RESUMEN
P4B (2-phenyl-1-[4-(6-(piperidin-1-yl) pyridazin-3-yl) piperazin-1-yl] butan-1-one) is a novel cellulose biosynthesis inhibitor (CBI) discovered in a screen for molecules to identify inhibitors of Arabidopsis (Arabidopsis thaliana) seedling growth. Growth and cellulose synthesis inhibition by P4B were greatly reduced in a novel mutant for the cellulose synthase catalytic subunit gene CESA3 (cesa3pbr1). Cross-tolerance to P4B was also observed for isoxaben-resistant (ixr) cesa3 mutants ixr1-1 and ixr1-2. P4B has an original mode of action as compared with most other CBIs. Indeed, short-term treatments with P4B did not affect the velocity of cellulose synthase complexes (CSCs) but led to a decrease in CSC density in the plasma membrane without affecting their accumulation in microtubule-associated compartments. This was observed in the wild type but not in a cesa3pbr1 background. This reduced density correlated with a reduced delivery rate of CSCs to the plasma membrane but also with changes in cortical microtubule dynamics and orientation. At longer timescales, however, the responses to P4B treatments resembled those to other CBIs, including the inhibition of CSC motility, reduced growth anisotropy, interference with the assembly of an extensible wall, pectin demethylesterification, and ectopic lignin and callose accumulation. Together, the data suggest that P4B either directly targets CESA3 or affects another cellular function related to CSC plasma membrane delivery and/or microtubule dynamics that is bypassed specifically by mutations in CESA3.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Celulosa , Glucosiltransferasas , Microtúbulos , Glucosiltransferasas/metabolismo , Glucosiltransferasas/genética , Microtúbulos/metabolismo , Microtúbulos/efectos de los fármacos , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Arabidopsis/metabolismo , Celulosa/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Mutación , Plantones/genética , Plantones/crecimiento & desarrollo , Plantones/efectos de los fármacos , Membrana Celular/metabolismo , Piridazinas/farmacología , Pared Celular/metabolismo , Pared Celular/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , BenzamidasRESUMEN
Pectin methylesterases (PMEs) modify homogalacturonan's chemistry and play a key role in regulating primary cell wall mechanical properties. Here, we report on Arabidopsis AtPME2, which we found to be highly expressed during lateral root emergence and dark-grown hypocotyl elongation. We showed that dark-grown hypocotyl elongation was reduced in knock-out mutant lines as compared to the control. The latter was related to the decreased total PME activity as well as increased stiffness of the cell wall in the apical part of the hypocotyl. To relate phenotypic analyses to the biochemical specificity of the enzyme, we produced the mature active enzyme using heterologous expression in Pichia pastoris and characterized it through the use of a generic plant PME antiserum. AtPME2 is more active at neutral compared to acidic pH, on pectins with a degree of 55-70% methylesterification. We further showed that the mode of action of AtPME2 can vary according to pH, from high processivity (at pH8) to low processivity (at pH5), and relate these observations to the differences in electrostatic potential of the protein. Our study brings insights into how the pH-dependent regulation by PME activity could affect the pectin structure and associated cell wall mechanical properties.
