RESUMEN
Cellulose is the main structural component in the plant cell walls. We show that two glycosyltransferase family 31 (GT31) enzymes of Arabidopsis thaliana, here named cellulose synthesis associated glycosyltransferases 1 and 2 (CAGE1 and 2), influence both primary and secondary cell wall cellulose biosynthesis. cage1cage2 mutants show primary cell wall defects manifesting as impaired growth and cell expansion in seedlings and etiolated hypocotyls, along with secondary cell wall defects, apparent as collapsed xylem vessels and reduced xylem wall thickness in the inflorescence stem. Single and double cage mutants also show increased sensitivity to the cellulose biosynthesis inhibitor isoxaben. The cage1cage2 phenotypes were associated with an approximately 30% reduction in cellulose content, an approximately 50% reduction in secondary cell wall CELLULOSE SYNTHASE (CESA) protein levels in stems and reduced cellulose biosynthesis rate in seedlings. CESA transcript levels were not significantly altered in cage1cage2 mutants, suggesting that the reduction in CESA levels was caused by a post-transcriptional mechanism. Both CAGE1 and 2 localize to the Golgi apparatus and are predicted to synthesize ß-1,3-galactans on arabinogalactan proteins. In line with this, the cage1cage2 mutants exhibit reduced levels of ß-Yariv binding to arabinogalactan protein linked ß-1,3-galactan. This leads us to hypothesize that defects in arabinogalactan biosynthesis underlie the cellulose deficiency of the mutants.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Pared Celular/metabolismo , Celulosa/metabolismo , Glucosiltransferasas/genética , Glucosiltransferasas/metabolismo , Glicosiltransferasas/genética , Glicosiltransferasas/metabolismo , Aparato de Golgi/metabolismo , Plantones/genética , Plantones/metabolismoRESUMEN
Biosynthesis of plant cell walls requires UDP-glucose as the substrate for cellulose biosynthesis, and as an intermediate for the synthesis of other matrix polysaccharides. The sucrose cleaving enzyme sucrose synthase (SUS) is thought to have a central role in UDP-glucose biosynthesis, and a long-held and much debated hypothesis postulates that SUS is required to supply UDP-glucose to cellulose biosynthesis. To investigate the role of SUS in cellulose biosynthesis of Arabidopsis thaliana we characterized mutants in which four or all six Arabidopsis SUS genes were disrupted. These sus mutants showed no growth phenotypes, vascular tissue cell wall defects, or changes in cellulose content. Moreover, the UDP-glucose content of rosette leaves of the sextuple sus mutants was increased by approximately 20% compared with wild type. It can thus be concluded that cellulose biosynthesis is able to employ alternative UDP-glucose biosynthesis pathway(s), and thereby the model of SUS requirements for cellulose biosynthesis in Arabidopsis can be refuted.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Pared Celular/metabolismo , Celulosa/metabolismo , Glucosa/metabolismo , Glucosiltransferasas/genética , Glucosiltransferasas/metabolismo , Sacarosa/metabolismo , Uridina Difosfato Glucosa/química , Uridina Difosfato Glucosa/metabolismoRESUMEN
Hydrogels of cellulose nanofibrils (CNFs) are promising wound dressing candidates due to their biocompatibility, high water absorption, and transparency. Herein, two different commercially available wood species, softwood and hardwood, were subjected to TEMPO-mediated oxidation to proceed with delignification and oxidation in a one-pot process, and thereafter, nanofibrils were isolated using a high-pressure microfluidizer. Furthermore, transparent nanofibril hydrogel networks were prepared by vacuum filtration. Nanofibril properties and network performance correlated with oxidation were investigated and compared with commercially available TEMPO-oxidized pulp nanofibrils and their networks. Softwood nanofibril hydrogel networks exhibited the best mechanical properties, and in vitro toxicological risk assessment showed no detrimental effect for any of the studied hydrogels on human fibroblast or keratinocyte cells. This study demonstrates a straightforward processing route for direct oxidation of different wood species to obtain nanofibril hydrogels for potential use as wound dressings, with softwood having the most potential.
