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1.
BMC Genomics ; 22(1): 804, 2021 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-34749656

RESUMO

BACKGROUND: Xyloglucan endotransglucosylase/hydrolase (XTH) family plays an important role in cell wall reconstruction and stress resistance in plants. However, the detailed characteristics of XTH family genes and their expression pattern under salt stress have not been reported in poplar. RESULTS: In this study, a total of 43 PtrXTH genes were identified from Populus simonii × Populus nigra, and most of them contain two conserved structures (Glyco_hydro_16 and XET_C domain). The promoters of the PtrXTH genes contain mutiple cis-acting elements related to growth and development and stress responses. Collinearity analysis revealed that the XTH genes from poplar has an evolutionary relationship with other six species, including Eucalyptus robusta, Solanum lycopersicum, Glycine max, Arabidopsis, Zea mays and Oryza sativa. Based on RNA-Seq analysis, the PtrXTH genes have different expression patterns in the roots, stems and leaves, and many of them are highly expressed in the roots. In addition, there are11 differentially expressed PtrXTH genes in the roots, 9 in the stems, and 7 in the leaves under salt stress. In addition, the accuracy of RNA-Seq results was verified by RT-qPCR. CONCLUSION: All the results indicated that XTH family genes may play an important role in tissue specificity and salt stress response. This study will lay a theoretical foundation for further study on molecular function of XTH genes in poplar.


Assuntos
Glicosiltransferases , Hidrolases/genética , Proteínas de Plantas , Populus , Regulação da Expressão Gênica de Plantas , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Populus/enzimologia , Populus/metabolismo , Estresse Salino
2.
BMC Genomics ; 22(1): 731, 2021 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-34625025

RESUMO

BACKGROUND: Trees such as Populus are planted extensively for reforestation and afforestation. However, their successful establishment greatly depends upon ambient environmental conditions and their relative resistance to abiotic and biotic stresses. Polyphenol oxidase (PPO) is a ubiquitous metalloproteinase in plants, which plays crucial roles in mediating plant resistance against biotic and abiotic stresses. Although the whole genome sequence of Populus trichocarpa has long been published, little is known about the PPO genes in Populus, especially those related to drought stress, mechanical damage, and insect feeding. Additionally, there is a paucity of information regarding hormonal responses at the whole genome level. RESULTS: A genome-wide analysis of the poplar PPO family was performed in the present study, and 18 PtrPPO genes were identified. Bioinformatics and qRT-PCR were then used to analyze the gene structure, phylogeny, chromosomal localization, gene replication, cis-elements, and expression patterns of PtrPPOs. Sequence analysis revealed that two-thirds of the PtrPPO genes lacked intronic sequences. Phylogenetic analysis showed that all PPO genes were categorized into 11 groups, and woody plants harbored many PPO genes. Eighteen PtrPPO genes were disproportionally localized on 19 chromosomes, and 3 pairs of segmented replication genes and 4 tandem repeat genomes were detected in poplars. Cis-acting element analysis identified numerous growth and developmental elements, secondary metabolism processes, and stress-related elements in the promoters of different PPO members. Furthermore, PtrPPO genes were expressed preferentially in the tissues and fruits of young plants. In addition, the expression of some PtrPPOs could be significantly induced by polyethylene glycol, abscisic acid, and methyl jasmonate, thereby revealing their potential role in regulating the stress response. Currently, we identified potential upstream TFs of PtrPPOs using bioinformatics. CONCLUSIONS: Comprehensive analysis is helpful for selecting candidate PPO genes for follow-up studies on biological function, and progress in understanding the molecular genetic basis of stress resistance in forest trees might lead to the development of genetic resources.


Assuntos
Catecol Oxidase , Proteínas de Plantas/genética , Populus , Catecol Oxidase/genética , Secas , Regulação da Expressão Gênica de Plantas , Filogenia , Populus/enzimologia , Populus/genética , Estresse Fisiológico
3.
Int J Mol Sci ; 22(18)2021 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-34576057