Asunto(s)
Arabidopsis , Hidrolasas de Éster Carboxílico , Hipocótilo , Hipocótilo/genética , Hipocótilo/metabolismo , Arabidopsis/metabolismo , Pared Celular/metabolismo , Mutación/genética , Pectinas/metabolismo , Concentración de Iones de HidrógenoRESUMEN
Polygalacturonases (PGs) fine-tune pectins to modulate cell wall chemistry and mechanics, impacting plant development. The large number of PGs encoded in plant genomes leads to questions on the diversity and specificity of distinct isozymes. Herein, we report the crystal structures of 2 Arabidopsis thaliana PGs, POLYGALACTURONASE LATERAL ROOT (PGLR), and ARABIDOPSIS DEHISCENCE ZONE POLYGALACTURONASE2 (ADPG2), which are coexpressed during root development. We first determined the amino acid variations and steric clashes that explain the absence of inhibition of the plant PGs by endogenous PG-inhibiting proteins (PGIPs). Although their beta helix folds are highly similar, PGLR and ADPG2 subsites in the substrate binding groove are occupied by divergent amino acids. By combining molecular dynamic simulations, analysis of enzyme kinetics, and hydrolysis products, we showed that these structural differences translated into distinct enzyme-substrate dynamics and enzyme processivities: ADPG2 showed greater substrate fluctuations with hydrolysis products, oligogalacturonides (OGs), with a degree of polymerization (DP) of ≤4, while the DP of OGs generated by PGLR was between 5 and 9. Using the Arabidopsis root as a developmental model, exogenous application of purified enzymes showed that the highly processive ADPG2 had major effects on both root cell elongation and cell adhesion. This work highlights the importance of PG processivity on pectin degradation regulating plant development.
Asunto(s)
Arabidopsis , Poligalacturonasa , Poligalacturonasa/genética , Poligalacturonasa/metabolismo , Arabidopsis/metabolismo , Pectinas/metabolismo , Proteínas/metabolismo , Pared Celular/metabolismoRESUMEN
Altering the content or composition of the cell wall polymer lignin is a favored approach to valorize lignin toward biomaterial and chemical production in the biorefinery. However, modifying lignin or cellulose in transgenic plants can induce expression of defense responses and negatively affect growth. Through genetic screening for suppressors of defense gene induction in the low lignin ccr1-3 mutant of Arabidopsis thaliana, we found that loss of function of the receptor-like kinase FERONIA, although not restoring growth, affected cell wall remodeling and blocked release of elicitor-active pectic polysaccharides as a result of the ccr1-3 mutation. Loss of function of multiple wall-associated kinases prevented perception of these elicitors. The elicitors are likely heterogeneous, with tri-galacturonic acid the smallest but not necessarily the most active component. Engineering of plant cell walls will require development of ways to bypass endogenous pectin signaling pathways.
Asunto(s)
Arabidopsis , Lignina , Lignina/metabolismo , Celulosa/metabolismo , Arabidopsis/genética , Polisacáridos/metabolismo , Pared Celular/genética , Regulación de la Expresión Génica de las PlantasRESUMEN
Pectins, complex polysaccharides and major components of the plant primary cell wall, can be degraded by pectate lyases (PLs). PLs cleave glycosidic bonds of homogalacturonans (HG), the main pectic domain, by ß-elimination, releasing unsaturated oligogalacturonides (OGs). To understand the catalytic mechanism and structure/function of these enzymes, we characterized VdPelB from Verticillium dahliae. We first solved the crystal structure of VdPelB at 1.2 Å resolution showing that it is a right-handed parallel ß-helix structure. Molecular dynamics (MD) simulations further highlighted the dynamics of the enzyme in complex with substrates that vary in their degree of methylesterification, identifying amino acids involved in substrate binding and cleavage of non-methylesterified pectins. We then biochemically characterized wild type and mutated forms of VdPelB. Pectate lyase VdPelB was most active on non-methylesterified pectins, at pH 8.0 in presence of Ca2+ ions. The VdPelB-G125R mutant was most active at pH 9.0 and showed higher relative activity compared to native enzyme. The OGs released by VdPelB differed to that of previously characterized PLs, showing its peculiar specificity in relation to its structure. OGs released from Verticillium-partially tolerant and sensitive flax cultivars differed which could facilitate the identification VdPelB-mediated elicitors of defence responses.
