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1.
Tree Physiol ; 44(1)2024 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-38123502

RESUMO

Anthocyanins are flavonoid-like substances that play important roles in plants' adaptation to various environmental stresses. In this research, we discovered that cytokinin (CK) alone could effectively induce the anthocyanin biosynthesis in Eucalyptus and many other perennial woody plant species, but not in tobacco and Arabidopsis, suggesting a diverse role of CK in regulating anthocyanin biosynthesis in different species. Transcriptomic and metabolomic strategies were used to further clarify the specific role of CK in regulating anthocyanin biosynthesis in Eucalyptus. The results showed that 801 and 2241 genes were differentially regulated at 6 and 24 h, respectively, after CK treatment. Pathway analysis showed that most of the differentially expressed genes were categorized into pathways related to cellular metabolism or transport of metabolites, including amino acids and sugars. The metabolomic results well supported the transcriptome data, which showed that most of the differentially regulated metabolites were related to the metabolism of sugar, amino acids and flavonoids. Moreover, CK treatment significantly induced the accumulation of sucrose in the CK-treated leaves, while sugar starvation mimicked by either defoliation or shading treatment of the basal leaves significantly reduced the sugar increase of the CK-treated leaves and thus inhibited CK-induced anthocyanin biosynthesis. The results of in vitro experiment also suggested that CK-induced anthocyanin in Eucalyptus was sugar-dependent. Furthermore, we identified an early CK-responsive transcription factor MYB113 in Eucalyptus, the expression of which was significantly upregulated by CK treatment in Eucalyptus, but was inhibited in Arabidopsis. Importantly, the overexpression of EgrMYB113 in the Eucalyptus hairy roots was associated with significant anthocyanin accumulation and upregulation of most of the anthocyanin biosynthetic genes. In conclusion, our study demonstrates a key role of CK in the regulation of anthocyanin biosynthesis in Eucalyptus, providing a molecular basis for further understanding the regulatory mechanism and diversity of hormone-regulated anthocyanin biosynthesis in different plant species.


Assuntos
Arabidopsis , Eucalyptus , Antocianinas/metabolismo , Arabidopsis/genética , Eucalyptus/genética , Eucalyptus/metabolismo , Açúcares/metabolismo , Citocininas/metabolismo , Flavonoides/metabolismo , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo
2.
BMC Plant Biol ; 23(1): 604, 2023 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-38030990

RESUMO

BACKGROUND: The WUSCHEL-related Homeobox (WOX) genes, which encode plant-specific homeobox (HB) transcription factors, play crucial roles in regulating plant growth and development. However, the functions of WOX genes are little known in Eucalyptus, one of the fastest-growing tree resources with considerable widespread cultivation worldwide. RESULTS: A total of nine WOX genes named EgWOX1-EgWOX9 were retrieved and designated from Eucalyptus grandis. From the three divided clades marked as Modern/WUS, Intermediate and Ancient, the largest group Modern/WUS (6 EgWOXs) contains a specific domain with 8 amino acids: TLQLFPLR. The collinearity, cis-regulatory elements, protein-protein interaction network and gene expression analysis reveal that the WUS proteins in E. grandis involve in regulating meristems development and regeneration. Furthermore, by externally adding of truncated peptides isolated from WUS specific domain, the transformation efficiency in E. urophylla × E. grandis DH32-29 was significant enhanced. The transcriptomics data further reveals that the use of small peptides activates metabolism pathways such as starch and sucrose metabolism, phenylpropanoid biosynthesis and flavonoid biosynthesis. CONCLUSIONS: Peptides isolated from WUS protein can be utilized to enhance the transformation efficiency in Eucalyptus, thereby contributing to the high-efficiency breeding of Eucalyptus.


Assuntos
Eucalyptus , Genes Homeobox , Eucalyptus/genética , Eucalyptus/metabolismo , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Melhoramento Vegetal , Peptídeos/genética , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Filogenia
3.
Plant Biol (Stuttg) ; 25(4): 541-550, 2023 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-36971569

