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1.
Plant Physiol ; 196(1): 124-136, 2024 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-38833284

RESUMO

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.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Celulose , Glucosiltransferases , Microtúbulos , Glucosiltransferases/metabolismo , Glucosiltransferases/genética , Microtúbulos/metabolismo , Microtúbulos/efeitos dos fármacos , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Celulose/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Mutação , Plântula/genética , Plântula/crescimento & desenvolvimento , Plântula/efeitos dos fármacos , Membrana Celular/metabolismo , Piridazinas/farmacologia , Parede Celular/metabolismo , Parede Celular/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Benzamidas
2.
Development ; 148(21)2021 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-34739031

RESUMO

Plant brassinosteroid hormones (BRs) regulate growth in part through altering the properties of the cell wall, the extracellular matrix of plant cells. Conversely, feedback signalling from the wall connects the state of cell wall homeostasis to the BR receptor complex and modulates BR activity. Here, we report that both pectin-triggered cell wall signalling and impaired BR signalling result in altered cell wall orientation in the Arabidopsis root meristem. Furthermore, both depletion of endogenous BRs and exogenous supply of BRs triggered these defects. Cell wall signalling-induced alterations in the orientation of newly placed walls appear to occur late during cytokinesis, after initial positioning of the cortical division zone. Tissue-specific perturbations of BR signalling revealed that the cellular malfunction is unrelated to previously described whole organ growth defects. Thus, tissue type separates the pleiotropic effects of cell wall/BR signals and highlights their importance during cell wall placement.


Assuntos
Arabidopsis/metabolismo , Brassinosteroides/metabolismo , Parede Celular/metabolismo , Meristema/metabolismo , Transdução de Sinais , Arabidopsis/citologia , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/metabolismo , Divisão Celular , Citocinese , Homeostase , Meristema/citologia , Pectinas/metabolismo , Raízes de Plantas/citologia , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo
3.
Proc Natl Acad Sci U S A ; 116(39): 19743-19752, 2019 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-31501325

RESUMO

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.


Assuntos
Arabidopsis/metabolismo , Botrytis/patogenicidade , Ácidos Hexurônicos/metabolismo , Proteínas de Arabidopsis/metabolismo , Botrytis/metabolismo , Regulação da Expressão Gênica de Plantas/genética , Pectinas/metabolismo , Doenças das Plantas/microbiologia , Folhas de Planta/metabolismo , Poligalacturonase/metabolismo , Transdução de Sinais
4.
Plant Physiol ; 182(1): 669-678, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31641077

RESUMO

Plants depend on various cell surface receptors to integrate extracellular signals with developmental programs. One of the best-studied receptors is BRASSINOSTEROID INSENSITIVE 1 (BRI1) in Arabidopsis (Arabidopsis thaliana). Upon binding of its hormone ligands, BRI1 forms a complex with a shape-complementary coreceptor and initiates a signal transduction cascade, which leads to a variety of responses. At the macroscopic level, brassinosteroid (BR) biosynthetic and receptor mutants have similar growth defects, which initially led to the assumption that the signaling pathways were largely linear. However, recent evidence suggests that BR signaling is interconnected with several other pathways through various mechanisms. We recently described that feedback from the cell wall is integrated at the level of the receptor complex through interaction with RECEPTOR-LIKE PROTEIN 44 (RLP44). Moreover, BRI1 is required for another function of RLP44: the control of procambial cell fate. Here, we report a BRI1 mutant, bri1 cnu4 , which differentially affects canonical BR signaling and RLP44 function in the vasculature. Although BR signaling is only mildly impaired, bri1 cnu4 mutants show ectopic xylem in place of procambium. Mechanistically, this is explained by an increased association between RLP44 and the mutated BRI1 protein, which prevents the former from acting in vascular cell fate maintenance. Consistent with this, the mild BR response phenotype of bri1 cnu4 is a recessive trait, whereas the RLP44-mediated xylem phenotype is semidominant. Our results highlight the complexity of plant plasma membrane receptor function and provide a tool to dissect BR signaling-related roles of BRI1 from its noncanonical functions.