Asunto(s)
Celulosa , Hidrogeles , Humanos , Vendajes , Oxidación-Reducción , FibroblastosRESUMEN
Currently one-third of the proteins encoded by the Arabidopsis (Arabidopsis thaliana) genome are of unknown function. Some of these unknown proteins are likely to be involved in uncharacterized vital biological processes. Evolutionarily conserved single copy genes in flowering plants have been shown to be enriched in essential housekeeping functions. This together with publicly available gene expression data allows for a focused search for uncharacterized essential genes. Here we identify an essential single copy gene called OPENER (OPNR) in Arabidopsis. We show that OPNR is predominantly expressed in actively dividing cells and performs essential functions in seed development and root meristem maintenance. Cell cycle tracking using 5-ethynyl-2'-deoxyuridine staining and fluorescent cell cycle markers together with the increased size of nucleolus and nucleus in opnr mutants indicate that OPNR is required for cell cycle progression through the S or G2 phases. Intriguingly, OPNR localizes to the nuclear envelope and mitochondria. Furthermore, the nuclear envelope localization of OPNR is dependent on its interaction with nuclear inner membrane Sad1/UNC-84 (SUN) domain proteins SUN1 and SUN2. Taken together our results open a line of investigation into an evolutionarily conserved essential cellular process occurring in both the nuclear envelopes and mitochondria of dividing cells.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/citología , Arabidopsis/metabolismo , Ciclo Celular , Mitocondrias/metabolismo , Proteínas Mitocondriales/metabolismo , Membrana Nuclear/metabolismo , Proteínas Nucleares/metabolismo , Alelos , Arabidopsis/embriología , Arabidopsis/genética , Proliferación Celular , Secuencia Conservada/genética , ADN Bacteriano/genética , Endospermo/metabolismo , Dosificación de Gen , Regulación de la Expresión Génica de las Plantas , Genes Esenciales , Pruebas Genéticas , Proteínas de la Membrana/metabolismo , Mitocondrias/ultraestructura , Tamaño Mitocondrial , Mutagénesis Insercional/genética , Mutación/genética , Fenotipo , Raíces de Plantas/citología , Transporte de ProteínasRESUMEN
Plant arabinogalactan proteins (AGPs) are a diverse group of cell surface- and wall-associated glycoproteins. Functionally important AGP glycans are synthesized in the Golgi apparatus, but the relationships among their glycosylation levels, processing, and functionalities are poorly understood. Here, we report the identification and functional characterization of two Golgi-localized exo-ß-1,3-galactosidases from the glycosyl hydrolase 43 (GH43) family in Arabidopsis thaliana GH43 loss-of-function mutants exhibited root cell expansion defects in sugar-containing growth media. This root phenotype was associated with an increase in the extent of AGP cell wall association, as demonstrated by Yariv phenylglycoside dye quantification and comprehensive microarray polymer profiling of sequentially extracted cell walls. Characterization of recombinant GH43 variants revealed that the exo-ß-1,3-galactosidase activity of GH43 enzymes is hindered by ß-1,6 branches on ß-1,3-galactans. In line with this steric hindrance, the recombinant GH43 variants did not release galactose from cell wall-extracted glycoproteins or AGP-rich gum arabic. These results indicate that the lack of exo-ß-1,3-galactosidase activity alters cell wall extensibility in roots, a phenotype that could be explained by the involvement of galactosidases in AGP glycan biosynthesis.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crecimiento & desarrollo , Galactosiltransferasas/metabolismo , Glicósido Hidrolasas/metabolismo , Mucoproteínas/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Galactosiltransferasas/genética , Glicósido Hidrolasas/genética , Mucoproteínas/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raíces de Plantas/genéticaRESUMEN
Cellulose microfibrils synthesized by CELLULOSE SYNTHASE COMPLEXES (CSCs) are the main load-bearing polymers in wood. CELLULOSE SYNTHASE INTERACTING1 (CSI1) connects CSCs with cortical microtubules, which align with cellulose microfibrils. Mechanical properties of wood are dependent on cellulose microfibril alignment and structure in the cell walls, but the molecular mechanism(s) defining these features is unknown. Herein, we investigated the role of CSI1 in hybrid aspen (Populus tremula × Populus tremuloides) by characterizing transgenic lines with significantly reduced CSI1 transcript abundance. Reduction in leaves (50-80%) caused leaf twisting and misshaped pavement cells, while reduction (70-90%) in developing xylem led to impaired mechanical wood properties evident as a decrease in the elastic modulus and rupture. X-ray diffraction measurements indicate that microfibril angle was not impacted by the altered CSI1 abundance in developing wood fibres. Instead, the augmented wood phenotype of the transgenic trees was associated with a reduced cellulose degree of polymerization. These findings establish a function for CSI1 in wood mechanics and in defining leaf cell shape. Furthermore, the results imply that the microfibril angle in wood is defined by CSI1 independent mechanism(s).