RESUMO

MicroRNAs (miRNAs) play a crucial role in the growth, development, morphogenesis, signal transduction, and stress response in plants. The ICE (Inducer of CBF expression)-CBF (C-repeat binding factor)-COR (Cold-regulated gene) regulatory cascade is an important signalling pathway in plant response to low temperature stress, and it remains unknown whether this pathway is regulated by miRNAs. In this study, high-throughput sequencing was employed for predicting and identifying the miRNAs that were likely to target the ICE-CBF-COR pathway in Eucalyptus camaldulensis. A novel ICE1-targeting miRNA, eca-novel-miR-259-5p (nov-miR259), was further analysed. A total of 392 conserved miRNAs and 97 novel miRNAs were predicted, including 80 differentially expressed miRNAs. Of these, 30 miRNAs were predicted to be associated with the ICE-CBF-COR pathway. The full-length of mature nov-miR259 was 22 bp and its precursor gene was 60 bp in length, with a typical hairpin structure. The RNA ligase-mediated 5' amplification of cDNA ends (5'-RLM-RACE) and Agrobacterium-mediated tobacco transient expression assays demonstrated that nov-miR259 could cleave EcaICE1 in vivo. Moreover, qRT-PCR and Pearson's correlation analysis further revealed that the expression levels of nov-miR259 were almost significantly negatively correlated with those of its target gene, EcaICE1, and the other genes in the ICE-CBF-COR pathway. We first identified the nov-miR259 as a novel ICE1-targeting miRNA, and the nov-miR259-ICE1 module may be involved in regulating the cold stress response in E. camaldulensis.


Assuntos
Eucalyptus , MicroRNAs , MicroRNAs/genética , MicroRNAs/metabolismo , Eucalyptus/genética , Eucalyptus/metabolismo , Temperatura , Temperatura Baixa , Plantas/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Regulação da Expressão Gênica de Plantas
4.
Genes (Basel) ; 13(10)2022 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-36292698

RESUMO

The transcriptional properties of artificial promoters are closely related to the type and arrangement position of cis-elements. GWSF (374-bp) was an effective SPIP with four cis-element dimers. There were four pathogen-inducible cis-elements in the GWSF promoter (GST1-boxes, W-boxes, S-boxes, and F-boxes) and a minimal cauliflower mosaic virus 35S promoter. V-element dimers were inserted into the upstream (VGWSF), midstream (GWVSF), and downstream (GWSFV) regions of the original GWSF promoter sequence to examine their affect on the position. The expression activity of promoters was analyzed and estimated using the histochemical staining of leaf discs of eucalyptus with transient expression, an image digitization method to extract the color features, and the induction treatment by a plant pathogenic microorganism/inducer and qPCR assays. The histochemical staining results of the adventitious buds indicated that the promoters had been successfully integrated into the E. urophylla genome and that they drove the expression of the gus gene. There was a noticeable difference in the intensity of color between the adventitious buds on the same callus block, as well as the intensity of color within the same adventitious bud. According to the established two-factor model of blue value, there was a greater difference between the levels of the genotype factor than the promoter factor in eucalyptus leaf discs. Further, the basal and inducible transcriptional levels of the three improved promoters were investigated by qPCR. With the basal transcriptional level of the GWSF promoter normalized to one, the relative basal levels of VGWSF, GWVSF, and GWSFV were 1.40, 1.45, and 4.15, respectively. The qPCR results were consistent with the staining results of GUS histochemical staining. The three improved promoters all had the properties of being induced by salicylic acid, Ralstonia solanacearum, and Phytophthora capsici. The three improved promoters demonstrated a significantly higher TMV induction activity: their induction activity from high to low was GWSFV > GWVSF > VGWSF. The findings will be beneficial to the construction and optimization of artificial promoters for transgenic plants.


Assuntos
Resistência à Doença , Eucalyptus , Resistência à Doença/genética , Eucalyptus/genética , Nicotiana/genética , Regiões Promotoras Genéticas , Ácido Salicílico/farmacologia
5.
Ecotoxicol Environ Saf ; 241: 113748, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-35696965

RESUMO

Forest ecosystems play an important role in environmental protection and maintaining ecological balance. Understanding the physiological mechanisms of tree species response to aluminum (Al) toxic is crucial to reveal the main causes of plantation decline in acid rain area. As an important afforestation tree species in tropical and subtropical areas, Eucalyptus has high economic value and plays crucial ecological roles. However, continuous fertilization and acid precipitation can exacerbate soil acidification and increase soil active Al, which has a significant negative impact on Eucalyptus growth. Hence, species and genotypes with high Al resistance are required to solve the problem of Al toxicity of acidic soils for sustainable forest production. In this study, E. urophylla was better adapted to Al stress than E. grandis or E. tereticornis; its high Al resistance was attributed to greater antioxidant enzyme activity and non-enzymatic antioxidant content, and a lower degree of membrane lipid peroxidation than E. grandis or E. tereticornis. The differences in adaptability among the three pure species were attributed to their distinct habitats. Eucalyptus urophylla × E. grandis inherited the outstanding adaptability to Al stress from its maternal species (E. urophylla), indicating that Al tolerance is highly heritable and can be selected in Eucalyptus breeding. Our results indicated that the response of Eucalyptus to Al stress may fluctuate according to the time under stress, and might be related to dynamic changes in ROS elimination and accumulation.


