RESUMO
Both filamentous pathogens' hyphae and pollen tube penetrate the host's outer layer and involve growth within the host tissues. Early epidermal responses are decisive for the outcome of these two-cell interaction processes. We identified a single cell type, the papilla of Arabidospis thaliana's stigma, as a tool to conduct a comprehensive comparative analysis on how an epidermal cell responds to the invasion of an unwanted pathogen or a welcomed pollen tube. We showed that Phytophtora parasitica, a root oomycete, effectively breaches the stigmatic cell wall and develops as a biotroph within the papilla cytoplasm. These invasive features resemble the behaviour exhibited by the pathogen within its natural host cells, but diverge from the manner in which the pollen tube progresses, being engulfed within the papilla cell wall. Quantitative analysis revealed that both invaders trigger reorganisation of the stigmatic endomembrane system and the actin cytoskeleton. While some remodelling processes are shared between the two interactions, others appear more specific towards the respective invader. These findings underscore the remarkable ability of an epidermal cell to differentiate between two types of invaders, thereby enabling it to trigger the most suitable response during the onset of invasion.
RESUMO
In plants, the first interaction that occurs between the male gametophytes (pollen grains) and the stigmatic epidermis of the female organ is crucial for successful reproduction. The stigma consists of a dome of flask-shaped cells specialized in pollen capture. In these stigmatic cells, the cytoskeleton network (cortical microtubules and actin microfilaments) actively responds to pollen contact and undergoes dynamic remodeling required for successful pollen acceptance to occur. Here, we have designed several microscopy mountings to monitor stigmatic cytoskeleton dynamics. These designs are based on the constraints linked to the tightly regulated pollen-stigma interaction and depend upon the experimental goal, either a static view or live-cell imaging.
Assuntos
Arabidopsis , Polinização , Tubo Polínico , Pólen , Citoesqueleto de Actina , FloresRESUMO
Following pollen deposition on the receptive surface of the stigma, pollen germinates a tube that carries male gametes toward the ovule where fertilization occurs. As soon as it emerges from the pollen grain, the pollen tube has to be properly guided through the pistil tissues so as to reach the ovule and ensure double fertilization. Chemical attractants, nutrients as well as receptor kinase-dependent signaling pathways have been implicated in this guidance. Recently, we showed in Arabidopsis that the microtubule severing enzyme KATANIN, by acting both on cortical microtubule (CMT) dynamics and cellulose microfibril (CMF) deposition, conferred particular mechanical properties to the papilla cell wall that act as active guidance factors. Here we confirm the importance of KATANIN and CMT orientation in pollen tube directionality by examining another katanin mutant.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Parede Celular , Katanina/metabolismo , Microtúbulos , Tubo Polínico , Polinização , Arabidopsis/fisiologia , Celulose , Fertilização , Flores , Óvulo Vegetal , Pólen , Tubo Polínico/crescimento & desenvolvimento , Tubo Polínico/metabolismoRESUMO
BACKGROUND: Fertilization in flowering plants depends on the early contact and acceptance of pollen grains by the receptive papilla cells of the stigma. Deciphering the specific transcriptomic response of both pollen and stigmatic cells during their interaction constitutes an important challenge to better our understanding of this cell recognition event. RESULTS: Here we describe a transcriptomic analysis based on single nucleotide polymorphisms (SNPs) present in two Arabidopsis thaliana accessions, one used as female and the other as male. This strategy allowed us to distinguish 80% of transcripts according to their parental origins. We also developed a tool which predicts male/female specific expression for genes without SNP. We report an unanticipated transcriptional activity triggered in stigma upon incompatible pollination and show that following compatible interaction, components of the pattern-triggered immunity (PTI) pathway are induced on the female side. CONCLUSIONS: Our work unveils the molecular signatures of compatible and incompatible pollinations both at the male and female side. We provide invaluable resource and tools to identify potential new molecular players involved in pollen-stigma interaction.
Assuntos
Arabidopsis , Polinização , Arabidopsis/genética , Pólen/genética , Polinização/genética , TranscriptomaRESUMO
The pollen tube grows inside the pistil, carrying male gametes to the ovule. During this journey, it invades diverse tissues, sensing and adapting to abrupt transitions in mechanical environment. In this issue of Developmental Cell, Zhou et al. identify a receptor-like kinase/Rho-GTPase module that regulates adaptation to such a transition.