Asunto(s)
Hojas de la Planta/anatomía & histología , Proteínas de Plantas/fisiología , Populus/anatomía & histología , Madera/anatomía & histología , Proteínas de Arabidopsis/genética , Proteínas Portadoras/genética , Celulosa/metabolismo , Filogenia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , Populus/genética , Populus/metabolismo , Resistencia a la Tracción , Árboles/anatomía & histología , Árboles/metabolismo , Xilema/anatomía & histologíaRESUMEN
Despite the ecological and industrial importance of biomass accumulation in wood, the control of carbon (C) allocation to this tissue and to other tree tissues remain poorly understood. We studied sucrose synthase (SUS) to clarify its role in biomass formation and C metabolism at the whole tree level in hybrid aspen (Populus tremula × tremuloides). To this end, we analysed source leaves, phloem, developing wood, and roots of SUSRNAi trees using a combination of metabolite profiling, 13 CO2 pulse labelling experiments, and long-term field experiments. The glasshouse grown SUSRNAi trees exhibited a mild stem phenotype together with a reduction in wood total C. The 13 CO2 pulse labelling experiments showed an alteration in the C flow in all the analysed tissues, indicating that SUS affects C metabolism at the whole tree level. This was confirmed when the SUSRNAi trees were grown in the field over a 5-yr period; their stem height, diameter and biomass were substantially reduced. These results establish that SUS influences C allocation to developing wood, and that it affects C metabolism at the whole tree level.
Asunto(s)
Populus , Madera , Carbono , Glucosiltransferasas , Populus/genética , ÁrbolesRESUMEN
Norway spruce is a boreal forest tree species of significant ecological and economic importance. Hence there is a strong imperative to dissect the genetics underlying important wood quality traits in the species. We performed a functional genome-wide association study (GWAS) of 17 wood traits in Norway spruce using 178 101 single nucleotide polymorphisms (SNPs) generated from exome genotyping of 517 mother trees. The wood traits were defined using functional modelling of wood properties across annual growth rings. We applied a Least Absolute Shrinkage and Selection Operator (LASSO-based) association mapping method using a functional multilocus mapping approach that utilizes latent traits, with a stability selection probability method as the hypothesis testing approach to determine a significant quantitative trait locus. The analysis provided 52 significant SNPs from 39 candidate genes, including genes previously implicated in wood formation and tree growth in spruce and other species. Our study represents a multilocus GWAS for complex wood traits in Norway spruce. The results advance our understanding of the genetics influencing wood traits and identifies candidate genes for future functional studies.
Asunto(s)
Genes de Plantas/genética , Estudio de Asociación del Genoma Completo/métodos , Picea/genética , Sitios de Carácter Cuantitativo/genética , Madera/genética , Algoritmos , Genómica/métodos , Genotipo , Desequilibrio de Ligamiento , Noruega , Fenotipo , Picea/clasificación , Polimorfismo de Nucleótido Simple , Madera/clasificaciónRESUMEN
Wood, or secondary xylem, is the product of xylogenesis, a developmental process that begins with the proliferation of cambial derivatives and ends with mature xylem fibers and vessels with lignified secondary cell walls. Fully mature xylem has undergone a series of cellular processes, including cell division, cell expansion, secondary wall formation, lignification and programmed cell death. A complex network of interactions between transcriptional regulators and signal transduction pathways controls wood formation. However, the role of metabolites during this developmental process has not been comprehensively characterized. To evaluate the role of metabolites during wood formation, we performed a high spatial resolution metabolomics study of the wood-forming zone of Populus tremula, including laser dissected aspen ray and fiber cells. We show that metabolites show specific patterns within the wood-forming zone, following the differentiation process from cell division to cell death. The data from profiled laser dissected aspen ray and fiber cells suggests that these two cell types host distinctly different metabolic processes. Furthermore, by integrating previously published transcriptomic and proteomic profiles generated from the same trees, we provide an integrative picture of molecular processes, for example, deamination of phenylalanine during lignification is of critical importance for nitrogen metabolism during wood formation.
