most tolerant genotype to water deficit, which maintained growth, had the lowest global phenotypic plasticity. Conversely, the most sensitive and avoidant genotype ceased growth until the season's end, had the highest plasticity level. All the remaining black poplar genotypes were close to avoidance with average levels of traits plasticity. These results underpinned the role of plasticity in black poplar response to drought and calls for its wider use into research on plants' responses to stress.     

Ozone-induced microscopical changes and quantitative carbohydrate contents of hybrid poplar (Populus × euramericana)

Summary Cuttings of hybrid poplar (Populus × euramericana var. Dorskamp) were exposed to ozone (80 g/m³ from 2100 hours to 0700 hours, 180 g/m³ from 0700 hours to 2100 hours) for 3 months. Ozone reduced the starch content in leaves and stem bark, whereas starch granules accumulated in bundle sheath cells along small leaf veins. At the same time, sucrose and inositol content increased in the leaves. Mesophyll cells in the vicinity of the stomata were injured first, and droplet-like material appeared on their walls. In the sieve plates of fumigated trees, the pores showed a higher degree of narrowing than those of the control treatment. Cell collapse in the leaves was accompanied by water loss and an increase in air space. In the stems, the ozone treatment led to a reduced radial width, particularly in the xylem tissue. These results are discussed in relation to reduced or inhibited phloem loading and ozone-induced drought stress. The plants injured by ozone showed quite distinct patterns of metabolite responses as well as enzyme activities (PEP- and RubP-carboxylase) in the leaves from the top to the bottom. There were also remarkable differences in the reaction of sucrose and inositol between leaves and stem bark. Future research should therefore increasingly follow a whole-plant approach for a better understanding of complex plant reactions.     

Paxillus involutus mycorrhiza attenuate NaCl-stress responses in the salt-sensitive hybrid poplar Populus×canescens

In order to characterise the effect of ectomycorrhiza on Na⁺-responses of the salt-sensitive poplar hybrid Populus × canescens, growth and stress responses of Paxillus involutus (strain MAJ) were tested in liquid cultures in the presence of 20 to 500 mM NaCl, and the effects of mycorrhization on mineral nutrient accumulation and oxidative stress were characterised in mycorrhizal and non-mycorrhizal poplar seedlings exposed to 150 mM NaCl. Paxillus involutus was salt tolerant, showing biomass increases in media containing up to 500 mM NaCl after 4 weeks growth. Mycorrhizal mantle formation on poplar roots was not affected by 150 mM NaCl. Whole plant performance was positively affected by the fungus because total biomass was greater and leaves accumulated less Na⁺ than non-mycorrhizal plants. Energy dispersive X-ray microanalysis using transmission electron microscopy analysis of the influence of mycorrhization on the subcellular localisation of Na⁺ and Cl⁻ in roots showed that the hyphal mantle did not diminish salt accumulation in root cell walls, indicating that mycorrhization did not provide a physical barrier against excess salinity. In the absence of salt stress, mycorrhizal poplar roots contained higher Na⁺ and Cl⁻ concentrations than non-mycorrhizal poplar roots. Paxillus involutus hyphae produced H₂O₂ in the mantle but not in the Hartig net or in pure culture. Salt exposure resulted in H₂O₂ formation in cortical cells of both non-mycorrhizal and mycorrhizal poplar and stimulated peroxidase but not superoxide dismutase activities. This shows that mature ectomycorrhiza was unable to suppress salt-induced oxidative stress. Element analyses suggest that improved performance of mycorrhizal poplar under salt stress may result from diminished xylem loading of Na⁺ and increased supply with K⁺.     

Improve freezing tolerance in transgenic poplar by overexpressing a ω-3 fatty acid desaturase gene

Polyunsaturated fatty acids are a major component of glycerolipids in plant cellular membranes, and play important role in the maintenance of its fluidity. In this study, we isolated a novel ω-3 fatty acid desaturase gene (PtFAD3) from Populus tomentosa Carr. and transformed it into hybrid poplar (Populus alba × P. glandulossa) K84 lines by overexpression and RNAi silencing methods. Quantitative PCR analysis revealed varied expression levels of PtFAD3 among the transgenic lines. Two overexpressing lines (OE lines) and two gene down-regulated RNAi lines (DR lines) were chosen for further analysis. The level of the major trienoic fatty acid species, linolenic acid, increased by an average of 10% in the two OE lines and decreased by an average of 5% in the two DR lines. When young poplar cuttings with three leaves were exposed to a freezing temperature (?4°C) for 3 h without cold acclimation, survival rates of the two independent OE lines (70 and 77%) were far superior to the wild type (36.7%) and the two DR lines (10% for both). Damage caused by freezing was alleviated significantly in the two OE lines compared to wild-type controls. Phenotypic evaluation indicated that the variable responses to freezing were attributable to genotypic variation. Therefore, in the case of poplar, a significant increase in freezing tolerance was achieved on a small scale following the introduction of PtFAD3. This strategy can be used for freezing tolerance breeding in hybrid poplar and other economically important forest trees.     