Asunto(s)
Simulación de Dinámica Molecular , Polisacárido Liasas , Polisacárido Liasas/química , Glicósidos , Pectinas/química , Especificidad por SustratoRESUMEN
Fungal pathogens grow in the apoplastic space, in constant contact with the plant cell wall (CW) that hinders microbe progression while representing a source of nutrients. Although numerous fungal CW modifying proteins have been identified, their role during host colonization remains underexplored. Here, we show that the root-infecting plant pathogen Fusarium oxysporum (Fo) does not require its complete arsenal of cellulases to infect the host plant. Quite the opposite: Fo mutants impaired in cellulose degradation become hypervirulent by enhancing the secretion of virulence factors. On the other hand, the reduction in cellulase activity had a severe negative effect on saprophytic growth and microconidia production during the final stages of the Fo infection cycle. These findings enhance our understanding of the function of plant CW degradation on the outcome of host-microbe interactions and reveal an unexpected role of cellulose degradation in a pathogen's reproductive success.
Asunto(s)
Aptitud Genética , Enfermedades de las Plantas , Celulosa , Proteínas Fúngicas , Fusarium , Enfermedades de las Plantas/microbiología , VirulenciaRESUMEN
Plant cells are encapsulated by cell walls whose properties largely determine cell growth. We have previously identified the rol1-2 mutant, which shows defects in seedling root and shoot development. rol1-2 is affected in the Rhamnose synthase 1 (RHM1) and shows alterations in the structures of Rhamnogalacturonan I (RG I) and RG II, two rhamnose-containing pectins. The data presented here shows that root tissue of the rol1-2 mutant fails to properly differentiate the cell wall in cell corners and accumulates excessive amounts of callose, both of which likely alter the physical properties of cells. A surr (suppressor of the rol1-2 root developmental defect) mutant was identified that alleviates the cell growth defects in rol1-2. The cell wall differentiation defect is re-established in the rol1-2 surr mutant and callose accumulation is reduced compared to rol1-2. The surr mutation is an allele of the cyclin-dependent kinase 8 (CDK8), which encodes a component of the mediator complex that influences processes central to plant growth and development. Together, the identification of the surr mutant suggests that changes in cell wall composition and turnover in the rol1-2 mutant have a significant impact on cell growth and reveals a function of CDK8 in cell wall architecture and composition.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Diferenciación Celular/fisiología , Quinasa 8 Dependiente de Ciclina/metabolismo , Proteínas de Arabidopsis/genética , Pared Celular/metabolismo , Quinasa 8 Dependiente de Ciclina/genética , Raíces de Plantas/genética , Ramnosa/análisis , Plantones/genéticaRESUMEN
Pectin, the major non-cellulosic component of primary cell wall can be degraded by polygalacturonases (PGs) and pectin methylesterases (PMEs) during pathogen attack on plants. We characterized two novel enzymes, VdPG2 and VdPME1, from the fungal plant pathogen Verticillium dahliae. VdPME1 was most active on citrus methylesterified pectin (55-70%) at pH 6 and a temperature of 40 °C, while VdPG2 was most active on polygalacturonic acid at pH 5 and a temperature of 50 °C. Using LC-MS/MS oligoprofiling, and various pectins, the mode of action of VdPME1 and VdPG2 were determined. VdPME1 was shown to be processive, in accordance with the electrostatic potential of the enzyme. VdPG2 was identified as endo-PG releasing both methylesterified and non-methylesterified oligogalacturonides (OGs). Additionally, when flax roots were used as substrate, acetylated OGs were detected. The comparisons of OGs released from Verticillium-susceptible and partially resistant flax cultivars identified new possible elicitor of plant defence responses.