Asunto(s)
Populus , Proteómica , Madera , Cámbium , Regulación de la Expresión Génica de las Plantas , Populus/genética , XilemaRESUMEN
The evolution of the plant vasculature was essential for the emergence of terrestrial life. Xylem vessels are solute-transporting elements in the vasculature that possess secondary wall thickenings deposited in intricate patterns. Evenly dispersed microtubule (MT) bands support the formation of these wall thickenings, but how the MTs direct cell wall synthesis during this process remains largely unknown. Cellulose is the major secondary wall constituent and is synthesized by plasma membrane-localized cellulose synthases (CesAs) whose catalytic activity propels them through the membrane. We show that the protein CELLULOSE SYNTHASE INTERACTING1 (CSI1)/POM2 is necessary to align the secondary wall CesAs and MTs during the initial phase of xylem vessel development in Arabidopsis thaliana and rice (Oryza sativa). Surprisingly, these MT-driven patterns successively become imprinted and sufficient to sustain the continued progression of wall thickening in the absence of MTs and CSI1/POM2 function. Hence, two complementary principles underpin wall patterning during xylem vessel development.
Asunto(s)
Pared Celular/metabolismo , Xilema/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Membrana Celular/metabolismo , Pared Celular/genética , Celulosa/metabolismo , Glucosiltransferasas/metabolismo , Microtúbulos/metabolismo , Xilema/genéticaRESUMEN
Trees represent the largest terrestrial carbon sink and a renewable source of ligno-cellulose. There is significant scope for yield and quality improvement in these largely undomesticated species, and efforts to engineer elite varieties will benefit from improved understanding of the transcriptional network underlying cambial growth and wood formation. We generated high-spatial-resolution RNA sequencing data spanning the secondary phloem, vascular cambium, and wood-forming tissues of Populus tremula The transcriptome comprised 28,294 expressed, annotated genes, 78 novel protein-coding genes, and 567 putative long intergenic noncoding RNAs. Most paralogs originating from the Salicaceae whole-genome duplication had diverged expression, with the exception of those highly expressed during secondary cell wall deposition. Coexpression network analyses revealed that regulation of the transcriptome underlying cambial growth and wood formation comprises numerous modules forming a continuum of active processes across the tissues. A comparative analysis revealed that a majority of these modules are conserved in Picea abies The high spatial resolution of our data enabled identification of novel roles for characterized genes involved in xylan and cellulose biosynthesis, regulators of xylem vessel and fiber differentiation and lignification. An associated web resource (AspWood, http://aspwood.popgenie.org) provides interactive tools for exploring the expression profiles and coexpression network.
Asunto(s)
Populus/genética , Transcriptoma , Madera/crecimiento & desarrollo , Madera/genética , Pared Celular/genética , Pared Celular/metabolismo , Regulación de la Expresión Génica de las Plantas , Internet , Meristema/genética , Polisacáridos/genética , Polisacáridos/metabolismo , Populus/citología , Populus/crecimiento & desarrollo , Madera/citología , Xilema/genéticaRESUMEN
Cellulose is synthesized at the plasma membrane by cellulose synthase complexes (CSCs) containing cellulose synthases (CESAs). Genetic analysis and CESA isoform quantification indicate that cellulose in the secondary cell walls of Arabidopsis (Arabidopsis thaliana) is synthesized by isoforms CESA4, CESA7, and CESA8 in equimolar amounts. Here, we used quantitative proteomics to investigate whether the CSC model based on Arabidopsis secondary cell wall CESA stoichiometry can be applied to the angiosperm tree aspen (Populus tremula) and the gymnosperm tree Norway spruce (Picea abies). In the developing xylem of aspen, the secondary cell wall CESA stoichiometry was 3:2:1 for PtCESA8a/b:PtCESA4:PtCESA7a/b, while in Norway spruce, the stoichiometry was 1:1:1, as observed previously in Arabidopsis. Furthermore, in aspen tension wood, the secondary cell wall CESA stoichiometry changed to 8:3:1 for PtCESA8a/b:PtCESA4:PtCESA7a/b. PtCESA8b represented 73% of the total secondary cell wall CESA pool, and quantitative polymerase chain reaction analysis of CESA transcripts in cryosectioned tension wood revealed increased PtCESA8b expression during the formation of the cellulose-enriched gelatinous layer, while the transcripts of PtCESA4, PtCESA7a/b, and PtCESA8a decreased. A wide-angle x-ray scattering analysis showed that the shift in CESA stoichiometry in tension wood coincided with an increase in crystalline cellulose microfibril diameter, suggesting that the CSC CESA composition influences microfibril properties. The aspen CESA stoichiometry results raise the possibility of alternative CSC models and suggest that homomeric PtCESA8b complexes are responsible for cellulose biosynthesis in the gelatinous layer in tension wood.