RESUMO

Stomatal regulation is crucial to reduce water consumption under drought conditions. Extracellular ATP (eATP) serves as a signaling agent in stomatal regulation; however, it is less known whether the eATP mediation of stomatal aperture is linked to apyrases (APYs), the principal enzymes that control the concentration of eATP. To clarify the role of APYs in stomatal control, PeAPY1 and PeAPY2 were isolated from Populus euphratica and transferred into Arabidopsis. Compared with the wild-type Arabidopsis and loss-of-function mutants (Atapy1 and Atapy2), PeAPY1- and PeAPY2-transgenic plants decreased stomatal aperture under mannitol treatment (200 mM, 2 h) and reduced water loss during air exposure (90 min). The role of apyrase in stomatal regulation resulted from its control in eATP-regulated stomatal movements and increased stomatal sensitivity to ABA. The bi-phasic dose-responses to applied nucleotides, i.e., the low ATP (0.3-1.0 mM)-promoted opening and high ATP (>2.0 mM)-promoted closure, were both restricted by P. euphratica apyrases. It is noteworthy that eATP at a low concentration (0.3 mM) counteracted ABA action in the regulation of stomatal aperture, while overexpression of PeAPY1 or PeAPY2 effectively diminished eATP promotion in opening, and consequently enhanced ABA action in closure. We postulate a speculative model of apyrase signaling in eATP- and ABA-regulated stomatal movements under drought.


Assuntos
Apirase/genética , Arabidopsis/genética , Plantas Geneticamente Modificadas/genética , Populus/enzimologia , Arabidopsis/crescimento & desenvolvimento , Secas , Regulação da Expressão Gênica de Plantas/genética , Estômatos de Plantas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Populus/genética , Estresse Fisiológico/genética
4.
New Phytol ; 231(4): 1478-1495, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33713445

RESUMO

Plant cellulose is synthesized by a large plasma membrane-localized cellulose synthase (CesA) complex. However, an overall functional determination of secondary cell wall (SCW) CesAs is still lacking in trees, especially one based on gene knockouts. Here, the Cas9/gRNA-induced knockouts of PtrCesA4, 7A, 7B, 8A and 8B genes were produced in Populus trichocarpa. Based on anatomical, immunohistochemical and wood composition evidence, we gained a comprehensive understanding of five SCW PtrCesAs at the genetic level. Complete loss of PtrCesA4, 7A/B or 8A/B led to similar morphological abnormalities, indicating similar and nonredundant genetic functions. The absence of the gelatinous (G) layer, one-layer-walled fibres and a 90% decrease in cellulose in these mutant woods revealed that the three classes of SCW PtrCesAs are essential for multilayered SCW structure and wood G-fibre. In addition, the mutant primary and secondary phloem fibres lost the n(G + L)- and G-layers and retained the thicker S-layers (L, lignified; S, secondary). Together with polysaccharide immunolocalization data, these findings suggest differences in the role of SCW PtrCesAs-synthesized cellulose in wood and phloem fibre wall structures. Overall, this functional understanding of the SCW PtrCesAs provides further insights into the impact of lacking cellulose biosynthesis on growth, SCW, wood G-fibre and phloem fibre wall structures in the tree.


Assuntos
Parede Celular/enzimologia , Glucosiltransferases/metabolismo , Populus , Sistemas CRISPR-Cas , Celulose/metabolismo , Técnicas de Inativação de Genes , Populus/enzimologia , Populus/genética , RNA Guia , Madeira/metabolismo
5.
Plant Cell Physiol ; 61(12): 2031-2042, 2021 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-32946565

RESUMO

Mitochondrial processes are implicated in plant response to biotic stress caused by viruses, actinomyces, bacteria and pests, but their function in defense against fungal invasion remains unclear. Here, we investigated the role and regulation of mitochondrial alternative oxidase (AOX) in response to black spot disease caused by the hemibiotrophic fungus Marssonina brunnea in poplar. M. brunnea inoculation induced the transcription of the AOX1a gene in the mitochondrial electron transport chain and of jasmonic acid (JA) and ethylene (ET) biosynthetic genes, with the accumulation of these phytohormones in poplar leaf, while inhibiting the transcript amount of the mitochondrial cytochrome c oxidase gene (COX6b) and genes related to salicylic acid (SA). Enhanced AOX reduced poplar susceptibility to M. brunnea with a higher ATP/ADP ratio while the repressed AOX caused the reverse effect. Exogenous JA and 1-aminocyclopropane-1-carboxylic acid (ACC, a biosynthetic precursor of ET) inhibited the transcript amount of COX6b and consequently increased the ratio of AOX pathway to total respiration. Furthermore, the transcription of CYS C1 and CYS D1 genes catalyzing cyanide metabolism was induced, while the cysteine (CYS) substrate levels reduced upon M. brunnea inoculation; exogenous JA and ACC mimicked the effect of M. brunnea infection on cysteine. Exogenous SA enhanced, while JA and ACC reduced, poplar susceptibility to M. brunnea. Moreover, inhibiting AOX completely prohibited JA- and ET-increased tolerance to M. brunnea in poplar. These observations indicate that the JA- and ET-induced mitochondrial AOX pathway triggers defense against M. brunnea in poplar. This effect probably involves cyanide. These findings deepen our understanding of plant-pathogenic fungi interactions.