Asunto(s)
Ascomicetos/enzimología , Hidrolasas de Éster Carboxílico/metabolismo , Proteínas Fúngicas/metabolismo , Poligalacturonasa/metabolismo , Ascomicetos/genética , Ascomicetos/patogenicidad , Hidrolasas de Éster Carboxílico/química , Hidrolasas de Éster Carboxílico/genética , Lino/metabolismo , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Cinética , Modelos Moleculares , Pectinas/metabolismo , Filogenia , Enfermedades de las Plantas/microbiología , Raíces de Plantas/metabolismo , Poligalacturonasa/química , Poligalacturonasa/genética , Electricidad Estática , Especificidad por SustratoRESUMEN
Plant cell wall remodeling plays a key role in the control of cell elongation and differentiation. In particular, fine-tuning of the degree of methylesterification of pectins was previously reported to control developmental processes as diverse as pollen germination, pollen tube elongation, emergence of primordia or elongation of dark-grown hypocotyls. However, how pectin degradation can modulate plant development has remained elusive. Here we report the characterization of a polygalacturonase (PG), AtPGLR, the gene for which is highly expressed at the onset of lateral root emergence in Arabidopsis. Due to gene compensation mechanisms, mutant approaches failed to determine the involvement of AtPGLR in plant growth. To overcome this issue, AtPGLR has been expressed heterologously in the yeast Pichia pastoris and biochemically characterized. We showed that AtPGLR is an endo-PG that preferentially releases non-methylesterified oligogalacturonides with a short degree of polymerization (< 8) at acidic pH. The application of the purified recombinant protein on Amaryllis pollen tubes, an excellent model for studying cell wall remodeling at acidic pH, induced abnormal pollen tubes or cytoplasmic leakage in the subapical dome of the pollen tube tip, where non-methylesterified pectin epitopes are detected. Those leaks could either be repaired by new ß-glucan deposits (mostly callose) in the cell wall or promoted dramatic burst of the pollen tube. Our work presents the full biochemical characterization of an Arabidopsis PG and highlights the importance of pectin integrity in pollen tube elongation.
Asunto(s)
Proteínas de Arabidopsis/fisiología , Tubo Polínico/fisiología , Poligalacturonasa/fisiología , Arabidopsis/efectos de los fármacos , Arabidopsis/enzimología , Arabidopsis/genética , Arabidopsis/fisiología , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/farmacología , Raíces de Plantas/metabolismo , Plantas Modificadas Genéticamente , Tubo Polínico/efectos de los fármacos , Poligalacturonasa/genética , Poligalacturonasa/farmacología , SaccharomycetalesRESUMEN
Ferulate (FA) units esterified to grass arabinoxylans are involved in cross-linking cell wall polymers. In this work, this contention is strengthened by the identification of FA homo- and heterodimers esterified to methyl arabinofuranoside (MeAra) units after their release from the xylan by mild acidolysis in dioxane/methanol/HCl. Acidolysis of poorly lignified maize bran cell walls provided diferulate (DFA) isomers, including those from 8-5, 8-O-4, and 5-5 interunit bonding, esterified to one or two MeAra units. Acidolysis of lignified grass samples released crossed dimers esterified to one MeAra unit and derived from the ß-O-4 coupling of coniferyl alcohol to FA esters. The evaluation of these heterodimeric esters by LC-UV of their aglycones revealed that the parent structures occur in significant amounts in lignified cell walls (0.5-1 mg/g expressed as FA equivalents). The present results position mild acidolysis as an efficient strategy to obtain improved details regarding the FA-mediated cross-linking of grass cell walls.
Asunto(s)
Arabinosa/química , Pared Celular/química , Ácidos Cumáricos/química , Poaceae/química , Ácidos/química , Dimerización , Ésteres/química , Hidrólisis , Lignina/química , Fenoles/química , Zea mays/químicaRESUMEN
Despite an ever-increasing interest for the use of pectin-derived oligogalacturonides (OGs) as biological control agents in agriculture, very little information exists-mainly for technical reasons-on the nature and activity of the OGs that accumulate during pathogen infection. Here we developed a sensitive OG profiling method, which revealed unsuspected features of the OGs generated during infection of Arabidopsis thaliana with the fungus Botrytis cinerea Indeed, in contrast to previous reports, most OGs were acetyl- and methylesterified, and 80% of them were produced by fungal pectin lyases, not by polygalacturonases. Polygalacturonase products did not accumulate as larger size OGs but were converted into oxidized GalA dimers. Finally, the comparison of the OGs and transcriptomes of leaves infected with B. cinerea mutants with reduced pectinolytic activity but with decreased or increased virulence, respectively, identified candidate OG elicitors. In conclusion, OG analysis provides insights into the enzymatic arms race between plant and pathogen and facilitates the identification of defense elicitors.