Assuntos
Proteínas de Arabidopsis/metabolismo , Brassinosteroides/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Proteínas Quinases/metabolismo , Alelos , Proteínas de Arabidopsis/genética , Plantas Geneticamente Modificadas/genética , Proteínas Quinases/genética , Transdução de Sinais
5.
Plant Cell ; 30(10): 2330-2351, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-30115738

RESUMO

Somatic polyploidy caused by endoreplication is observed in arthropods, molluscs, and vertebrates but is especially prominent in higher plants, where it has been postulated to be essential for cell growth and fate maintenance. However, a comprehensive understanding of the physiological significance of plant endopolyploidy has remained elusive. Here, we modeled and experimentally verified a high-resolution DNA endoploidy map of the developing Arabidopsis thaliana root, revealing a remarkable spatiotemporal control of DNA endoploidy levels across tissues. Fitting of a simplified model to publicly available data sets profiling root gene expression under various environmental stress conditions suggested that this root endoploidy patterning may be stress-responsive. Furthermore, cellular and transcriptomic analyses revealed that inhibition of endoreplication onset alters the nuclear-to-cellular volume ratio and the expression of cell wall-modifying genes, in correlation with the appearance of cell structural changes. Our data indicate that endopolyploidy might serve to coordinate cell expansion with structural stability and that spatiotemporal endoreplication pattern changes may buffer for stress conditions, which may explain the widespread occurrence of the endocycle in plant species growing in extreme or variable environments.


Assuntos
Adaptação Fisiológica/genética , Arabidopsis/fisiologia , Raízes de Plantas/genética , Poliploidia , Arabidopsis/citologia , Arabidopsis/genética , Tamanho Celular , DNA de Plantas , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Células Vegetais/fisiologia , Raízes de Plantas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas , Reprodutibilidade dos Testes , Análise Espaço-Temporal , Estresse Fisiológico/genética
6.
Plant Cell Physiol ; 61(6): 1191-1203, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32333782

RESUMO

Growth of etiolated Arabidopsis hypocotyls is biphasic. During the first phase, cells elongate slowly and synchronously. At 48 h after imbibition, cells at the hypocotyl base accelerate their growth. Subsequently, this rapid elongation propagates through the hypocotyl from base to top. It is largely unclear what regulates the switch from slow to fast elongation. Reverse genetics-based screening for hypocotyl phenotypes identified three independent mutant lines of At1g70990, a short extensin (EXT) family protein that we named EXT33, with shorter etiolated hypocotyls during the slow elongation phase. However, at 72 h after imbibition, these dark-grown mutant hypocotyls start to elongate faster than the wild type (WT). As a result, fully mature 8-day-old dark-grown hypocotyls were significantly longer than WTs. Mutant roots showed no growth phenotype. In line with these results, analysis of native promoter-driven transcriptional fusion lines revealed that, in dark-grown hypocotyls, expression occurred in the epidermis and cortex and that it was strongest in the growing part. Confocal and spinning disk microscopy on C-terminal protein-GFP fusion lines localized the EXT33-protein to the ER and cell wall. Fourier-transform infrared microspectroscopy identified subtle changes in cell wall composition between WT and the mutant, reflecting altered cell wall biomechanics measured by constant load extensometry. Our results indicate that the EXT33 short EXT family protein is required during the first phase of dark-grown hypocotyl elongation and that it regulates the moment and extent of the growth acceleration by modulating cell wall extensibility.