Assuntos
Flores , Tubo Polínico , Masculino , Óvulo Vegetal , FosfotransferasesRESUMO
Successful fertilization in angiosperms depends on the proper trajectory of pollen tubes through the pistil tissues to reach the ovules. Pollen tubes first grow within the cell wall of the papilla cells, applying pressure to the cell. Mechanical forces are known to play a major role in plant cell shape by controlling the orientation of cortical microtubules (CMTs), which in turn mediate deposition of cellulose microfibrils (CMFs). Here, by combining imaging, genetic and chemical approaches, we show that isotropic reorientation of CMTs and CMFs in aged Col-0 and katanin1-5 (ktn1-5) papilla cells is accompanied by a tendency of pollen tubes to coil around the papillae. We show that this coiled phenotype is associated with specific mechanical properties of the cell walls that provide less resistance to pollen tube growth. Our results reveal an unexpected role for KTN1 in pollen tube guidance on the stigma by ensuring mechanical anisotropy of the papilla cell wall.
Flowering plants produce small particles known as pollen that with the help of the wind, bees and other animals carry male sex cells (sperm) to female sex cells (eggs) contained within flowers. When a grain of pollen lands on the female organ of a flower, called the pistil, it gives rise to a tube that grows through the pistil towards the egg cells at the base. The surface of the pistil is covered in a layer of long cells named papillae. Like most plant cells, the papillae are surrounded by a rigid structure known as the cell wall, which is mainly composed of strands known as microfibrils. The pollen tube exerts pressure on a papilla to allow it to grow through the cell wall towards the base of the pistil. Previous studies have shown that the pistil produces signals that guide pollen tubes to the eggs. However, it remains unclear how pollen tubes orient themselves on the surface of papillae to grow in the right direction through the pistil. Riglet et al. combined microscopy, genetic and chemical approaches to study how pollen tubes grow through the surface of the pistils of a small weed known as Arabidopsis thaliana. The experiments showed that an enzyme called KATANIN conferred mechanical properties to the cell walls of papillae that allowed pollen tubes to grow towards the egg cells, and also altered the orientation of the microfibrils in these cell walls. In A. thaliana plants that were genetically modified to lack KATANIN the pollen tubes coiled around the papillae and sometimes grew in the opposite direction to where the eggs were. KATANIN is known to cut structural filaments inside the cells of plants, animals and most other living things. By revealing an additional role for KATANIN in regulating the mechanical properties of the papilla cell wall, these findings indicate this enzyme may also regulate the mechanical properties of cells involved in other biological processes.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Katanina/metabolismo , Tubo Polínico/crescimento & desenvolvimento , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Parede Celular/metabolismo , Flores , Regulação da Expressão Gênica de Plantas , Katanina/genética , Microfibrilas/metabolismo , Microtúbulos/metabolismoRESUMO
Small noncoding RNAs are central regulators of genome activity and stability. Their regulatory function typically involves sequence similarity with their target sites, but understanding the criteria by which they specifically recognize and regulate their targets across the genome remains a major challenge in the field, especially in the face of the diversity of silencing pathways involved. The dominance hierarchy among self-incompatibility alleles in Brassicaceae is controlled by interactions between a highly diversified set of small noncoding RNAs produced by dominant S-alleles and their corresponding target sites on recessive S-alleles. By controlled crosses, we created numerous heterozygous combinations of S-alleles in Arabidopsis halleri and developed an real-time quantitative PCR assay to compare allele-specific transcript levels for the pollen determinant of self-incompatibility (SCR). This provides the unique opportunity to evaluate the precise base-pairing requirements for effective transcriptional regulation of this target gene. We found strong transcriptional silencing of recessive SCR alleles in all heterozygote combinations examined. A simple threshold model of base pairing for the small RNA-target interaction captures most of the variation in SCR transcript levels. For a subset of S-alleles, we also measured allele-specific transcript levels of the determinant of pistil specificity (SRK), and found sharply distinct expression dynamics throughout flower development between SCR and SRK In contrast to SCR, both SRK alleles were expressed at similar levels in the heterozygote genotypes examined, suggesting no transcriptional control of dominance for this gene. We discuss the implications for the evolutionary processes associated with the origin and maintenance of the dominance hierarchy among self-incompatibility alleles.