Asunto(s)
Arabidopsis/metabolismo , Botrytis/patogenicidad , Ácidos Hexurónicos/metabolismo , Proteínas de Arabidopsis/metabolismo , Botrytis/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , Pectinas/metabolismo , Enfermedades de las Plantas/microbiología , Hojas de la Planta/metabolismo , Poligalacturonasa/metabolismo , Transducción de SeñalRESUMEN
The arabinosyl side chains of grass arabinoxylans are partially acylated by p-coumarate ( pCA) and ferulate (FA). These aromatic side chains can cross-couple wall polymers resulting in modulation of cell wall physical properties. The determination of p-coumaroylated and feruloylated arabinose units has been the target of analytical efforts with trifluoroacetic acid hydrolysis the standard method to release feruloylated and p-coumaroylated arabinose units from arabinoxylans. Herein, we report on a more robust method to measure these acylated units. Acidolysis of extractive-free grass samples in a dioxane/methanol/aqueous 2 M HCl mixture provided the methyl 5- O- p-coumaroyl- and 5- O-feruloyl-l-arabinofuranoside anomers ( pCA-MeAra and FA-MeAra). These conjugates were readily analyzed by liquid chromatography combined with both UV and MS detection. The method revealed the variability of the relative acylation of arabinose units by pCA or FA in grass cell walls. This methodology will permit delineation of hydroxycinnamate acylation patterns in arabinoxylans.
Asunto(s)
Arabinosa/análisis , Arabinosa/química , Ácidos Cumáricos/química , Poaceae/química , Propionatos/química , Xilanos/química , Acilación , Pared Celular/química , Cromatografía Líquida de Alta Presión/métodos , Dioxanos , Ácido Clorhídrico/química , Lignina/aislamiento & purificación , MetanolRESUMEN
In the plant cell wall, boron links two pectic domain rhamnogalacturonan II (RG-II) chains together to form a dimer and thus contributes to the reinforcement of cell adhesion. We studied the mur1-1 mutant of Arabidopsis thaliana which has lost the ability to form GDP-fucose in the shoots and show that the extent of RG-II cross-linking is reduced in the lignified stem of this mutant. Surprisingly, MUR1 mutation induced an enrichment of resistant interunit bonds in lignin and triggered the overexpression of many genes involved in lignified tissue formation and in jasmonic acid signaling. The defect in GDP-fucose synthesis induced a loss of cell adhesion at the interface between stele and cortex, as well as between interfascicular fibers. This led to the formation of regenerative xylem, where tissue detachment occurred, and underlined a loss of resistance to mechanical forces. Similar observations were also made on bor1-3 mutant stems which are altered in boron xylem loading, leading us to suggest that diminished RG-II dimerization is responsible for regenerative xylem formation.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/metabolismo , Guanosina Difosfato Fucosa/metabolismo , Lignina/metabolismo , Mutación , Pectinas/metabolismo , Arabidopsis/genética , Pectinas/químicaRESUMEN
Plants are able to generate large leaf surfaces that act as two-dimensional solar panels with a minimum investment in building material, thanks to a hydrostatic skeleton. This requires high intracellular pressures (up to 1 MPa), which depend on the presence of strong cell walls. The walls of growing cells (also called primary walls), are remarkably able to reconcile extreme tensile strength (up to 100 MPa) with the extensibility necessary for growth. All walled organisms are confronted with this dilemma - the need to balance strength and extensibility - and bacteria, fungi and plants have evolved independent solutions to cope. In this Primer, we discuss how plant cells have solved this problem, allowing them to support often very large increases in volume and to develop a broad variety of shapes (Figure 1A,B,D). This shape variation reflects the targeted deposition of wall material combined with local variations in cell-wall extensibility, processes that remain incompletely understood. Once the cell has reached its final size, it can lay down secondary wall layers, the composition and architecture of which are optimized to exert specific functions in different cell types (Figure 1E-G). Such functions include: providing mechanical support, for instance, for fibre cells in tree trunks or grass internodes; impermeabilising and strengthening vascular tissue to resist the negative pressure of the transpiration stream; increasing the surface area of the plasma membrane to facilitate solute exchange between cells (Figure 1C); or allowing important elastic deformation, for instance, to support the opening and closing of stomates. Specialized secondary walls, such as those constituting seed mucilage, are stored in a dehydrated form in seedcoat epidermis cells and show rapid swelling upon hydration of the seed. Other walls, in particular in reserve tissues, can accommodate large amounts of storage polysaccharides, which can be easily mobilized as a carbon source. Here we will discuss some general principles underlying wall architecture and wall growth that have emerged from recent studies, as well as future questions for investigation (Box 1).