Assuntos
Proteínas de Arabidopsis/fisiologia , Arabidopsis/crescimento & desenvolvimento , Hipocótilo/crescimento & desenvolvimento , Proteínas de Membrana/fisiologia , Alelos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Parede Celular/metabolismo , Cotilédone/metabolismo , Estiolamento , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas/genética , Hipocótilo/metabolismo , Proteínas de Membrana/genética , Filogenia , Raízes de Plantas/metabolismo , Alinhamento de Sequência , Espectroscopia de Infravermelho com Transformada de Fourier
7.
PLoS Genet ; 13(6): e1006832, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28604776

RESUMO

Plants actively perceive and respond to perturbations in their cell walls which arise during growth, biotic and abiotic stresses. However, few components involved in plant cell wall integrity sensing have been described to date. Using a reverse-genetic approach, we identified the Arabidopsis thaliana leucine-rich repeat receptor kinase MIK2 as an important regulator of cell wall damage responses triggered upon cellulose biosynthesis inhibition. Indeed, loss-of-function mik2 alleles are strongly affected in immune marker gene expression, jasmonic acid production and lignin deposition. MIK2 has both overlapping and distinct functions with THE1, a malectin-like receptor kinase previously proposed as cell wall integrity sensor. In addition, mik2 mutant plants exhibit enhanced leftward root skewing when grown on vertical plates. Notably, natural variation in MIK2 (also named LRR-KISS) has been correlated recently to mild salt stress tolerance, which we could confirm using our insertional alleles. Strikingly, both the increased root skewing and salt stress sensitivity phenotypes observed in the mik2 mutant are dependent on THE1. Finally, we found that MIK2 is required for resistance to the fungal root pathogen Fusarium oxysporum. Together, our data identify MIK2 as a novel component in cell wall integrity sensing and suggest that MIK2 is a nexus linking cell wall integrity sensing to growth and environmental cues.


Assuntos
Proteínas de Arabidopsis/genética , Parede Celular/genética , Raízes de Plantas/genética , Proteínas Quinases/genética , Receptores de Superfície Celular/genética , Estresse Fisiológico/genética , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/biossíntese , Parede Celular/efeitos dos fármacos , Celulose/biossíntese , Ciclopentanos/metabolismo , Resistência à Doença/genética , Fusarium/patogenicidade , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Lignina/biossíntese , Oxilipinas/metabolismo , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Raízes de Plantas/efeitos dos fármacos , Proteínas Quinases/biossíntese , Cloreto de Sódio/toxicidade , Estresse Fisiológico/efeitos dos fármacos
8.
Plant Cell ; 28(9): 2276-2290, 2016 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-27543091

RESUMO

Because the plant cell wall provides the first line of defense against biotic and abiotic assaults, its functional integrity needs to be maintained under stress conditions. Through a phenotype-based compound screening approach, we identified a novel cellulose synthase inhibitor, designated C17. C17 administration depletes cellulose synthase complexes from the plasma membrane in Arabidopsis thaliana, resulting in anisotropic cell elongation and a weak cell wall. Surprisingly, in addition to mutations in CELLULOSE SYNTHASE1 (CESA1) and CESA3, a forward genetic screen identified two independent defective genes encoding pentatricopeptide repeat (PPR)-like proteins (CELL WALL MAINTAINER1 [CWM1] and CWM2) as conferring tolerance to C17. Functional analysis revealed that mutations in these PPR proteins resulted in defective cytochrome c maturation and activation of mitochondrial retrograde signaling, as evidenced by the induction of an alternative oxidase. These mitochondrial perturbations increased tolerance to cell wall damage induced by cellulose deficiency. Likewise, administration of antimycin A, an inhibitor of mitochondrial complex III, resulted in tolerance toward C17. The C17 tolerance of cwm2 was partially lost upon depletion of the mitochondrial retrograde regulator ANAC017, demonstrating that ANAC017 links mitochondrial dysfunction with the cell wall. In view of mitochondria being a major target of a variety of stresses, our data indicate that plant cells might modulate mitochondrial activity to maintain a functional cell wall when subjected to stresses.