Assuntos
Ascomicetos , Ciclopentanos/metabolismo , Resistência à Doença/efeitos dos fármacos , Etilenos/metabolismo , Proteínas Mitocondriais/fisiologia , Oxirredutases/fisiologia , Oxilipinas/metabolismo , Doenças das Plantas/imunologia , Reguladores de Crescimento de Plantas/fisiologia , Proteínas de Plantas/fisiologia , Populus/imunologia , Proteínas Mitocondriais/metabolismo , Oxirredutases/metabolismo , Doenças das Plantas/microbiologia , Reguladores de Crescimento de Plantas/metabolismo , Folhas de Planta/metabolismo , Folhas de Planta/microbiologia , Proteínas de Plantas/metabolismo , Populus/enzimologia , Populus/microbiologia
6.
Science ; 369(6507): 1089-1094, 2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32646917

RESUMO

Cellulose is an essential plant cell wall component and represents the most abundant biopolymer on Earth. Supramolecular plant cellulose synthase complexes organize multiple linear glucose polymers into microfibrils as load-bearing wall components. We determined the structure of a poplar cellulose synthase CesA homotrimer that suggests a molecular basis for cellulose microfibril formation. This complex, stabilized by cytosolic plant-conserved regions and helical exchange within the transmembrane segments, forms three channels occupied by nascent cellulose polymers. Secretion steers the polymers toward a common exit point, which could facilitate protofibril formation. CesA's N-terminal domains assemble into a cytosolic stalk that interacts with a microtubule-tethering protein and may thus be involved in CesA localization. Our data suggest how cellulose synthase complexes assemble and provide the molecular basis for plant cell wall engineering.


Assuntos
Glucosiltransferases/química , Complexos Multienzimáticos/química , Proteínas de Plantas/química , Populus/enzimologia , Biocatálise , Domínio Catalítico , Multimerização Proteica
7.
Methods Mol Biol ; 2149: 145-164, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32617934

RESUMO

Wood is an important source of biomass for materials and chemicals, and a target for genetic engineering of its properties for different applications or for research. Wood properties can be altered by using different enzymes acting on cell wall polymers postsynthetically in cell walls. This approach allows for a precise polymer structure modification thanks to the specificity of enzymes used. Such enzymes can originate from all kinds of organisms, or even be modified in a desired way for novel attributes. Here we present a general strategy for expressing a microbial enzyme in aspen and targeting it to cell wall, using an example of fungal glucuronoyl esterase. We describe methods of vector cloning, plant transformation, transgenic line selection and multiplication, testing for the presence of enzymatic activity in different cell compartments, and finally the method of plant transferring from sterile culture to the greenhouse conditions.


Assuntos
Parede Celular/enzimologia , Lignina/metabolismo , Populus/enzimologia , Agrobacterium/metabolismo , DNA Complementar/genética , Fungos/genética , Expressão Gênica , Vetores Genéticos/metabolismo , Proteínas de Plantas/isolamento & purificação , Plantas Geneticamente Modificadas , Populus/genética , Regiões Promotoras Genéticas/genética , Sinais Direcionadores de Proteínas , Transformação Genética , Transgenes , Madeira/genética
8.
BMC Plant Biol ; 20(1): 251, 2020 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-32493269

RESUMO

BACKGROUND: Recombinant carbohydrases genes are used to produce transgenic woody plants with improved phenotypic traits. However, cultivation of such plants in open field is challenging due to a number of problems. Therefore, additional research is needed to alleviate them. RESULTS: Results of successful cultivation of the transgenic aspens (Populus tremula) carrying the recombinant xyloglucanase gene (sp-Xeg) from Penicillium canescens in semi-natural conditions are reported in this paper for the first time. Change of carbohydrate composition of wood was observed in transgenic aspens carrying the sp-Xeg gene. The transformed transgenic line Xeg-2-1b demonstrated accelerated growth and increased content of cellulose in wood of trees growing in both greenhouse and outside in comparison with the control untransformed line Pt. The accelerated growth was observed also in the transgenic line Xeg-1-1c. Thicker cell-wall and longer xylem fiber were also observed in both these transgenic lines. Undescribed earlier considerable reduction in the wood decomposition rate of the transgenic aspen stems was also revealed for the transformed transgenic lines. The decomposition rate was approximately twice as lower for the transgenic line Xeg-2-3b in comparison with the control untransformed line Pt. CONCLUSION: A direct dependence of the phenotypic and biochemical traits on the expression of the recombinant gene sp-Xeg was demonstrated. The higher was the level of the sp-Xeg gene expression, the more pronounced were changes in the phenotypic and biochemical traits. All lines showed phenotypic changes in the leave traits. Our results showed that the plants carrying the recombinant sp-Xeg gene do not demonstrate a decrease in growth parameters in semi-natural conditions. In some transgenic lines, a change in the carbohydrate composition of the wood, an increase in the cell wall thickness, and a decrease in the rate of decomposition of wood were observed.