Asunto(s)
Pared Celular/fisiología , Células Vegetales/metabolismo , Plantas/química , Plantas/metabolismo , Sustancias Macromoleculares/metabolismo , Células Vegetales/química , Polisacáridos/metabolismoRESUMEN
Plants, like yeast, have the ability to monitor alterations in the cell wall architecture that occur during normal growth or in changing environments and to trigger compensatory changes in the cell wall. We discuss how recent advances in our understanding of the cell wall architecture provide new insights into the role of cell wall integrity sensing in growth control. Next we review the properties of membrane receptor-like kinases that have roles in pH control, mechano-sensing and reactive oxygen species accumulation in growing cells and which may be the plant equivalents of the yeast cell wall integrity (CWI) sensors. Finally, we discuss recent findings showing an increasing role for CWI signaling in plant immunity and the adaptation to changes in the ionic environment of plant cells.
Asunto(s)
Pared Celular/genética , Células Vegetales/metabolismo , Plantas/genética , Pared Celular/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Transducción de Señal/genéticaRESUMEN
Lignins are cell wall phenolic polymers resulting from monolignol radical coupling. They have characteristically high diversity in their structures which is a direct consequence of the versatile character of the lignification mechanisms discussed in this review. We will relate the latest discoveries regarding the main participants involved in lignin deposition in various tissues. Lignification is often described as a cell autonomous event occurring progressively in all cell wall layers during lignifying cell life and stopping with the cell death. However, recent data combined to old data from studies of tree lignification and zinnia cultures challenged these entrenched views and showed that the lignification process is cell-type dependent and can involve neighboring cells. Therefore, we consider recent data on cell-autonomous and non-cell autonomous lignification processes. We conclude that the role of lignins still need to be assessed during plant development and that control of polymerization/lignin deposition remains elusive and need to be investigated.
Asunto(s)
Lignina/metabolismo , Polimerizacion , Regulación de la Expresión Génica de las Plantas , Modelos Biológicos , Células Vegetales/metabolismoRESUMEN
Polysaccharides were extracted from seven plants endemic to Gabon to study their potential immunological activities. Peripheral blood mononuclear cell (PBMC) (5×105 cells/mL) proliferation, cytokine and immunoglobulin G (IgG) assays were performed after stimulation with different concentrations of polysaccharide fractions compared with lipopolysaccharides (LPS) and concanavalin A (ConA) from healthy volunteers. The culture supernatants were used for cytokine and IgG detection by enzyme-linked immunosorbent assay (ELISA). The results show that pectin and hemicellulose extracts from Uvaria klainei, Petersianthus macrocarpus, Trichoscypha addonii, Aphanocalyx microphyllus, Librevillea klaineana, Neochevalierodendron stephanii and Scorodophloeus zenkeri induced production levels that were variable from one individual to another for IL-12 (3-40 pg/mL), IL-10 (6-443 pg/mL), IL-6 (7-370 pg/mL), GM-CSF (3-170 pg/mL) and IFN-γ (5-80 pg/mL). Only hemicelluloses from Aphanocalyx microphyllus produce a small amount of IgG (OD=0.034), while the proliferation of cells stimulated with these polysaccharides increased up to 318% above the proliferation of unstimulated cells. However, this proliferation of PBMCs was abolished when the pectin of some of these plants was treated with endopolygalacturonase (p<0.05), but the trend of cytokine synthesis remained the same, both before and after enzymatic treatment or saponification. This study suggests that these polysaccharides stimulate cells in a structure-dependent manner. The rhamnogalacturonan-I (RGI) fragment alone was not able to induce the proliferation of PBMC.