Assuntos
Alelos , Proteínas de Arabidopsis/genética , Pareamento de Bases , Inativação Gênica , RNA Interferente Pequeno/genética , Autoincompatibilidade em Angiospermas/genética , Arabidopsis , Proteínas de Arabidopsis/metabolismo , Genes Recessivos , Heterozigoto , RNA Interferente Pequeno/química , RNA Interferente Pequeno/metabolismoRESUMO
Early events occurring at the surface of the female organ are critical for plant reproduction, especially in species with a dry stigma. After landing on the stigmatic papilla cells, the pollen hydrates and germinates a tube, which penetrates the cell wall and grows towards the ovules to convey the male gametes to the embryo sac. In self-incompatible species within the Brassicaceae, these processes are blocked when the stigma encounters an incompatible pollen. Based on the generation of self-incompatible Arabidopsis lines and by setting up a live imaging system, we showed that control of pollen hydration has a central role in pollen selectivity. The faster the pollen pumps water from the papilla during an initial period of 10 min, the faster it germinates. Furthermore, we found that the self-incompatibility barriers act to block the proper hydration of incompatible pollen and, when hydration is promoted by high humidity, an additional control prevents pollen tube penetration into the stigmatic wall. In papilla cells, actin bundles focalize at the contact site with the compatible pollen but not with the incompatible pollen, raising the possibility that stigmatic cells react to the mechanical pressure applied by the invading growing tube.
Assuntos
Arabidopsis , Percepção , Pólen , Tubo Polínico , PolinizaçãoRESUMO
Although the transition to selfing in the model plant Arabidopsis thaliana involved the loss of the self-incompatibility (SI) system, it clearly did not occur due to the fixation of a single inactivating mutation at the locus determining the specificities of SI (the S-locus). At least three groups of divergent haplotypes (haplogroups), corresponding to ancient functional S-alleles, have been maintained at this locus, and extensive functional studies have shown that all three carry distinct inactivating mutations. However, the historical process of loss of SI is not well understood, in particular its relation with the last glaciation. Here, we took advantage of recently published genomic resequencing data in 1,083 Arabidopsis thaliana accessions that we combined with BAC sequencing to obtain polymorphism information for the whole S-locus region at a species-wide scale. The accessions differed by several major rearrangements including large deletions and interhaplogroup recombinations, forming a set of haplogroups that are widely distributed throughout the native range and largely overlap geographically. "Relict" A. thaliana accessions that directly derive from glacial refugia are polymorphic at the S-locus, suggesting that the three haplogroups were already present when glacial refugia from the last Ice Age became isolated. Interhaplogroup recombinant haplotypes were highly frequent, and detailed analysis of recombination breakpoints suggested multiple independent origins. These findings suggest that the complete loss of SI in A. thaliana involved independent self-compatible mutants that arose prior to the last Ice Age, and experienced further rearrangements during postglacial colonization.
Assuntos
Arabidopsis/genética , Autofertilização/genética , Alelos , Sequência de Aminoácidos/genética , Proteínas de Arabidopsis/genética , Evolução Molecular , Genes de Plantas/genética , Haplótipos/genética , Mutação , Filogenia , Proteínas de Plantas/genética , Polimorfismo Genético/genéticaRESUMO
Lipid droplets/oil bodies (OBs) are lipid-storage organelles that play a crucial role as an energy resource in a variety of eukaryotic cells. Lipid stores are mobilized in the case of food deprivation or high energy demands--for example, during certain developmental processes in animals and plants. OB degradation is achieved by lipases that hydrolyze triacylglycerols (TAGs) into free fatty acids and glycerol. In the model plant Arabidopsis thaliana, Sugar-Dependent 1 (SDP1) was identified as the major TAG lipase involved in lipid reserve mobilization during seedling establishment. Although the enzymatic activity of SDP1 is associated with the membrane of OBs, its targeting to the OB surface remains uncharacterized. Here we demonstrate that the core retromer, a complex involved in protein trafficking, participates in OB biogenesis, lipid store degradation, and SDP1 localization to OBs. We also report an as-yet-undescribed mechanism for lipase transport in eukaryotic cells, with SDP1 being first localized to the peroxisome membrane at early stages of seedling growth and then possibly moving to the OB surface through peroxisome tubulations. Finally, we show that the timely transfer of SDP1 to the OB membrane requires a functional core retromer. In addition to revealing previously unidentified functions of the retromer complex in plant cells, our work provides unanticipated evidence for the role of peroxisome dynamics in interorganelle communication and protein transport.