10.
Plant J ; 87(2): 230-42, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27121260

RESUMO

The localization of proteins in specific domains or compartments in the 3D cellular space is essential for many fundamental processes in eukaryotic cells. Deciphering spatial organization principles within cells is a challenging task, in particular because of the large morphological variations between individual cells. We present here an approach for normalizing variations in cell morphology and for statistically analyzing spatial distributions of intracellular compartments from collections of 3D images. The method relies on the processing and analysis of 3D geometrical models that are generated from image stacks and that are used to build representations at progressively increasing levels of integration, ultimately revealing statistical significant traits of spatial distributions. To make this methodology widely available to end-users, we implemented our algorithmic pipeline into a user-friendly, multi-platform, and freely available software. To validate our approach, we generated 3D statistical maps of endomembrane compartments at subcellular resolution within an average epidermal root cell from collections of image stacks. This revealed unsuspected polar distribution patterns of organelles that were not detectable in individual images. By reversing the classical 'measure-then-average' paradigm, one major benefit of the proposed strategy is the production and display of statistical 3D representations of spatial organizations, thus fully preserving the spatial dimension of image data and at the same time allowing their integration over individual observations. The approach and software are generic and should be of general interest for experimental and modeling studies of spatial organizations at multiple scales (subcellular, cellular, tissular) in biological systems.


Assuntos
Células/ultraestrutura , Imageamento Tridimensional/métodos , Arabidopsis/ultraestrutura , Proteínas de Fluorescência Verde/metabolismo , Software , Análise Espacial , Frações Subcelulares/ultraestrutura
11.
Plant J ; 88(3): 468-475, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27354251

RESUMO

A major challenge in plant systems biology is the development of robust, predictive multiscale models for organ growth. In this context it is important to bridge the gap between the, rather well-documented molecular scale and the organ scale by providing quantitative methods to study within-organ growth patterns. Here, we describe a simple method for the analysis of the evolution of growth patterns within rod-shaped organs that does not require adding markers at the organ surface. The method allows for the simultaneous analysis of root and hypocotyl growth, provides spatio-temporal information on curvature, growth anisotropy and relative elemental growth rate and can cope with complex organ movements. We demonstrate the performance of the method by documenting previously unsuspected complex growth patterns within the growing hypocotyl of the model species Arabidopsis thaliana during normal growth, after treatment with a growth-inhibiting drug or in a mechano-sensing mutant. The method is freely available as an intuitive and user-friendly Matlab application called KymoRod.


Assuntos
Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Hipocótilo/genética , Hipocótilo/metabolismo , Raízes de Plantas/genética , Raízes de Plantas/metabolismo
12.
Development ; 141(24): 4841-50, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25395456

RESUMO

Polysaccharides are major components of extracellular matrices and are often extensively modified post-synthetically to suit local requirements and developmental programmes. However, our current understanding of the spatiotemporal dynamics and functional significance of these modifications is limited by a lack of suitable molecular tools. Here, we report the development of a novel non-immunological approach for producing highly selective reciprocal oligosaccharide-based probes for chitosan (the product of chitin deacetylation) and for demethylesterified homogalacturonan. Specific reciprocal binding is mediated by the unique stereochemical arrangement of oppositely charged amino and carboxy groups. Conjugation of oligosaccharides to fluorophores or gold nanoparticles enables direct and rapid imaging of homogalacturonan and chitosan with unprecedented precision in diverse plant, fungal and animal systems. We demonstrated their potential for providing new biological insights by using them to study homogalacturonan processing during Arabidopsis thaliana root cap development and by analyzing sites of chitosan deposition in fungal cell walls and arthropod exoskeletons.