Asunto(s)
Arabidopsis/enzimología , Glucosiltransferasas/metabolismo , Picea/enzimología , Proteínas de Plantas/metabolismo , Populus/enzimología , Arabidopsis/citología , Proteínas de Arabidopsis/metabolismo , Pared Celular/enzimología , Glucosiltransferasas/aislamiento & purificación , Péptidos/análisis , Péptidos/metabolismo , Picea/citología , Proteínas de Plantas/aislamiento & purificación , Populus/citología , Proteómica/métodos , Dispersión de Radiación , Especificidad de la Especie , Xilema/metabolismoRESUMEN
Conifers have dominated forests for more than 200 million years and are of huge ecological and economic importance. Here we present the draft assembly of the 20-gigabase genome of Norway spruce (Picea abies), the first available for any gymnosperm. The number of well-supported genes (28,354) is similar to the >100 times smaller genome of Arabidopsis thaliana, and there is no evidence of a recent whole-genome duplication in the gymnosperm lineage. Instead, the large genome size seems to result from the slow and steady accumulation of a diverse set of long-terminal repeat transposable elements, possibly owing to the lack of an efficient elimination mechanism. Comparative sequencing of Pinus sylvestris, Abies sibirica, Juniperus communis, Taxus baccata and Gnetum gnemon reveals that the transposable element diversity is shared among extant conifers. Expression of 24-nucleotide small RNAs, previously implicated in transposable element silencing, is tissue-specific and much lower than in other plants. We further identify numerous long (>10,000 base pairs) introns, gene-like fragments, uncharacterized long non-coding RNAs and short RNAs. This opens up new genomic avenues for conifer forestry and breeding.
Asunto(s)
Evolución Molecular , Genoma de Planta/genética , Picea/genética , Secuencia Conservada/genética , Elementos Transponibles de ADN/genética , Silenciador del Gen , Genes de Plantas/genética , Genómica , Internet , Intrones/genética , Fenotipo , ARN no Traducido/genética , Análisis de Secuencia de ADN , Secuencias Repetidas Terminales/genética , Transcripción Genética/genéticaRESUMEN
Wood biosynthesis defines the chemical and structural properties of wood. The metabolic pathways that produce the precursors of wood cell wall polymers have a central role in defining wood properties. To make rational design of wood properties feasible, we need not only to understand the cell wall biosynthetic machinery, but also how sucrose transport and metabolism in developing wood connect to cell wall biosynthesis and how they respond to genetic and environmental cues. Here, we review the current understanding of the sucrose transport and primary metabolism pathways leading to the precursors of cell wall biosynthesis in woody plant tissues. We present both old, persistent questions and new emerging themes with a focus on wood formation in trees and draw upon evidence from the xylem tissues of herbaceous plants when it is relevant.
Asunto(s)
Pared Celular/metabolismo , Sacarosa/metabolismo , Madera , Transporte Biológico/fisiología , Ciclo del Carbono/fisiología , Regulación de la Expresión Génica de las PlantasRESUMEN
Carbon for cellulose biosynthesis is derived from sucrose. Cellulose is synthesized from uridine 5'-diphosphoglucose (UDP-glucose), but the enzyme(s) responsible for the initial sucrose cleavage and the source of UDP-glucose for cellulose biosynthesis in developing wood have not been defined. We investigated the role of CYTOSOLIC INVERTASEs (CINs) during wood formation in hybrid aspen (Populus tremula × tremuloides) and characterized transgenic lines with reduced CIN activity during secondary cell wall biosynthesis. Suppression of CIN activity by 38-55% led to a 9-13% reduction in crystalline cellulose. The changes in cellulose were reflected in reduced diameter of acid-insoluble cellulose microfibrils and increased glucose release from wood upon enzymatic digestion of cellulose. Reduced CIN activity decreased the amount of the cellulose biosynthesis precursor UDP-glucose in developing wood, pointing to the likely cause of the cellulose phenotype. The findings suggest that CIN activity has an important role in the cellulose biosynthesis of trees, and indicate that cellulose biosynthesis in wood relies on a quantifiable UDP-glucose pool. The results also introduce a concept of altering cellulose microfibril properties by modifying substrate supply to cellulose biosynthesis.