Assuntos
Glicosídeo Hidrolases/genética , Penicillium/genética , Populus/genética , Carboidratos/análise , Parede Celular/genética , Celulose/análise , Penicillium/enzimologia , Plantas Geneticamente Modificadas/genética , Populus/enzimologia , Populus/crescimento & desenvolvimento , Madeira/análise , Xilema/genética
9.
Plant Sci ; 294: 110459, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32234218

RESUMO

Glutathione reductase (GR; EC 1.6.4.2) is a key NADPH-dependent flavo-protein oxidoreductase which can catalyze the oxidized glutathione (GSSG) to reduced glutathione (GSH) to protect plant cells from oxidative damage induced by Reactive oxygen species (ROS) burst. To investigate the biochemical characteristics and functional divergence of Populus GR family, three GR genes (PtGR1.1/1.2/2) were cloned from Populus trichocarpa and their biochemical characteristics were analyzed in this study. All the three genes were expressed in root, stem, leaf and bud, and the expression of PtGR genes were general upregulated under salicylic acid and alamethicin treatment. PtGR1.1 and PtGR1.2 were localized in cytoplasm, while PtGR2 was in chloroplast. The three PtGR proteins showed different enzymatic activities, apparent kinetic characteristic and thermal stability profiles. However, they have similar bivalent metal ions (Cu2+, Cd2+, Zn2+ and Pb2+) sensitivity and optimum pH profiles. Our study sheds light on a comprehensive information of glutathione reductase family in P. trichocarpa, and proved PtGR genes play critical roles when suffering different stresses.


Assuntos
Glutationa Redutase/metabolismo , Proteínas de Plantas/metabolismo , Populus/enzimologia , Alameticina/metabolismo , Cádmio/metabolismo , Cobre/metabolismo , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Glutationa Redutase/genética , Cinética , Chumbo/metabolismo , Proteínas de Plantas/genética , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Caules de Planta/genética , Caules de Planta/metabolismo , Populus/genética , Populus/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Ácido Salicílico/metabolismo , Zinco/metabolismo
10.
Microbiol Res ; 231: 126371, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31734584

RESUMO

Eleven soil samples were collected from different plantations at the Forestry Model Base, Northeast Forestry University, China (45°43'10″N, 126°37'15″E), and 122 Trichoderma strains (T1-T122) were isolated. Nine Trichoderma species were identified based on morphological and molecular classification methods. The diversity of woody fungi was analyzed based on the type and quantity of Trichoderma spp. in the soil samples isolated from each plantation. Subdominant T. pseudoharzianum T17 (TpsT17) was screened and its biocontrol potential against Fusarium oxysporum CFCC86068 (Fox68) and growth promotion of Populus davidiana × P. alba var. pyramidalis (PdPap) seedlings were investigated. Compared with PdPap + Fox68 treatment, PdPap + TpsT17 + Fox68 treatment had an obvious antagonistic effect on Fox68 based on the status of roots and stomata of the poplar seedlings. In addition, pretreatment with TpsT17 increased catalase activity 14-fold and decreased hydrogen peroxide and malondialdehyde concentrations 2.57- and 7-fold, respectively, in the PdPap + TpsT17 + Fox68 treatment compared with the PdPap + Fox68 treatment. The transcription levels of PR1, JAZ6751, MYC2, MP, and JAR1 in PdPap + TpsT17+Fox68-treated plants were upregulated 5.75-, 5.63-, 14.88-, 8.24-, and 10.45-fold, respectively, at 3 d, while LAX2 exhibited little change in comparison with the level in PdPap + Fox-treated plants. TpsT17 was detected in the roots and stems of PdPap + TpsT17- and PdPap + TpsT17+Fox68-treated PdPap 28 d after inoculation, which demonstrated the endogenous capacity of TpsT17.