Asunto(s)
Proliferación Celular/efectos de los fármacos , Citocinas/biosíntesis , Inmunoglobulina G/biosíntesis , Leucocitos Mononucleares/metabolismo , Extractos Vegetales/química , Polisacáridos , Femenino , Gabón , Humanos , Leucocitos Mononucleares/citología , Masculino , Polisacáridos/química , Polisacáridos/aislamiento & purificación , Polisacáridos/farmacologíaRESUMEN
The scientific presentations at the First International Brachypodium Conference (abstracts available at http://www.brachy2013.unimore.it) are evidence of the widespread adoption of Brachypodium distachyon as a model system. Furthermore, the wide range of topics presented (genome evolution, roots, abiotic and biotic stress, comparative genomics, natural diversity, and cell walls) demonstrates that the Brachypodium research community has achieved a critical mass of tools and has transitioned from resource development to addressing biological questions, particularly those unique to grasses.
Asunto(s)
Brachypodium/fisiología , Genoma de Planta/genética , Genómica , Enfermedades de las Plantas , Biomasa , Brachypodium/genética , Brachypodium/crecimiento & desarrollo , Pared Celular/metabolismo , Evolución Molecular , Modelos Biológicos , Poaceae/genética , Poaceae/crecimiento & desarrollo , Poaceae/fisiología , Estrés FisiológicoRESUMEN
BACKGROUND AND AIMS: Rhamnogalacturonan-II (RG-II) is one of the pectin motifs found in the cell wall of all land plants. It contains sugars such as 2-keto-3-deoxy-d-lyxo-heptulosaric acid (Dha) and 2-keto-3-deoxy-d-manno-octulosonic acid (Kdo), and within the wall RG-II is mostly found as a dimer via a borate diester cross-link. To date, little is known regarding the biosynthesis of this motif. Here, after a brief review of our current knowledge on RG-II structure, biosynthesis and function in plants, this study explores the implications of the presence of a Golgi-localized sialyltransferase-like 2 (SIA2) protein that is possibly involved in the transfer of Dha or Kdo in the RG-II of Arabidopsis thaliana pollen tubes, a fast-growing cell type used as a model for the study of cell elongation. METHODS: Two heterozygous mutant lines of arabidopsis (sia2-1+/- and qrt1 × sia2-2+/-) were investigated. sia2-2+/- was in a quartet1 background and the inserted T-DNA contained the reporter gene ß-glucuronidase (GUS) under the pollen-specific promoter LAT52. Pollen germination and pollen tube phenotype and growth were analysed both in vitro and in vivo by microscopy. KEY RESULTS: Self-pollination of heterozygous lines produced no homozygous plants in the progeny, which may suggest that the mutation could be lethal. Heterozygous mutants displayed a much lower germination rate overall and exhibited a substantial delay in germination (20 h of delay to reach 30 % of pollen grain germination compared with the wild type). In both lines, mutant pollen grains that were able to produce a tube had tubes that were either bursting, abnormal (swollen or dichotomous branching tip) or much shorter compared with wild-type pollen tubes. In vivo, mutant pollen tubes were restricted to the style, whereas the wild-type pollen tubes were detected at the base of the ovary. CONCLUSIONS: This study highlights that the mutation in arabidopsis SIA2 encoding a sialyltransferase-like protein that may transfer Dha or Kdo on the RG-II motif has a dramatic effect on the stability of the pollen tube cell wall.