Assuntos
Arabidopsis/genética , Hidrolases de Éster Carboxílico/metabolismo , Gotículas Lipídicas/química , Peroxissomos/química , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Genes de Plantas , Germinação , Lipase/metabolismo , Lipídeos/química , Microscopia Confocal , Mutação , Oxigênio/química , Peroxissomos/metabolismo , Fenótipo , Raízes de Plantas/metabolismo , Transporte Proteico , Plântula/crescimento & desenvolvimento , Sementes/metabolismoRESUMO
The prevention of fertilization through self-pollination (or pollination by a close relative) in the Brassicaceae plant family is determined by the genotype of the plant at the self-incompatibility locus (S locus). The many alleles at this locus exhibit a dominance hierarchy that determines which of the two allelic specificities of a heterozygous genotype is expressed at the phenotypic level. Here, we uncover the evolution of how at least 17 small RNA (sRNA)-producing loci and their multiple target sites collectively control the dominance hierarchy among alleles within the gene controlling the pollen S-locus phenotype in a self-incompatible Arabidopsis species. Selection has created a dynamic repertoire of sRNA-target interactions by jointly acting on sRNA genes and their target sites, which has resulted in a complex system of regulation among alleles.
Assuntos
Arabidopsis/genética , Evolução Biológica , Regulação da Expressão Gênica de Plantas , Redes Reguladoras de Genes , Genes Dominantes , Genes Recessivos , Pequeno RNA não Traduzido/genética , Alelos , Loci Gênicos , Modelos Moleculares , Filogenia , Polinização , Pequeno RNA não Traduzido/classificação , Seleção GenéticaRESUMO
The retromer complex localizes to endosomal membranes and is involved in protein trafficking. In mammals, it is composed of a dimer of sorting nexins and of the core retromer consisting of vacuolar protein sorting (VPS)26, VPS29, and VPS35. Although homologs of these proteins have been identified in plants, how the plant retromer functions remains elusive. To better understand the role of VPS components in the assembly and function of the core retromer, we characterize here Arabidopsis vps26-null mutants. We show that impaired VPS26 function has a dramatic effect on VPS35 levels and causes severe phenotypic defects similar to those observed in vps29-null mutants. This implies that functions of plant VPS26, VPS29, and VPS35 are tightly linked. Then, by combining live-cell imaging with immunochemical and genetic approaches, we report that VPS35 alone is able to bind to endosomal membranes and plays an essential role in VPS26 and VPS29 membrane recruitment. We also show that the Arabidopsis Rab7 homolog RABG3f participates in the recruitment of the core retromer to the endosomal membrane by interacting with VPS35. Altogether our data provide original information on the molecular interactions that mediate assembly of the core retromer in plants.
Assuntos
Arabidopsis/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Citosol/metabolismo , Endossomos/metabolismo , Genótipo , Imunoquímica/métodos , Microscopia Confocal/métodos , Mutagênese Sítio-Dirigida , Mutação , Fenótipo , Fenômenos Fisiológicos Vegetais , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Plasmídeos/metabolismo , Frações Subcelulares/metabolismo , Técnicas do Sistema de Duplo-Híbrido , Proteínas de Transporte Vesicular/genética , Proteínas de Transporte Vesicular/metabolismo , Proteínas rab de Ligação ao GTP/genéticaRESUMO
Self-incompatibility has been considered by geneticists a model system for reproductive biology and balancing selection, but our understanding of the genetic basis and evolution of this molecular lock-and-key system has remained limited by the extreme level of sequence divergence among haplotypes, resulting in a lack of appropriate genomic sequences. In this study, we report and analyze the full sequence of eleven distinct haplotypes of the self-incompatibility locus (S-locus) in two closely related Arabidopsis species, obtained from individual BAC libraries. We use this extensive dataset to highlight sharply contrasted patterns of molecular evolution of each of the two genes controlling self-incompatibility themselves, as well as of the genomic region surrounding them. We find strong collinearity of the flanking regions among haplotypes on each side of the S-locus together with high levels of sequence similarity. In contrast, the S-locus region itself shows spectacularly deep gene genealogies, high variability in size and gene organization, as well as complete absence of sequence similarity in intergenic sequences and striking accumulation of transposable elements. Of particular interest, we demonstrate that dominant and recessive S-haplotypes experience sharply contrasted patterns of molecular evolution. Indeed, dominant haplotypes exhibit larger size and a much higher density of transposable elements, being matched only by that in the centromere. Overall, these properties highlight that the S-locus presents many striking similarities with other regions involved in the determination of mating-types, such as sex chromosomes in animals or in plants, or the mating-type locus in fungi and green algae.
Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Evolução Molecular , Haplótipos , Autoincompatibilidade em Angiospermas/genética , Elementos de DNA Transponíveis/genética , Rearranjo Gênico , Genes Dominantes , Genes Recessivos , Filogenia , Recombinação Genética , Análise de Sequência de DNARESUMO
Sorting nexins (SNXs) are conserved eukaryotic proteins that associate with three types of vacuolar protein sorting (VPS) proteins to form the retromer complex. How SNXs act in this complex and whether they might work independently of the retromer remains elusive. Here, we show by genetic and cell imaging approaches that the Arabidopsis thaliana SNX1 protein recruits SNX2 at the endosomal membrane, a process required for SNX1-SNX2 dimer activity. We report that, in contrast with the mammalian retromer, SNXs are dispensable for membrane binding and function of the retromer complex. We also show that VPS retromer components can work with or independently of SNXs in the trafficking of seed storage proteins, which reveals distinct functions for subcomplexes of the plant retromer. Finally, we provide compelling evidence that the combined loss of function of SNXs and VPS29 leads to embryo or seedling lethality, underlining the essential role of these proteins in development.
Assuntos
Arabidopsis/metabolismo , Transporte Proteico , Nexinas de Classificação/fisiologia , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/fisiologia , Dimerização , Endossomos/metabolismo , Membranas Intracelulares/metabolismoRESUMO
Flowering plants (angiosperms) are the most prevalent and evolutionarily advanced group of plants. Reproductive strategies that promote cross-fertilization have played an essential role in the success of angiosperms as they contribute to genetic variability among plant species. A major genetic barrier to self-fertilization is self-incompatibility (SI), which allows female reproductive cells to discriminate between self- and non-self pollen and specifically reject self-pollen. In this review, we describe three SI mechanisms showing that different flowering plant families use distinct molecules for recognition of self as well as diverse biochemical pathways to arrest pollen tube growth.
Assuntos
Fertilização/fisiologia , Magnoliopsida/fisiologia , Pólen/fisiologia , Brassicaceae/enzimologia , Brassicaceae/genética , Brassicaceae/fisiologia , Fertilização/genética , Magnoliopsida/enzimologia , Magnoliopsida/genética , Papaver/genética , Papaver/fisiologia , Pólen/crescimento & desenvolvimento , Polinização/genética , Polinização/fisiologia , Reprodução/fisiologia , Ribonucleases/genética , Ribonucleases/metabolismoRESUMO
More than half of the flowering plants have a sophisticated mechanism for self-pollen rejection, named self-incompatibility (SI). In Brassicaceae, recognition specificity is achieved by the interaction of the stigmatic S-RECEPTOR KINASE (SRK) and its ligand S-LOCUS CYSTEINE-RICH PROTEIN (SCR). Recent years have seen significant advances in understanding the SI response. Progress has been made on elucidating the regulation and function of proteins that act as either molecular partners of SRK or modulators of SI. Thus, modules controlling the specificity of the central receptor-ligand interaction have been identified on both SRK and SCR proteins. A role for intracellular protein trafficking in SI has also been demonstrated. Here, we integrate the novel findings into the existing model to present the current understanding of SI signaling.
Assuntos
Brassicaceae/fisiologia , Polinização , Animais , Proteínas Nucleares/metabolismo , Proteínas de Plantas/metabolismo , Ligação Proteica , Proteínas Quinases/metabolismo , Transdução de SinaisRESUMO
In eukaryotic cells, the endocytic and secretory pathways are key players in several physiological processes. These pathways are largely inter-connected in animal and yeast cells through organelles named sorting endosomes. Sorting endosomes are multi-vesicular compartments that redirect proteins towards various destinations, such as the lysosomes or vacuoles for degradation, the trans-Golgi network for retrograde transport and the plasma membrane for recycling. In contrast, cross-talk between the endocytic and secretory pathways has not been clearly established in plants, especially in terms of cargo protein trafficking. Here we show by co-localization analyses that endosomes labelled with the AtSORTING NEXIN1 (AtSNX1) protein overlap with the pre-vacuolar compartment in Arabidopsis root cells. In addition, alteration of the routing functions of AtSNX1 endosomes by drug treatments leads to mis-routing of endocytic and secretory cargo proteins. Based on these results, we propose that the AtSNX1 endosomal compartment represents a sorting endosome in root cells, and that this specialized organelle is conserved throughout eukaryotes.