Asunto(s)
Celulosa/biosíntesis , Citosol/enzimología , Populus/enzimología , Madera/enzimología , beta-Fructofuranosidasa/metabolismo , Cristalización , Regulación de la Expresión Génica de las Plantas , Glucosa/metabolismo , Concentración de Iones de Hidrógeno , Metaboloma , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , Interferencia de ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , Solubilidad , Especificidad por Sustrato , Transcriptoma/genética , Árboles/genéticaRESUMEN
The contribution of transcriptional and post-transcriptional regulation to modifying carbon allocation to developing wood of trees is not well defined. To clarify the role of transcriptional regulation, the enzyme activity patterns of eight central primary metabolism enzymes across phloem, cambium, and developing wood of aspen (Populus tremula L.) were compared with transcript levels obtained by RNA sequencing of sequential stem sections from the same trees. Enzymes were selected on the basis of their importance in sugar metabolism and in linking primary metabolism to lignin biosynthesis. Existing enzyme assays were adapted to allow measurements from ~1 mm3 sections of dissected stem tissue. These experiments provided high spatial resolution of enzyme activity changes across different stages of wood development, and identified the gene transcripts probably responsible for these changes. In most cases, there was a clear positive relationship between transcripts and enzyme activity. During secondary cell wall formation, the increases in transcript levels and enzyme activities also matched with increased levels of glucose, fructose, hexose phosphates, and UDP-glucose, emphasizing an important role for transcriptional regulation in carbon allocation to developing aspen wood. These observations corroborate the efforts to increase carbon allocation to wood by engineering gene regulatory networks.
Asunto(s)
Carbono/metabolismo , Regulación de la Expresión Génica de las Plantas , Proteínas de Plantas/genética , Populus/genética , Transcripción Genética , Cámbium/enzimología , Cámbium/crecimiento & desarrollo , Floema/enzimología , Floema/crecimiento & desarrollo , Proteínas de Plantas/metabolismo , Populus/enzimología , Madera/enzimología , Madera/crecimiento & desarrolloRESUMEN
Wood of trees is formed from carbon assimilated in the photosynthetic tissues. Determining the temporal dynamics of carbon assimilation, subsequent transport into developing wood, and incorporation to cell walls would further our understanding of wood formation in particular and tree growth in general. To investigate these questions, we designed a (13)CO2 labeling system to study carbon transport and incorporation to developing wood of hybrid aspen (Populus tremula × tremuloides). Tracking of (13)C incorporation to wood over a time course using nuclear magnetic resonance spectroscopy revealed diurnal patterns in wood cell wall biosynthesis. The dark period had a differential effect on (13)C incorporation to lignin and cell wall carbohydrates. No (13)C was incorporated into aromatic amino acids of cell wall proteins in the dark, suggesting that cell wall protein biosynthesis ceased during the night. The results show previously unrecognized temporal patterns in wood cell wall biosynthesis, suggest diurnal cycle as a possible cue in the regulation of carbon incorporation to wood, and establish a unique (13)C labeling method for the analysis of wood formation and secondary growth in trees.
Asunto(s)
Dióxido de Carbono/metabolismo , Ritmo Circadiano , Populus/fisiología , Madera/crecimiento & desarrollo , Análisis de Varianza , Isótopos de Carbono , Pared Celular/metabolismo , Celulosa/metabolismo , Espectroscopía de Resonancia Magnética , Redes y Vías Metabólicas , Metaboloma , Modelos Biológicos , Floema/metabolismo , Hojas de la Planta/metabolismo , Análisis de Componente Principal , Sacarosa/metabolismoRESUMEN
The biosynthesis of wood in aspen (Populus) depends on the metabolism of sucrose, which is the main transported form of carbon from source tissues. The largest fraction of the wood biomass is cellulose, which is synthesized from UDP-glucose. Sucrose synthase (SUS) has been proposed previously to interact directly with cellulose synthase complexes and specifically supply UDP-glucose for cellulose biosynthesis. To investigate the role of SUS in wood biosynthesis, we characterized transgenic lines of hybrid aspen with strongly reduced SUS activity in developing wood. No dramatic growth phenotypes in glasshouse-grown trees were observed, but chemical fingerprinting with pyrolysis-GC/MS, together with micromechanical analysis, showed notable changes in chemistry and ultrastructure of the wood in the transgenic lines. Wet chemical analysis showed that the dry weight percentage composition of wood polymers was not changed significantly. However, a decrease in wood density was observed and, consequently, the content of lignin, hemicellulose and cellulose was decreased per wood volume. The decrease in density was explained by a looser structure of fibre cell walls as shown by increased wall shrinkage on drying. The results show that SUS is not essential for cellulose biosynthesis, but plays a role in defining the total carbon incorporation to wood cell walls.