Assuntos
Eucalyptus , Alumínio/toxicidade , Antioxidantes , Ecossistema , Eucalyptus/genética , Estresse Oxidativo , Melhoramento Vegetal , Solo , Árvores
6.
Tree Physiol ; 42(10): 2020-2039, 2022 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-35512354

RESUMO

Eucalypts engage in a mutualistic endosymbiosis with arbuscular mycorrhizal (AM) fungi to acquire mineral nutrients from soils, particularly inorganic phosphate (Pi). In return, the host plant provides organic carbons to its fungal partners. However, the mechanism by which the Eucalyptus plants acquire Pi released from the AM fungi has remained elusive. In this study, we investigated the characterization of potential PHOSPHATE TRANSPORTER1 (PHT1) family Pi transporters in AM symbiosis in Eucalyptus grandis W. Hill ex Maiden. We show that multiple PHT1 family Pi transporters were recruited for AM symbiosis in E. grandis. We further report that EgPT4, an E. grandis member of the PHT1 family, is conserved across angiosperms and is exclusively expressed in AM roots with arbuscule-containing cells and localizes to the periarbuscular membrane (PAM). EgPT4 was able to complement a yeast mutant strain defective in all inorganic Pi transporters and mediate Pi uptake. Importantly, EgPT4 is essential for improved E. grandis growth, total phosphorus concentration and arbuscule development during symbiosis. Moreover, silencing of EgPT4 led to the induction of polyphosphate accumulation relevant genes of Rhizophagus irregularis DAOM 197198. Collectively, our results unravel a pivotal role for EgPT4 in symbiotic Pi transport across the PAM required for arbuscule development in E. grandis.


Assuntos
Eucalyptus , Micorrizas , Eucalyptus/genética , Eucalyptus/metabolismo , Regulação da Expressão Gênica de Plantas , Minerais , Proteínas de Transporte de Fosfato/genética , Fósforo/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , Polifosfatos , Solo , Simbiose/genética
7.
Sci Rep ; 11(1): 24408, 2021 12 23.
Artigo em Inglês | MEDLINE | ID: mdl-34949763

RESUMO

Some forest trees have been polyploidized to improve their traits and to supply new germplasms for breeding programs. As trees have a long juvenile stage, the early characterization of the chromosome set doubling effects is crucial for previous selection. Thus, we aimed to characterize the chemical variability of essential oils from diploid and autotetraploid germplasms (autotetraploid A and B) of Eucalyptus benthamii, as well as to evaluate their larvicidal and allelopathic effects. Autotetraploid A showed a higher essential oil yield than diploid and autotetraploid B, which did not differ quantitatively. Aromadendrene, viridiflorol and α-pinene were the major compounds in the diploid essential oil. In contrast, compounds were present in autotetraploids, such as 1,8-cineole, limonene, α-terpineol, and α-terpinyl-acetate. Essential oils from the diploid at 50-200 ppm were twice as larvicidal than those from autotetraploids against Aedes aegypti larvae. Considering the phytotoxicity bioassays using Lactuca sativa, essential oils from both ploidy levels affected root growth. Moreover, the essential oils inhibited shoot growth at all concentrations tested (187.5; 375; 750; 1500; and 3000 ppm). Autotetraploid A and B had the same effect on shoot growth as glyphosate. The essential oils had no cytogenotoxic effect on root meristematic cells of L. sativa, whereas phytotoxic potential was identified mainly in shoot growth. This work demonstrated a dramatic change in secondary metabolism (terpene composition) related to an increase in the ploidy level in Eucalyptus germplasms. In addition, we report the novelty of the chemical composition of essential oils among germplasms and their potential use as larvicidal and post-emergence weed control agents.


Assuntos
Óleo de Eucalipto/química , Óleo de Eucalipto/farmacologia , Eucalyptus/química , Eucalyptus/genética , Herbicidas , Inseticidas , Óleos Voláteis/química , Óleos Voláteis/farmacologia , Tetraploidia , Aedes/efeitos dos fármacos , Alelopatia/efeitos dos fármacos , Animais , Bioensaio , Relação Dose-Resposta a Droga , Larva/efeitos dos fármacos , Lactuca/efeitos dos fármacos , Lactuca/crescimento & desenvolvimento , Melhoramento Vegetal , Brotos de Planta/efeitos dos fármacos , Brotos de Planta/crescimento & desenvolvimento
8.
Planta ; 253(1): 3, 2020 Dec 21.
Artigo em Inglês | MEDLINE | ID: mdl-33346890