Assuntos
Arabidopsis/fisiologia , Endossomos/fisiologia , Proteínas de Membrana/metabolismo , Raízes de Plantas/fisiologia , Proteínas de Transporte Vesicular/metabolismo , Proteínas de Arabidopsis/metabolismo , Membrana Celular/metabolismo , Vesículas Citoplasmáticas/fisiologia , Endocitose , Raízes de Plantas/ultraestrutura , Transporte Proteico/fisiologia , Rede trans-Golgi/fisiologiaRESUMO
A key feature of plants (as opposed to animals) is their ability to establish new organs not only during embryogenesis, but also throughout their development. A master regulator of organ initiation in plants is the phytohormone auxin. Auxin acts locally as a morphogen and is directionally transported from cell to cell by polarized auxin efflux carriers, termed PIN-FORMED (PIN) proteins. Here we report that the Arabidopsis ortholog of the yeast and mammalian vacuolar protein sorting 29 (VPS29), a member of the retromer complex, mediates the formation of new axes of development. Furthermore, we show that VPS29 is required for endosome homeostasis, PIN protein cycling, and dynamic PIN1 repolarization during development. We propose a model that links VPS29 function, PIN1 polarity, and organ initiation in plants.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Polaridade Celular , Ácidos Indolacéticos/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Proteínas de Transporte Vesicular/metabolismo , Arabidopsis/embriologia , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Proteínas de Transporte/metabolismo , Diferenciação Celular , Proliferação de Células , Cotilédone/metabolismo , Endossomos/metabolismo , Genótipo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Proteínas de Membrana Transportadoras/genética , Meristema/embriologia , Meristema/metabolismo , Complexos Multiproteicos/metabolismo , Mutação , Fenótipo , Plantas Geneticamente Modificadas/embriologia , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Transporte Proteico , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais , Nexinas de Classificação , Proteínas de Transporte Vesicular/genética , Xilema/metabolismoRESUMO
Arabidopsis thaliana has become widely used as a model system for plant biology. Recent phylogenetic studies led to a severe revision of the systematic relationships across species of the Brassicaceae family. This provided an opportunity to examine close relatives of A. thaliana and to study the function and molecular evolution of genes that play roles in ecology and speciation. In this context, developing tools to genetically transform "non-model plants" appears as a major issue to ascertain gene function. Here, we report a method to transform A. lyrata, one of the closest relatives of A. thaliana.
Assuntos
Arabidopsis/genética , Proteínas de Fluorescência Verde/genética , Transformação Genética , Sequência de Bases , Primers do DNA , Microscopia Confocal , Microscopia de Fluorescência , Plantas Geneticamente ModificadasRESUMO
Polarized cellular distribution of the phytohormone auxin and its carriers is essential for normal plant growth and development. Polar auxin transport is maintained by a network of auxin influx (AUX) and efflux (PIN) carriers. Both auxin transport and PIN protein cycling between the plasma membrane and endosomes require the activity of the endosomal GNOM; however, intracellular routes taken by these carriers remain largely unknown. Here we show that Arabidopsis thaliana SORTING NEXIN 1 (AtSNX1) is involved in the auxin pathway and that PIN2, but not PIN1 or AUX1, is transported through AtSNX1-containing endosomes. We demonstrate that the snx1-null mutant exhibits multiple auxin-related defects and that loss of function of AtSNX1 severely enhances the phenotype of a weak gnom mutant. In root cells, we further show that AtSNX1 localizes to an endosomal compartment distinct from GNOM-containing endosomes, and that PIN2 accumulates in this compartment after treatment with the phosphatidylinositol-3-OH kinase inhibitor wortmannin or after a gravity stimulus. Our data reveal the existence of a novel endosomal compartment involved in PIN2 endocytic sorting and plant development.