Assuntos
Quitina/metabolismo , Matriz Extracelular/metabolismo , Sondas Moleculares , Oligossacarídeos , Pectinas/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Parede Celular/ultraestrutura , Quitina/isolamento & purificação , Desmidiales/ultraestrutura , Nanopartículas Metálicas , Análise em Microsséries , Microscopia Eletrônica de Transmissão , Sondas Moleculares/metabolismo , Estrutura Molecular , Oligossacarídeos/química , Oligossacarídeos/metabolismo , Imagem Óptica/métodos , Pectinas/isolamento & purificação , Coifa/crescimento & desenvolvimento , Coifa/metabolismo
13.
New Phytol ; 215(3): 1009-1025, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28617955

RESUMO

While Brachypodium distachyon (Brachypodium) is an emerging model for grasses, no expression atlas or gene coexpression network is available. Such tools are of high importance to provide insights into the function of Brachypodium genes. We present a detailed Brachypodium expression atlas, capturing gene expression in its major organs at different developmental stages. The data were integrated into a large-scale coexpression database ( www.gene2function.de), enabling identification of duplicated pathways and conserved processes across 10 plant species, thus allowing genome-wide inference of gene function. We highlight the importance of the atlas and the platform through the identification of duplicated cell wall modules, and show that a lignin biosynthesis module is conserved across angiosperms. We identified and functionally characterised a putative ferulate 5-hydroxylase gene through overexpression of it in Brachypodium, which resulted in an increase in lignin syringyl units and reduced lignin content of mature stems, and led to improved saccharification of the stem biomass. Our Brachypodium expression atlas thus provides a powerful resource to reveal functionally related genes, which may advance our understanding of important biological processes in grasses.


Assuntos
Brachypodium/citologia , Brachypodium/genética , Parede Celular/genética , Regulação da Expressão Gênica de Plantas , Redes Reguladoras de Genes , Genes de Plantas , Lignina/metabolismo , Arabidopsis/genética , Bases de Dados Genéticas , Oryza/genética , Caules de Planta/metabolismo , Plantas Geneticamente Modificadas , Transcriptoma/genética
14.
J Exp Bot ; 68(16): 4583-4593, 2017 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-28981771

RESUMO

Perturbation of cellulose synthesis in plants triggers stress responses, including growth retardation, mediated by the cell wall integrity-sensing receptor-like kinase (RLK) THESEUS1 (THE1). The analysis of two alleles carrying T-DNA insertions at comparable positions has led to conflicting conclusions concerning the impact of THE1 signaling on growth. Here we confirm that, unlike the1-3 and other the1 alleles in which cellular responses to genetic or pharmacological inhibition of cellulose synthesis are attenuated, the1-4 showed enhanced responses, including growth inhibition, ectopic lignification, and stress gene expression. Both the1-3 and the1-4 express a transcript encoding a predicted membrane-associated truncated protein lacking the kinase domain. However, the1-3, in contrast to the1-4, strongly expresses antisense transcripts, which are expected to prevent the expression of the truncated protein as suggested by the genetic interactions between the two alleles. Seedlings overexpressing such a truncated protein react to isoxaben treatment similarly to the1-4 and the full-length THE overexpressor. We conclude that the1-4 is a hypermorphic allele; that THE1 signaling upon cell wall damage has a negative impact on cell expansion; and that caution is required when interpreting the phenotypic effects of T-DNA insertions in RLK genes.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/citologia , Parede Celular/metabolismo , Proteínas Quinases/genética , Receptores de Superfície Celular/genética , Alelos , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Benzamidas/farmacologia , Parede Celular/genética , Celulose/biossíntese , DNA Bacteriano , Regulação da Expressão Gênica de Plantas , Genes Dominantes , Lignina/metabolismo , Plantas Geneticamente Modificadas , Proteínas Quinases/metabolismo , Receptores de Superfície Celular/metabolismo , Plântula/efeitos dos fármacos , Plântula/genética , Plântula/metabolismo , Transdução de Sinais
15.
Plant Cell ; 26(5): 1848-1856, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24808052