The Fast-track breeding approach can be improved by heat-induced expression of the FLOWERING LOCUS T genes from poplar (Populus trichocarpa) in apple (Malus × domestica Borkh.)

The Fast-track breeding approach in apple is based on the utilization of the BpMADS4 gene from Betula pendula. However, this approach has several disadvantages which could be solved using other flowering inducing genes and inducible promoters. The FLOWERING LOCUS T genes (PtFT1 and PtFT2) from poplar (Populus trichocarpa) driven by the heat-inducible Gmhsp 17.5-E (HSP)-promoter from soybean (Glycine max) were transferred into apple (Malus × domestica Borkh.) cv. ‘Pinova’ in order to induce flowering. Seven transgenic apple lines were obtained. All transgenic apple lines micrografted onto ‘Golden Delicious’ seedlings used as rootstocks were transferred to the greenhouse. Six out of seven transgenic lines developed flowers after a heat treatment at 42 °C for 1 h daily over a period of 28 days. The transgenic line T836 failed to flower. Flower morphology and pollen vitality of transgenic lines appeared normal. Transgenic plants were successfully used for hybridizations. Pollen from Malus ×robusta 5 applied to flowers of transgenic plants resulted in fruit formation. Heat induced PtFT1, respectively PtFT2 over-expressing rootstocks did not cause flowering in micrografted non-transgenic ‘Pinova’ scions. The mRNA of the PtFT genes was transported from transgenic rootstocks to non-transgenic scions only in one case. As a balance between plant development and flowering is important for the production of early flowering plants usable for a fast-track breeding program the new approach based on heat-induced flowering could be a refinement of the fast breeding program using the possibility of turning-on-turning-off flowering in physiological well developed plants.     

Effect of in vitro storage at 4 °C on survival and proliferation of poplar shoots

Shoot explants of in vitro proliferating cultures of Populus tremula (L.) x Populus tremuloides (L.) were stored for three months at 4°C, in dark or light, in basal culture medium with or without 2-isopentenyladenine (2iP), and in rooting medium with naphthalene acetic acid. They were transferred to cold at different times after subculturing. One hundred percent of shoots survived all tested conditions, in spite of leaf browing and necrosis. After transfer to 24°C for 2 weeks and a normal multiplication cycle, the shoots proliferated at a rate similar to controls or at a higher rate in the case of shoots introduced into the cold 7 or 14 days after subculture and stored in dark on medium containing 2iP.Abbreviations 2iP 2-isopentenyladenine - NAA naphthaleneacetic acid - MS Murashige & Skoog (1962) medium     

Influence of over-expression of the FLOWERING PROMOTING FACTOR 1 gene (FPF1) from Arabidopsis on wood formation in hybrid poplar (Populus tremula L. × P. tremuloides Michx.)

Constitutive expression of the FPF1 gene in hybrid aspen (Populus tremula L. × P. tremuloides Michx.) showed a strong effect on wood formation but no effect on flowering time. Gene expression studies showed that activity of flowering time genes PtFT1, PtCO2, and PtFUL was not increased in FPF1 transgenic plants. However, the SOC1/TM3 class gene PTM5, which has been related to wood formation and flowering time, showed a strong activity in stems of all transgenic lines studied. Wood density was lower in transgenic plants, despite significantly reduced vessel frequency which was overcompensated by thinner fibre cell walls. Chemical screening of the wood by pyrolysis GC/MS showed that FPF1 transgenics have higher fractions of cellulose and glucomannan products as well as lower lignin content. The latter observation was confirmed by UV microspectrophotometry on a cellular level. Topochemical lignin distribution revealed a slower increase of lignin incorporation in the developing xylem of the transgenics when compared with the wild-type plants. In line with the reduced wood density, micromechanical wood properties such as stiffness and ultimate stress were also significantly reduced in all transgenic lines. Thus, we provide evidence that FPF1 class genes may play a regulatory role in both wood formation and flowering in poplar.