Assuntos
Endófitos/isolamento & purificação , Fusarium/crescimento & desenvolvimento , Populus , Trichoderma , Antibiose , Antifúngicos , Agentes de Controle Biológico , Catalase/metabolismo , DNA Espaçador Ribossômico , Endófitos/genética , Endófitos/metabolismo , Agricultura Florestal , Genes Fúngicos , Micoses/microbiologia , Filogenia , Doenças das Plantas/microbiologia , Imunidade Vegetal , Populus/enzimologia , Populus/microbiologia , Plântula/metabolismo , Microbiologia do Solo , Trichoderma/genética , Trichoderma/isolamento & purificação , Trichoderma/metabolismo
11.
Int J Biol Macromol ; 155: 1277-1287, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-31730960

RESUMO

Xyloglucan endotransglycosylase/hydrolase (XTH) belongs to the GH16 subfamily of the glycoside hydrolases of carbohydrate active enzymes and plays an important role in the structure and function of plant cell walls. In this study, 11 members of the XTH gene family were cloned from Populus tomentosa. A bioinformatics analysis revealed that 11 PtoXTHs could be classified into three groups, where PtoXTH27 and PtoXTH34 were most likely to exhibit XTH activity. Biochemical analyses of purified PtoXTHs demonstrated that PtoXTH27 and PtoXTH34 had detectable xyloglucan endotransglucosylase (XET) activity, while the others did not exhibit XET or XEH activity. Moreover, enzymatic assays revealed that the optimum reaction temperature of both PtoXTH27 and PtoXTH34 was 37 °C, while their optimum pH values differed, such that PtoXTH27 was 6.0 and PtoXTH34 was 5.0. Enzyme kinetic parameters indicated that PtoXTH34 had higher affinity for the receptor substrate, XXXG, implying that PtoXTH34 and PtoXTH27 in plants have different substrate structure specificity. Finally, heterologous expression of XTH significantly increased intracellular total sugar content and osmotolerance of yeast cells, indicating that PtoXTH27 and PtoXTH34 are potentially involved in osmotic stress responses. These results clearly demonstrate the enzymatic characteristics and putative role of XTH in osmotic stress responses.


Assuntos
Biologia Computacional , Glicosídeo Hidrolases/metabolismo , Glicosiltransferases/metabolismo , Populus/enzimologia , Parede Celular/enzimologia , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/isolamento & purificação , Glicosiltransferases/genética , Glicosiltransferases/isolamento & purificação , Osmorregulação , Proteínas de Plantas/genética , Proteínas de Plantas/isolamento & purificação , Proteínas de Plantas/metabolismo , Populus/genética , Especificidade por Substrato
12.
Plant J ; 102(1): 99-115, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31736216

RESUMO

The salicinoids are anti-herbivore phenolic glycosides unique to the Salicaceae (Populus and Salix). They consist of a salicyl alcohol glucoside core, which is usually further acylated with benzoic, cinnamic or phenolic acids. While salicinoid structures are well known, their biosynthesis remains enigmatic. Recently, two enzymes from poplar, salicyl alcohol benzoyl transferase and benzyl alcohol benzoyl transferase, were shown to catalyze the production of salicyl benzoate, a predicted potential intermediate in salicinoid biosynthesis. Here, we used transcriptomics and co-expression analysis with these two genes to identify two UDP-glucose-dependent glycosyltransferases (UGT71L1 and UGT78M1) as candidate enzymes in this pathway. Both recombinant enzymes accepted only salicyl benzoate, salicylaldehyde and 2-hydroxycinnamic acid as glucose acceptors. Knocking out the UGT71L1 gene by CRISPR/Cas9 in poplar hairy root cultures led to the complete loss of salicortin, tremulacin and tremuloidin, and a partial reduction of salicin content. This demonstrated that UGT71L1 is required for synthesis of the major salicinoids, and suggested that an additional route can lead to salicin. CRISPR/Cas9 knockouts for UGT78M1 were not successful, and its in vivo role thus remains to be determined. Although it has a similar substrate preference and predicted structure as UGT71L1, it appears not to contribute to the synthesis of salicortin, tremulacin and tremuloidin, at least in roots. The demonstration of UGT71L1 as an enzyme of salicinoid biosynthesis will open up new avenues for the elucidation of this pathway.


Assuntos
Glicosídeos/biossíntese , Glicosiltransferases/metabolismo , Proteínas de Plantas/metabolismo , Populus/enzimologia , Álcoois Benzílicos , Glucosídeos/biossíntese , Glicosiltransferases/química , Glicosiltransferases/genética , Redes e Vias Metabólicas , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Populus/genética , Populus/metabolismo , Estrutura Terciária de Proteína
13.
Sci Rep ; 9(1): 19541, 2019 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-31862975

RESUMO

Plant mitochondria are important energy-producing structure and ROS are generated as byproducts. APX is one enzyme of the AsA-GSH cycle to reduces H2O2 to water. We identified both PtomtAPX and PtosAPX are located in mitochondria of Populus tomentosa Carr. PtomtAPX is specifically targeted to mitochondria, while PtosAPX is dual targeted to both chloroplast and mitochondria. The expression of PtomtAPX in mitochondria was 60-fold that of PtosAPX by ELISA and qPCR analysis. Under high light stress, the expression levels of PtosAPX increased, while that of PtomtAPX only slightly changed. Compared to the WT, the antisense transgenic PtomtAPX cell lines showed slowed growth, smaller cells impaired mitochondria in MS medium under normal growth. RNA-seq results showed 3121 genes significantly altered expression in the antisense cells, and most of them are important for mitochondrial function, particularly in oxidative phosphorylation. Our findings demonstrates a mitochondrial location for one APX isoform, and provide valuable insight into the mechanism which ROS balance is modulated by AsA-GSH cycle in mitochondria.