RESUMO

MAIN CONCLUSION: Eucalyptus camaldulensis EcDQD/SDH2 and 3 combine gallate formation, dehydroquinate dehydratase, and shikimate dehydrogenase activities. They are candidates for providing the essential gallate for the biosynthesis of the aluminum-detoxifying metabolite oenothein B. The tree species Eucalyptus camaldulensis shows exceptionally high tolerance against aluminum, a widespread toxic metal in acidic soils. In the roots of E. camaldulensis, aluminum is detoxified via the complexation with oenothein B, a hydrolyzable tannin. In our approach to elucidate the biosynthesis of oenothein B, we here report on the identification of E. camaldulensis enzymes that catalyze the formation of gallate, which is the phenolic constituent of hydrolyzable tannins. By systematical screening of E. camaldulensis dehydroquinate dehydratase/shikimate dehydrogenases (EcDQD/SDHs), we found two enzymes, EcDQD/SDH2 and 3, catalyzing the NADP+-dependent oxidation of 3-dehydroshikimate to produce gallate. Based on extensive in vitro assays using recombinant EcDQD/SDH2 and 3 enzymes, we present for the first time a detailed characterization of the enzymatic gallate formation activity, including the cofactor preferences, pH optima, and kinetic constants. Sequence analyses and structure modeling suggest the gallate formation activity of EcDQD/SDHs is based on the reorientation of 3-dehydroshikimate in the catalytic center, which facilitates the proton abstraction from the C5 position. Additionally, EcDQD/SDH2 and 3 maintain DQD and SDH activities, resulting in a 3-dehydroshikimate supply for gallate formation. In E. camaldulensis, EcDQD/SDH2 and 3 are co-expressed with UGT84A25a/b and UGT84A26a/b involved in hydrolyzable tannin biosynthesis. We further identified EcDQD/SDH1 as a "classical" bifunctional plant shikimate pathway enzyme and EcDQD/SDH4a/b as functional quinate dehydrogenases of the NAD+/NADH-dependent clade. Our data indicate that in E. camaldulensis the enzymes EcDQD/SDH2 and 3 provide the essential gallate for the biosynthesis of the aluminum-detoxifying metabolite oenothein B.


Assuntos
Oxirredutases do Álcool , Eucalyptus , Ácido Gálico , Oxirredutases do Álcool/metabolismo , Alumínio/toxicidade , Vias Biossintéticas/fisiologia , Eucalyptus/efeitos dos fármacos , Eucalyptus/enzimologia , Eucalyptus/genética , Ácido Gálico/metabolismo , Hidroliases/metabolismo
9.
Planta ; 252(3): 45, 2020 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-32880001

RESUMO

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


Assuntos
Eucalyptus/crescimento & desenvolvimento , Eucalyptus/genética , Proteínas de Plantas/genética , Brotos de Planta/crescimento & desenvolvimento , Xilema/fisiologia , Parede Celular/genética , Celulose/metabolismo , Eucalyptus/citologia , Regulação da Expressão Gênica de Plantas , Ácidos Indolacéticos/metabolismo , Lignina/metabolismo , Filogenia , Proteínas de Plantas/metabolismo , Brotos de Planta/genética , Plantas Geneticamente Modificadas/genética , Regiões Promotoras Genéticas , Nicotiana/genética
10.
Int J Mol Sci ; 21(8)2020 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-32344718

RESUMO

Eucalypts are the most planted trees worldwide, but most of them are frost sensitive. Overexpressing transcription factors for CRT-repeat binding factors (CBFs) in transgenic Eucalyptus confer cold resistance both in leaves and stems. While wood plays crucial roles in trees and is affected by environmental cues, its potential role in adaptation to cold stress has been neglected. Here, we addressed this question by investigating the changes occurring in wood in response to the overexpression of two CBFs, taking advantage of available transgenic Eucalyptus lines. We performed histological, biochemical, and transcriptomic analyses on xylem samples. CBF ectopic expression led to a reduction of both primary and secondary growth, and triggered changes in xylem architecture with smaller and more frequent vessels and fibers exhibiting reduced lumens. In addition, lignin content and syringyl/guaiacyl (S/G) ratio increased. Consistently, many genes of the phenylpropanoid and lignin branch pathway were upregulated. Most of the features of xylem remodeling induced by CBF overexpression are reminiscent of those observed after long exposure of Eucalyptus trees to chilling temperatures. Altogether, these results suggest that CBF plays a central role in the cross-talk between response to cold and wood formation and that the remodeling of wood is part of the adaptive strategies to face cold stress.