RESUMO

Despite an increasingly detailed understanding of endogenous and environmental growth-controlling signals and their signaling networks, little is known on how these networks are integrated with the cell expansion machinery. Members of the CrRLK1L family control cell wall properties and cell expansion in a variety of developmental and environmental contexts. Two recent reports provide exciting new insights into the mode of action of these RLKs. One study shows that one family member, FERONIA (FER), is required for the production of hydroxyl radicals in the female gametophyte, which causes pollen tube rupture and sperm cell release during fertilization. Another study shows that FER is a receptor for a signaling peptide (Rapid Alkalinization Factor 1 [RALF1]) that triggers cell wall alkalinization and growth arrest, possibly through the inhibition of plasma membrane H+-ATPase activity. RALF1 belongs to a large gene family, with a wide range of expression patterns. Other CrRLK1L family members therefore may also be receptors for RALF-like peptides. These findings have important implications for our understanding of the control of cell wall integrity during growth and raise new intriguing questions.

16.
Proc Natl Acad Sci U S A ; 111(40): 14601-6, 2014 Oct 07.
Artigo em Inglês | MEDLINE | ID: mdl-25246540

RESUMO

Lignocellulosic plant biomass is an attractive feedstock for the production of sustainable biofuels, but the commercialization of such products is hampered by the high costs of processing this material into fermentable sugars (saccharification). One approach to lowering these costs is to produce crops with cell walls that are more susceptible to hydrolysis to reduce preprocessing and enzyme inputs. To deepen our understanding of the molecular genetic basis of lignocellulose recalcitrance, we have screened a mutagenized population of the model grass Brachypodium distachyon for improved saccharification with an industrial polysaccharide-degrading enzyme mixture. From an initial screen of 2,400 M2 plants, we selected 12 lines that showed heritable improvements in saccharification, mostly with no significant reduction in plant size or stem strength. Characterization of these putative mutants revealed a variety of alterations in cell-wall components. We have mapped the underlying genetic lesions responsible for increased saccharification using a deep sequencing approach, and here we report the mapping of one of the causal mutations to a narrow region in chromosome 2. The most likely candidate gene in this region encodes a GT61 glycosyltransferase, which has been implicated in arabinoxylan substitution. Our work shows that forward genetic screening provides a powerful route to identify factors that impact on lignocellulose digestibility, with implications for improving feedstock for cellulosic biofuel production.


Assuntos
Brachypodium/genética , Brachypodium/metabolismo , Metabolismo dos Carboidratos , Parede Celular/metabolismo , Mutação , Biocombustíveis , Biomassa , Brachypodium/crescimento & desenvolvimento , Celulose/metabolismo , Mapeamento Cromossômico , Cromossomos de Plantas/genética , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Lignina/metabolismo , Monossacarídeos/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Caules de Planta/genética , Caules de Planta/crescimento & desenvolvimento , Caules de Planta/metabolismo , Polissacarídeos/metabolismo , Análise de Componente Principal , Espectroscopia de Infravermelho com Transformada de Fourier
17.
Proc Natl Acad Sci U S A ; 111(42): 15261-6, 2014 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-25288746

RESUMO

The brassinosteroid (BR) signaling module is a central regulator of plant morphogenesis, as indicated by the large number of BR-responsive cell wall-related genes and the severe growth defects of BR mutants. Despite a detailed knowledge of the signaling components, the logic of this auto-/paracrine signaling module in growth control remains poorly understood. Recently, extensive cross-talk with other signaling pathways has been shown, suggesting that the outputs of BR signaling, such as gene-expression changes, are subject to complex control mechanisms. We previously provided evidence for a role of BR signaling in a feedback loop controlling the integrity of the cell wall. Here, we identify the first dedicated component of this feedback loop: a receptor-like protein (RLP44), which is essential for the compensatory triggering of BR signaling upon inhibition of pectin de-methylesterification in the cell wall. RLP44 is required for normal growth and stress responses and connects with the BR signaling pathway, presumably through a direct interaction with the regulatory receptor-like kinase BAK1. These findings corroborate a role for BR in controlling the sensitivity of a feedback signaling module involved in maintaining the physico-chemical homeostasis of the cell wall during cell expansion.