Asunto(s)
Pared Celular/metabolismo , Celulosa/biosíntesis , Glucosiltransferasas/deficiencia , Populus/enzimología , Populus/crecimiento & desarrollo , Madera/enzimología , Madera/crecimiento & desarrollo , Arabidopsis/enzimología , Fenómenos Biomecánicos , Cruzamientos Genéticos , Regulación de la Expresión Génica de las Plantas , Glucosiltransferasas/genética , Glucosiltransferasas/metabolismo , Populus/anatomía & histología , Populus/genética , Interferencia de ARN , Solubilidad , Transcriptoma/genética , Madera/anatomía & histología , Madera/genéticaRESUMEN
Wood formation in trees requires carbon import from the photosynthetic tissues. In several tree species, including Populus species, the majority of this carbon is derived from sucrose (Suc) transported in the phloem. The mechanism of radial Suc transport from phloem to developing wood is not well understood. We investigated the role of active Suc transport during secondary cell wall formation in hybrid aspen (Populus tremula × Populus tremuloides). We show that RNA interference-mediated reduction of PttSUT3 (for Suc/H(+) symporter) during secondary cell wall formation in developing wood caused thinner wood fiber walls accompanied by a reduction in cellulose and an increase in lignin. Suc content in the phloem and developing wood was not significantly changed. However, after (13)CO2 assimilation, the SUT3RNAi lines contained more (13)C than the wild type in the Suc-containing extract of developing wood. Hence, Suc was transported into developing wood, but the Suc-derived carbon was not efficiently incorporated to wood fiber walls. A yellow fluorescent protein:PttSUT3 fusion localized to plasma membrane, suggesting that reduced Suc import into developing wood fibers was the cause of the observed cell wall phenotype. The results show the importance of active Suc transport for wood formation in a symplasmically phloem-loading tree species and identify PttSUT3 as a principal transporter for carbon delivery into secondary cell wall-forming wood fibers.
Asunto(s)
Carbono/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Proteínas de Plantas/metabolismo , Populus/metabolismo , Madera/metabolismo , Dióxido de Carbono/metabolismo , Isótopos de Carbono , Membrana Celular/metabolismo , Fructosa/metabolismo , Regulación de la Expresión Génica de las Plantas , Glucosa/metabolismo , Proteínas de Transporte de Membrana/genética , Fenotipo , Floema/metabolismo , Hojas de la Planta/anatomía & histología , Hojas de la Planta/metabolismo , Proteínas de Plantas/genética , Tallos de la Planta/crecimiento & desarrollo , Tallos de la Planta/metabolismo , Populus/genética , Populus/crecimiento & desarrollo , Populus/ultraestructura , Transporte de Proteínas , Interferencia de ARN , ARN Mensajero/genética , ARN Mensajero/metabolismo , Fracciones Subcelulares/metabolismo , Sacarosa/metabolismo , Madera/anatomía & histología , Madera/crecimiento & desarrollo , Madera/ultraestructuraRESUMEN
Sucrose is the main transported form of carbon in several plant species, including Populus species. Sucrose metabolism in developing wood has therefore a central role in carbon partitioning to stem biomass. Half of the sucrose-derived carbon is in the form of fructose, but metabolism of fructose has received little attention as a factor in carbon partitioning to walls of wood cells. We show that RNAi-mediated reduction of FRK2 activity in developing wood of hybrid aspen (Populus tremula × tremuloides) led to the accumulation of soluble neutral sugars and a decrease in hexose phosphates and UDP-glucose, indicating that carbon flux to cell-wall polysaccharide precursors is decreased. Reduced FRK2 activity also led to thinner fiber cell walls with a reduction in the proportion of cellulose. No pleiotropic effects on stem height or diameter were observed. The results establish a central role for FRK2 activity in carbon flux to wood cellulose.