Assuntos
Resposta ao Choque Frio , Fatores de Ligação ao Core/genética , Eucalyptus/genética , Expressão Gênica , Fatores de Transcrição/genética , Madeira/anatomia & histologia , Madeira/genética , Fatores de Ligação ao Core/metabolismo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Lignina/metabolismo , Fenótipo , Plantas Geneticamente Modificadas , Fatores de Transcrição/metabolismo , Madeira/química , Xilema/genética , Xilema/metabolismo
11.
New Phytol ; 223(4): 1952-1972, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31144333

RESUMO

Acetyl- and methylglucuronic acid decorations of xylan, the dominant hemicellulose in secondary cell walls (SCWs) of woody dicots, affect its interaction with cellulose and lignin to determine SCW structure and extractability. Genes and pathways involved in these modifications may be targets for genetic engineering; however, little is known about the regulation of xylan modifications in woody plants. To address this, we assessed genetic and gene expression variation associated with xylan modification in developing xylem of Eucalyptus grandis × Eucalyptus urophylla interspecific hybrids. Expression quantitative trait locus (eQTL) mapping identified potential regulatory polymorphisms affecting gene expression modules associated with xylan modification. We identified 14 putative xylan modification genes that are members of five expression modules sharing seven trans-eQTL hotspots. The xylan modification genes are prevalent in two expression modules. The first comprises nucleotide sugar interconversion pathways supplying the essential precursors for cellulose and xylan biosynthesis. The second contains genes responsible for phenylalanine biosynthesis and S-adenosylmethionine biosynthesis required for glucuronic acid and monolignol methylation. Co-expression and co-regulation analyses also identified four metabolic sources of acetyl coenxyme A that appear to be transcriptionally coordinated with xylan modification. Our systems genetics analysis may provide new avenues for metabolic engineering to alter wood SCW biology for enhanced biomass processability.


Assuntos
Eucalyptus/genética , Redes e Vias Metabólicas , Análise de Sistemas , Madeira/crescimento & desenvolvimento , Xilanos/metabolismo , Acetilcoenzima A/metabolismo , Vias Biossintéticas/genética , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Redes e Vias Metabólicas/genética , Modelos Biológicos , Modelos Genéticos , Anotação de Sequência Molecular , Locos de Características Quantitativas/genética , Transcrição Gênica , Madeira/genética
12.
New Phytol ; 223(3): 1489-1504, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31066055

RESUMO

Terpenoid-based essential oils are economically important commodities, yet beyond their biosynthetic pathways, little is known about the genetic architecture of terpene oil yield from plants. Transport, storage, evaporative loss, transcriptional regulation and precursor competition may be important contributors to this complex trait. Here, we associate 2.39 million single nucleotide polymorphisms derived from shallow whole-genome sequencing of 468 Eucalyptus polybractea individuals with 12 traits related to the overall terpene yield, eight direct measures of terpene concentration and four biomass-related traits. Our results show that in addition to terpene biosynthesis, development of secretory cavities, where terpenes are both synthesized and stored, and transport of terpenes were important components of terpene yield. For sesquiterpene concentrations, the availability of precursors in the cytosol was important. Candidate terpene synthase genes for the production of 1,8-cineole and α-pinene, and ß-pinene (which comprised > 80% of the total terpenes) were functionally characterized as a 1,8-cineole synthase and a ß/α-pinene synthase. Our results provide novel insights into the genomic architecture of terpene yield and we provide candidate genes for breeding or engineering of crops for biofuels or the production of industrially valuable terpenes.


Assuntos
Eucalyptus/genética , Genoma de Planta , Estudo de Associação Genômica Ampla , Óleos de Plantas/metabolismo , Terpenos/metabolismo , Alquil e Aril Transferases/genética , Vias Biossintéticas , Genes de Plantas , Genótipo , Padrões de Herança/genética , Análise Multivariada , Polimorfismo de Nucleotídeo Único/genética , Locos de Características Quantitativas/genética , Reprodutibilidade dos Testes , Terpenos/química
13.
Plant Physiol ; 180(1): 124-152, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30760638

RESUMO

Isoprene synthase converts dimethylallyl diphosphate to isoprene and appears to be necessary and sufficient to allow plants to emit isoprene at significant rates. Isoprene can protect plants from abiotic stress but is not produced naturally by all plants; for example, Arabidopsis (Arabidopsis thaliana) and tobacco (Nicotiana tabacum) do not produce isoprene. It is typically present at very low concentrations, suggesting a role as a signaling molecule; however, its exact physiological role and mechanism of action are not fully understood. We transformed Arabidopsis with a Eucalyptus globulus isoprene synthase The regulatory mechanisms of photosynthesis and isoprene emission were similar to those of native emitters, indicating that regulation of isoprene emission is not specific to isoprene-emitting species. Leaf chlorophyll and carotenoid contents were enhanced by isoprene, which also had a marked positive effect on hypocotyl, cotyledon, leaf, and inflorescence growth in Arabidopsis. By contrast, leaf and stem growth was reduced in tobacco engineered to emit isoprene. Expression of genes belonging to signaling networks or associated with specific growth regulators (e.g. gibberellic acid that promotes growth and jasmonic acid that promotes defense) and genes that lead to stress tolerance was altered by isoprene emission. Isoprene likely executes its effects on growth and stress tolerance through direct regulation of gene expression. Enhancement of jasmonic acid-mediated defense signaling by isoprene may trigger a growth-defense tradeoff leading to variations in the growth response. Our data support a role for isoprene as a signaling molecule.