Assuntos
Ascorbato Peroxidases/metabolismo , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Populus/enzimologia , Populus/metabolismo , Ascorbato Peroxidases/genética , Cloroplastos/metabolismo , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Regulação da Expressão Gênica de Plantas/genética , Immunoblotting , Mitocôndrias/metabolismo , Oxirredução , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Reação em Cadeia da Polimerase , Populus/genética
14.
Nat Plants ; 5(7): 676-680, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31285560

RESUMO

The molecular mechanisms underlying mycorrhizal symbioses, the most ubiquitous and impactful mutualistic plant-microbial interaction in nature, are largely unknown. Through genetic mapping, resequencing and molecular validation, we demonstrate that a G-type lectin receptor-like kinase (lecRLK) mediates the symbiotic interaction between Populus and the ectomycorrhizal fungus Laccaria bicolor. This finding uncovers an important molecular step in the establishment of symbiotic plant-fungal associations and provides a molecular target for engineering beneficial mycorrhizal relationships.


Assuntos
Laccaria/fisiologia , Micorrizas/fisiologia , Proteínas de Plantas/metabolismo , Populus/enzimologia , Populus/microbiologia , Proteínas Quinases/metabolismo , Simbiose , Laccaria/genética , Micorrizas/genética , Proteínas de Plantas/genética , Raízes de Plantas/enzimologia , Raízes de Plantas/genética , Raízes de Plantas/microbiologia , Raízes de Plantas/fisiologia , Populus/genética , Populus/fisiologia , Proteínas Quinases/genética
15.
Molecules ; 24(13)2019 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-31261889

RESUMO

In response to insect herbivory, poplar releases a blend of volatiles that plays important roles in plant defense. Although the volatile bouquet is highly complex and comprises several classes of compounds, it is dominated by mono- and sesquiterpenes. The most common precursors for mono- and sesquiterpenes, geranyl diphosphate (GPP) and (E,E)-farnesyl diphosphate (FPP), respectively, are in general produced by homodimeric or heterodimeric trans-isopentenyl diphosphate synthases (trans-IDSs) that belong to the family of prenyltransferases. To understand the molecular basis of herbivory-induced terpene formation in poplar, we investigated the trans-IDS gene family in the western balsam poplar Populus trichocarpa. Sequence comparisons suggested that this species possesses a single FPP synthase gene (PtFPPS1) and four genes encoding two large subunits (PtGPPS1.LSU and PtGPPS2.LSU) and two small subunits (PtGPPS.SSU1 and PtGPPS.SSU2) of GPP synthases. Transcript accumulation of PtGPPS1.LSU and PtGPPS.SSU1 was significantly upregulated upon leaf herbivory, while the expression of PtFPPS1, PtGPPS2.LSU, and PtGPPS.SSU2 was not influenced by the herbivore treatment. Heterologous expression and biochemical characterization of recombinant PtFPPS1, PtGPPS1.LSU, and PtGPPS2.LSU confirmed their respective IDS activities. Recombinant PtGPPS.SSU1 and PtGPPS.SSU2, however, had no enzymatic activity on their own, but PtGPPS.SSU1 enhanced the GPP synthase activities of PtGPPS1.LSU and PtGPPS2.LSU in vitro. Altogether, our data suggest that PtGPPS1.LSU and PtGPPS2.LSU in combination with PtGPPS.SSU1 may provide the substrate for herbivory-induced monoterpene formation in P. trichocarpa. The sole FPP synthase PtFPPS1 likely produces FPP for both primary and specialized metabolism in this plant species.