Assuntos
Brassinosteroides/química , Pectinas/química , Proteínas de Plantas/fisiologia , Proteínas de Arabidopsis/fisiologia , Parede Celular/metabolismo , Clonagem Molecular , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Homeostase , Ligantes , Microscopia Confocal , Mutação , Fenótipo , Ligação Proteica , Mapeamento de Interação de Proteínas , Proteínas Serina-Treonina Quinases/fisiologia , Transdução de Sinais
18.
Glycobiology ; 26(9): 950-960, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-26945038

RESUMO

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.


Assuntos
Parede Celular/genética , Células Vegetais/metabolismo , Plantas/genética , Parede Celular/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Transdução de Sinais/genética
19.
Plant J ; 77(1): 71-84, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24147885

RESUMO

During cytokinesis a new crosswall is rapidly laid down. This process involves the formation at the cell equator of a tubulo-vesicular membrane network (TVN). This TVN evolves into a tubular network (TN) and a planar fenestrated sheet, which extends at its periphery before fusing to the mother cell wall. The role of cell wall polymers in cell plate assembly is poorly understood. We used specific stains and GFP-labelled cellulose synthases (CESAs) to show that cellulose, as well as three distinct CESAs, accumulated in the cell plate already at the TVN stage. This early presence suggests that cellulose is extruded into the tubular membrane structures of the TVN. Co-localisation studies using GFP-CESAs suggest the delivery of cellulose synthase complexes (CSCs) to the cell plate via phragmoplast-associated vesicles. In the more mature TN part of the cell plate, we observed delivery of GFP-CESA from doughnut-shaped organelles, presumably Golgi bodies. During the conversion of the TN into a planar fenestrated sheet, the GFP-CESA density diminished, whereas GFP-CESA levels remained high in the TVN zone at the periphery of the expanding cell plate. We observed retrieval of GFP-CESA in clathrin-containing structures from the central zone of the cell plate and from the plasma membrane of the mother cell, which may contribute to the recycling of CESAs to the peripheral growth zone of the cell plate. These observations, together with mutant phenotypes of cellulose-deficient mutants and pharmacological experiments, suggest a key role for cellulose synthesis already at early stages of cell plate assembly.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/metabolismo , Parede Celular/metabolismo , Celulose/metabolismo , Glucosiltransferases/genética , Arabidopsis/citologia , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Divisão Celular , Membrana Celular/metabolismo , Membrana Celular/ultraestrutura , Parede Celular/ultraestrutura , Clatrina/metabolismo , Citocinese , Genes Reporter , Glucosiltransferases/metabolismo , Complexo de Golgi/metabolismo , Complexo de Golgi/ultraestrutura , Isoenzimas , Microscopia Confocal , Microtúbulos/ultraestrutura , Modelos Biológicos , Raízes de Plantas/citologia , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Proteínas Recombinantes de Fusão , Plântula/citologia , Plântula/genética , Plântula/metabolismo
20.
Plant Cell Physiol ; 56(2): 224-31, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25481004

RESUMO

Understanding how developmental and environmental signals control plant cell expansion requires an intimate knowledge of the architecture of the primary cell wall and the chemo-rheological processes that underlie cell wall relaxation. In this review I discuss recent findings that reveal a more prominent role than previously suspected for covalent bonds and pectin cross-links in primary cell wall architecture. In addition, genetic studies have uncovered a role for receptor kinases in the control of cell wall homeostasis in growing cells. The emerging view is that, upon cell wall disruption, compensatory changes are induced in the cell wall through the interplay between the brassinosteroid signaling module, which positively regulates wall extensibility and receptor kinases of the CrRLKL1 family, which may act as negative regulators of cell wall stiffness. These findings lift the tip of the veil of a complex signaling network allowing normal homeostasis in walls of growing cells but also crisis management under stress conditions.


Assuntos
Brassinosteroides/metabolismo , Parede Celular/metabolismo , Proteínas de Plantas/metabolismo , Transdução de Sinais , Modelos Biológicos , Desenvolvimento Vegetal
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