Assuntos
Alquil e Aril Transferases/genética , Arabidopsis/genética , Hemiterpenos/fisiologia , Nicotiana/genética , Estresse Fisiológico , Arabidopsis/efeitos dos fármacos , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Butadienos/farmacologia , Carotenoides/metabolismo , Clorofila/metabolismo , Eucalyptus/genética , Regulação da Expressão Gênica de Plantas , Hemiterpenos/biossíntese , Hemiterpenos/farmacologia , Fotossíntese , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Transdução de Sinais , Nicotiana/crescimento & desenvolvimento , Nicotiana/metabolismo , Transformação Genética
14.
Genome ; 61(11): 777-786, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30354691

RESUMO

SWEET proteins are essential for the maintenance of nectar production, as well as seed and pollen development, in plants. A search within the Eucalyptus genome identified 52 putative genes belonging to the SWEET gene family based on sequence similarity. The expression of two of these genes, EcSWEET2 and EcSWEET5, was analyzed in vegetative and reproductive tissues of Eucalyptus camaldulensis. The expression of EcSWEET5 was specific to male reproductive tissues, and transcripts were detected only at certain stages of flower development. Tobacco Rattle Virus (TRV)-mediated suppression of EcSWEET5 resulted in a significant reduction in pollen germination percentage in Nicotiana benthamiana without adverse effect on vegetative growth. A promoter sequence 1 kb upstream of the start codon of EcSWEET5 contained many elements suggestive of pollen specificity of the promoter. This specificity was confirmed in transgenic tobacco lines harboring a GUS gene whose expression was controlled by the EcSWEET5 gene promoter. GUS expression was limited to pollen alone in transgenic tobacco as evidenced by histochemical staining. The expression of a cytotoxic gene, barnase under the control of the EcSWEET5 gene promoter, showed pollen ablation in transgenic tobacco with normal vegetative growth.


Assuntos
Eucalyptus/genética , Genes de Plantas , Regiões Promotoras Genéticas , Proteínas de Transporte de Monossacarídeos/genética , Família Multigênica , Plantas Geneticamente Modificadas , Pólen/genética , Reprodução/genética , Nicotiana/genética
15.
PLoS One ; 13(10): e0206085, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30335843

RESUMO

Modern genotyping techniques, such as SNP analysis and genotyping by sequencing (GBS), are hampered by poor DNA quality and purity, particularly in challenging plant species, rich in secondary metabolites. We therefore investigated the utility of a pre-wash step using a buffered sorbitol solution, prior to DNA extraction using a high salt CTAB extraction protocol, in a high throughput or miniprep setting. This pre-wash appears to remove interfering metabolites, such as polyphenols and polysaccharides, from tissue macerates. We also investigated the adaptability of the sorbitol pre-wash for RNA extraction using a lithium chloride-based protocol. The method was successfully applied to a variety of tissues, including leaf, cambium and fruit of diverse plant species including annual crops, forest and fruit trees, herbarium leaf material and lyophilized fungal mycelium. We consistently obtained good yields of high purity DNA or RNA in all species tested. The protocol has been validated for thousands of DNA samples by generating high data quality in dense SNP arrays. DNA extracted from Eucalyptus spp. leaf and cambium as well as mycelium from Trichoderma spp. was readily digested with restriction enzymes and performed consistently in AFLP assays. Scaled-up DNA extractions were also suitable for long read sequencing. Successful RNA quality control and good RNA-Seq data for Eucalyptus and cashew confirms the effectiveness of the sorbitol buffer pre-wash for high quality RNA extraction.


Assuntos
DNA/normas , Eucalyptus/genética , Polimorfismo de Nucleotídeo Único , RNA/normas , Trichoderma/genética , Soluções Tampão , Câmbio/genética , DNA/isolamento & purificação , DNA Fúngico/isolamento & purificação , DNA Fúngico/normas , DNA de Plantas/isolamento & purificação , DNA de Plantas/normas , Técnicas de Genotipagem , Micélio/genética , Folhas de Planta/genética , RNA/isolamento & purificação , RNA Fúngico/normas , RNA de Plantas/isolamento & purificação , RNA de Plantas/normas , Análise de Sequência de DNA , Análise de Sequência de RNA , Sorbitol/química
16.
PLoS One ; 13(1): e0190900, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29324770