Assuntos
Dimetilaliltranstransferase/genética , Insetos/fisiologia , Populus/química , Animais , Dimetilaliltranstransferase/metabolismo , Regulação Enzimológica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Herbivoria , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Populus/enzimologia , Populus/genética , Terpenos/química , Regulação para Cima , Compostos Orgânicos Voláteis/química
16.
J Biotechnol ; 302: 92-100, 2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31233773

RESUMO

4-Coumarate: CoA ligase (4CL) is an important branch point directing metabolites to flavonoid or monolignol pathways in plants. It plays a vital role in the biosynthesis of plant nature products in microbes. Herein, Ptr4CL4, Ptr4CL5 and Ptr4CL7 from Populus trichocarpa were cloned and expressed in Escherichia coli. Two recombinant proteins Ptr4CL4 and Ptr4CL5 showed distinct activities for different substrates. The Ptr4CL4, not previously reported, showed the highest affinity and activity for p-coumaric acid, but a unique substrate self-inhibition was observed at high concentration of p-coumaric acid. Ptr4CL5 was suitable for pathway construction due to no self-substrate inhibition and high initial reaction rate. To explore the potential of Ptr4CL5 in biosynthesis of cinnamyl alcohol, a biosynthesis pathway established with Ptr4CL5, PtrCCR2, endogenous reductases was constructed in E. coli and the titer of cinnamyl alcohol reached 4.8 mM which is higher than other reports. The result indicates that the wood-derived Ptr4CL5 has signification potential in the biosynthesis of cinnamyl alcohol and other monolignol derivatives.


Assuntos
Coenzima A Ligases/metabolismo , Populus/enzimologia , Propanóis/metabolismo , Propionatos/metabolismo , Cinética
17.
BMC Plant Biol ; 19(1): 276, 2019 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-31234799

RESUMO

BACKGROUND: Aspartic protease (AP) is one of four large proteolytic enzyme families that are involved in plant growth and development. Little is known about the AP gene family in tree species, although it has been characterized in Arabidopsis, rice and grape. The AP genes that are involved in tree wood formation remain to be determined. RESULTS: A total of 67 AP genes were identified in Populus trichocarpa (PtAP) and classified into three categories (A, B and C). Chromosome mapping analysis revealed that two-thirds of the PtAP genes were located in genome duplication blocks, indicating the expansion of the AP family by segmental duplications in Populus. The microarray data from the Populus eFP browser demonstrated that PtAP genes had diversified tissue expression patterns. Semi-qRT-PCR analysis further determined that more than 10 PtAPs were highly or preferentially expressed in the developing xylem. When the involvement of the PtAPs in wood formation became the focus, many SCW-related cis-elements were found in the promoters of these PtAPs. Based on PtAPpromoter::GUS techniques, the activities of PtAP66 promoters were observed only in fiber cells, not in the vessels of stems as the xylem and leaf veins developed in the transgenic Populus tree, and strong GUS signals were detected in interfascicular fiber cells, roots, anthers and sepals of PtAP17promoter::GUS transgenic plants. Intensive GUS activities in various secondary tissues implied that PtAP66 and PtAP17 could function in wood formation. In addition, most of the PtAP proteins were predicted to contain N- and (or) O-glycosylation sites, and the integration of PNGase F digestion and western blotting revealed that the PtAP17 and PtAP66 proteins were N-glycosylated in Populus. CONCLUSIONS: Comprehensive characterization of the PtAP genes suggests their functional diversity during Populus growth and development. Our findings provide an overall understanding of the AP gene family in trees and establish a better foundation to further describe the roles of PtAPs in wood formation.


Assuntos
Ácido Aspártico Proteases/genética , Genes de Plantas , Família Multigênica , Proteínas de Plantas/genética , Populus/genética , Madeira/crescimento & desenvolvimento , Parede Celular/genética , Sequência Conservada , Duplicação Gênica , Perfilação da Expressão Gênica , Glicosilação , Filogenia , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas , Populus/enzimologia , Populus/crescimento & desenvolvimento , Regiões Promotoras Genéticas
18.
Genes (Basel) ; 10(5)2019 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-31035411

RESUMO

Asparagine synthetase (AS), a key enzyme in plant nitrogen metabolism, plays an important role in plant nitrogen assimilation and distribution. Asparagine (Asn), the product of asparagine synthetase, is one of the main compounds responsible for organic nitrogen transport and storage in plants. In this study, we performed complementation experiments using an Asn-deficient Escherichia coli strain to demonstrate that three putative asparagine synthetase family members in poplar (Populus simonii × P. nigra) function in Asn synthesis. Quantitative real-time PCR revealed that the three members had high expression levels in different tissues of poplar and were regulated by exogenous nitrogen. PnAS1 and PnAS2 were also affected by diurnal rhythm. Long-term dark treatment resulted in a significant increase in PnAS1 and PnAS3 expression levels. Under long-term light conditions, however, PnAS2 expression decreased significantly in the intermediate region of leaves. Exogenous application of ammonium nitrogen, glutamine, and a glutamine synthetase inhibitor revealed that PnAS3 was more sensitive to exogenous glutamine, while PnAS1 and PnAS2 were more susceptible to exogenous ammonium nitrogen. Our results suggest that the various members of the PnAS gene family have distinct roles in different tissues and are regulated in different ways.