RESUMO

Under acidic conditions, aluminum (Al) toxicity is an important factor limiting plant productivity; however, the application of phosphorus (P) might alleviate the toxic effects of Al. In this study, seedlings of two vegetatively propagated Eucalyptus clones, E. grandis × E. urophylla 'G9' and E. grandis × E. urophylla 'DH32-29'were subjected to six treatments (two levels of Al stress and three levels of P). Under excessive Al stress, root Al content was higher, whereas shoot and leaf Al contents were lower with P application than those without P application. Further, Al accumulation was higher in the roots, but lower in the shoots and leaves of G9 than in those of DH32-29. The secretion of organic acids was higher under Al stress than under no Al stress. Further, under Al stress, the roots of G9 secreted more organic acids than those of DH32-29. With an increase in P supply, Al-induced secretion of organic acids from roots decreased. Under Al stress, some enzymes, including PEPC, CS, and IDH, played important roles in organic acid biosynthesis and degradation. Thus, our results indicate that P can reduce Al toxicity via the fixation of elemental Al in roots and restriction of its transport to stems and leaves, although P application cannot promote the secretion of organic acid anions. Further, the higher Al-resistance of G9 might be attributed to the higher Al accumulation in and organic acid anion secretion from roots and the lower levels of Al in leaves.


Assuntos
Alumínio/toxicidade , Eucalyptus/efeitos dos fármacos , Eucalyptus/metabolismo , Fósforo/farmacologia , Folhas de Planta/metabolismo , Raízes de Plantas/metabolismo , Substâncias Protetoras/farmacologia , Cloreto de Alumínio , Compostos de Alumínio/farmacologia , Biomassa , Cloretos/farmacologia , Enzimas/metabolismo , Eucalyptus/genética , Fosfatos/administração & dosagem , Fosfatos/farmacologia , Fósforo/administração & dosagem , Folhas de Planta/efeitos dos fármacos , Proteínas de Plantas/metabolismo , Raízes de Plantas/efeitos dos fármacos , Caules de Planta/efeitos dos fármacos , Caules de Planta/metabolismo , Compostos de Potássio/administração & dosagem , Compostos de Potássio/farmacologia , Substâncias Protetoras/administração & dosagem , Distribuição Aleatória , Plântula/efeitos dos fármacos , Plântula/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/fisiologia
17.
J Integr Plant Biol ; 60(5): 397-411, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29247597

RESUMO

Species and hybrids of Eucalyptus are the world's most widely planted hardwood trees. They are cultivated across a wide range of latitudes and therefore environmental conditions. In this context, comprehensive metabolomics approaches have been used to assess how different temperature regimes may affect the metabolism of three species of Eucalyptus, E. dunnii, E. grandis and E. pellita. Young plants were grown for 53 d in the greenhouse and then transferred to growth chambers at 10°C, 20°C or 30°C for another 7 d. In all three species the leaf chlorophyll content was positively correlated to temperature, and in E. pellita the highest temperature also resulted in a significant increase in stem biomass. Comprehensive metabolomics was performed using untargeted gas chromatography mass spectrometry (GC-MS) and liquid chromatography (LC)-MS. This approach enabled the comparison of the relative abundance of 88 polar primary metabolites from GC-MS and 625 semi-polar secondary metabolites from LC-MS. Using principal components analysis, a major effect of temperature was observed in each species which was larger than that resulting from the genetic background. Compounds mostly affected by temperature treatment were subsequently selected using partial least squares discriminant analysis and were further identified. These putative annotations indicated that soluble sugars and several polyphenols, including tannins, triterpenes and alkaloids were mostly influenced.


Assuntos
Eucalyptus/metabolismo , Metabolômica , Temperatura , Clorofila/metabolismo , Cromatografia Líquida , Análise Discriminante , Eucalyptus/genética , Cromatografia Gasosa-Espectrometria de Massas , Genótipo , Análise dos Mínimos Quadrados , Metaboloma , Anotação de Sequência Molecular , Análise Multivariada , Análise de Componente Principal , Especificidade da Espécie , Açúcares/metabolismo
18.
Transgenic Res ; 26(4): 447-463, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28349287