Assuntos
Asparagina/metabolismo , Aspartato-Amônia Ligase/genética , Nitrogênio/metabolismo , Populus/enzimologia , Aspartato-Amônia Ligase/química , Escherichia coli/genética , Regulação da Expressão Gênica de Plantas , Glutamato-Amônia Ligase/química , Glutamato-Amônia Ligase/genética , Glutamina/metabolismo , Populus/genética
19.
Int J Mol Sci ; 20(6)2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30875880

RESUMO

Oxidation of methionine to methionine sulfoxide is a type of posttranslational modification reversed by methionine sulfoxide reductases (Msrs), which present an exceptionally high number of gene copies in plants. The side-form general antioxidant function-specific role of each Msr isoform has not been fully studied. Thirty homologous genes of Msr type A (MsrA) and type B (MsrB) that originate from the genomes of Arabidopsis thaliana, Populus trichocarpa, and Oryza sativa were analyzed in silico. From 109 to 201 transcription factors and responsive elements were predicted for each gene. Among the species, 220 and 190 common transcription factors and responsive elements were detected for the MsrA and MsrB isoforms, respectively. In a comparison of 14 MsrA and 16 MsrB genes, 424 transcription factors and responsive elements were reported in both types of genes, with almost ten times fewer unique elements. The transcription factors mainly comprised plant growth and development regulators, transcription factors important in stress responses with significant overrepresentation of the myeloblastosis viral oncogene homolog (MYB) and no apical meristem, Arabidopsis transcription activation factor and cup-shaped cotyledon (NAC) families and responsive elements sensitive to ethylene, jasmonate, sugar, and prolamine. Gene Ontology term-based functional classification revealed that cellular, metabolic, and developmental process terms and the response to stimulus term dominated in the biological process category. Available experimental transcriptomic and proteomic data, in combination with a set of predictions, gave coherent results validating this research. Thus, new manners Msr gene expression regulation, as well as new putative roles of Msrs, are proposed.


Assuntos
Arabidopsis/enzimologia , Metionina Sulfóxido Redutases/genética , Oryza/enzimologia , Populus/enzimologia , Arabidopsis/genética , Biologia Computacional/métodos , Simulação por Computador , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica de Plantas , Ontologia Genética , Metionina Sulfóxido Redutases/metabolismo , Oryza/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Populus/genética , Proteômica/métodos , Fatores de Transcrição/metabolismo
20.
Ecotoxicol Environ Saf ; 174: 35-42, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-30818258

RESUMO

Pollution with both heavy metal and ground-level ozone (O3) has been steadily increasing, especially in the cities with heavy industry. Little information is known about their combined impacts on urban tree. This study was aimed at characterizing the interactive effects of soil cadmium (Cd) addition and O3 fumigation on visible injury and growth, photosynthesis, oxidative stress, antioxidant enzyme activities, abscisic acid (ABA) content and bioaccumulation of Cd in one-year-old Populus alba 'Berolinensis' saplings by using open top chambers in Shenyang city with developed heavy industry, Northeast China. In this study, poplar saplings were grown in the pots containing soil with different concentrations of Cd (0, 100 and 500 mg kg-1) under ambient air (40 µg L-1) and elevated O3 (120 µg L-1). The results showed that EO and its combination with high Cd (500 mg kg-1) induced significant foliar injury symptoms, decreased root weight (by 41.6%) and total biomass (by 17.4%), inhibited net photosynthetic rate and stomatal conductance, and increased malondialdehyde and ABA contents after 4 weeks of O3 exposure. Elevated O3 exacerbated the accumulation of Cd in leaves and stems of poplar plants grown in the pots with high Cd-polluted soil. Our results also indicated that high Cd pollution in soil increased the susceptibility of plants to O3 and exacerbated the adverse impact of elevated O3 on physiological metabolisms of poplar species, which implied that it was very necessary to take into consideration for O3-tolerance of tree species during phytoremediation of Cd-polluted soil in heavy industrial areas.


Assuntos
Poluentes Atmosféricos/toxicidade , Cádmio/toxicidade , Ozônio/toxicidade , Populus/efeitos dos fármacos , Poluentes do Solo/toxicidade , Ácido Abscísico/metabolismo , Biomassa , Cádmio/metabolismo , Malondialdeído/metabolismo , Estresse Oxidativo , Fotossíntese/efeitos dos fármacos , Populus/enzimologia , Populus/metabolismo
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