RESUMO

The expression of cell-wall-targeted Carbohydrate Binding Modules (CBMs) can alter cell wall properties and modulate growth and development in plants such as tobacco and potato. CBM2a identified in xylanase 10A from Cellulomonas fimi is of particular interest for its ability to bind crystalline cellulose. However, its potential for promoting plant growth has not been explored. In this work, we tested the ability of CBM2a to promote growth when expressed using both CaMV35S and a vascular tissue-specific promoter derived from Arabidopsis expansin4 (AtEXP4) in three plant species: Arabidopsis, Nicotiana tabacum and Eucalyptus camaldulensis. In Arabidopsis, the expression of AtEXP4pro:CBM2a showed trends for growth promoting effects including the increase of root and hypocotyl lengths and the enlargements of the vascular xylem area, fiber cells and vessel cells. However, in N. tabacum, the expression of CBM2a under the control of either CaMV35S or AtEXP4 promoter resulted in subtle changes in the plant growth, and the thickness of secondary xylem and vessel and fiber cell sizes were generally reduced in the transgenic lines with AtEXP4pro:CBM2a. In Eucalyptus, while transgenics expressing CaMV35S:CBM2a showed very subtle changes compared to wild type, those transgenics with AtEXP4pro:CBM2a showed increases in plant height, enlargement of xylem areas and xylem fiber and vessel cells. These data provide comparative effects of expressing CBM2a protein in different plant species, and this finding can be applied for plant biomass improvement.


Assuntos
Carboidratos/genética , Endo-1,4-beta-Xilanases/genética , Proteínas de Plantas/genética , Plantas Geneticamente Modificadas/genética , Xilema/genética , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Parede Celular/genética , Celulose/genética , Eucalyptus/genética , Eucalyptus/crescimento & desenvolvimento , Regulação da Expressão Gênica de Plantas , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Regiões Promotoras Genéticas , Nicotiana/genética , Nicotiana/crescimento & desenvolvimento , Xilema/crescimento & desenvolvimento
19.
Genet Mol Res ; 15(4)2016 Dec 02.
Artigo em Inglês | MEDLINE | ID: mdl-27966751

RESUMO

Cinnamyl alcohol dehydrogenase (CAD) catalyzes the final step in lignin biosynthesis. The genus Eucalyptus belongs to the family Myrtaceae, which is the main cultivated species in China. Eucalyptus urophylla GLU4 (GLU4) is widely grown in Guangxi. It is preferred for pulping because of its excellent cellulose content and fiber length. Based on GLU4 and CAD gene expression, a Eucalyptus variety low in lignin content should be obtained using transgenic technology, which could reduce the cost of pulp and improve the pulping rate, and have favorable prospects for application. However, the role and function of CAD in GLU4 is still unclear. In the present study, EuCAD was cloned from GLU4 and identified using bioinformatic tools. Subsequently, in order to evaluate its impact on lignin synthesis, a full-length EuCAD RNAi vector was constructed, and transgenic tobacco was obtained via Agrobacterium-mediated transformation. A significant decrease in CAD expression and lignin content in transgenic tobacco demonstrated a key role for EuCAD in lignin biosynthesis and established a regulatory role for RNAi. In our study, the direct molecular basis of EuCAD expression was determined, and the potential regulatory effects of this RNAi vector on lignin biosynthesis in E. urophylla GLU4 were demonstrated. Our results provide a theoretical basis for the study of lignin biosynthesis in Eucalyptus.


Assuntos
Oxirredutases do Álcool/genética , Clonagem Molecular/métodos , Eucalyptus/enzimologia , Nicotiana/genética , Oxirredutases do Álcool/metabolismo , China , Eucalyptus/genética , Regulação da Expressão Gênica de Plantas , Lignina/biossíntese , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Nicotiana/crescimento & desenvolvimento
20.
Mol Phylogenet Evol ; 105: 63-85, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27530705

RESUMO

We used genome-wide markers and an unprecedented scale of sampling to construct a phylogeny for a globally significant Eucalyptus lineage that has been impacted by hybridisation, recent radiation and morphological convergence. Our approach, using 3109 DArT markers distributed throughout the genome and 540 samples covering 185 terminal taxa in sections Maidenaria, Exsertaria, Latoangulatae and related smaller sections, with multiple geographically widespread samples per terminal taxon, produced a phylogeny that largely matched the morphological treatment of sections, though sections Exsertaria and Latoangulatae were polyphyletic. At lower levels there were numerous inconsistencies between the morphological treatment and the molecular phylogeny, and taxa within the three main sections were generally not monophyletic at the series (at least 62% polyphyly) or species (at least 52% polyphyly) level. Some of the discrepancies appear to be the result of morphological convergence or misclassifications, and we propose some taxonomic reassessments to address this. However, many inconsistencies appear to be the products of incomplete speciation and/or hybridisation. Our analysis represents a significant advance on previous phylogenies of these important eucalypt sections (which have mainly used single samples to represent each species), thus providing a robust phylogenetic framework for evolutionary and ecological studies.


Assuntos
Eucalyptus/classificação , Eucalyptus/genética , Genoma de Planta , Internacionalidade , Filogenia , Marcadores Genéticos , Análise de Componente Principal , Especificidade da Espécie
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