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1.
Nat Commun ; 13(1): 6190, 2022 Oct 19.
Artigo em Inglês | MEDLINE | ID: mdl-36261416

RESUMO

Plant-parasitic nematodes are a major threat to crop production in all agricultural systems. The scarcity of classical resistance genes highlights a pressing need to find new ways to develop nematode-resistant germplasm. Here, we sequence and assemble a high-quality phased genome of the model cyst nematode Heterodera schachtii to provide a platform for the first system-wide dual analysis of host and parasite gene expression over time, covering all major parasitism stages. Analysis of the hologenome of the plant-nematode infection site identified metabolic pathways that were incomplete in the parasite but complemented by the host. Using a combination of bioinformatic, genetic, and biochemical approaches, we show that a highly atypical completion of vitamin B5 biosynthesis by the parasitic animal, putatively enabled by a horizontal gene transfer from a bacterium, is required for full pathogenicity. Knockout of either plant-encoded or now nematode-encoded steps in the pathway significantly reduces parasitic success. Our experiments establish a reference for cyst nematodes, further our understanding of the evolution of plant-parasitism by nematodes, and show that congruent differential expression of metabolic pathways in the infection hologenome represents a new way to find nematode susceptibility genes. The approach identifies genome-editing-amenable targets for future development of nematode-resistant crops.


Assuntos
Cistos , Parasitos , Tylenchida , Animais , Ácido Pantotênico , Transcriptoma
2.
J Plant Physiol ; 272: 153680, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35338957

RESUMO

Sedentary plant parasitic nematodes have developed competences to reprogram host plant cell metabolism via sophisticated manipulation of gene expression, leading to the formation of permanent feeding sites for an unlimited source of food. Arabidopsis thaliana and the beet cyst nematode Heterodera schachtii is a good model for studying the mechanisms of compatible plant-nematode interactions and basic plant responses to nematode infection. Transcription factors are proteins that modulate plant reactions during regular development and under different biotic and abiotic stresses via direct binding to promoter regions of genes. Here, we report on the AtHRS1 gene encoding a MYB-related transcription factor belonging to the GARP family, whose expression is downregulated in syncytia, as confirmed by gene expression analysis. Constitutive overexpression of AtHRS1 disturbed the development of nematode-induced syncytia and led to a reduction in the number of developed females in transgenic A. thaliana roots. In contrast, the hrs1 mutant with decreased expression of AtHRS1 was more susceptible to cyst nematode infection. The influence of AtHRS1 on selected elements of the JA-dependent defence pathway suggests its mode of action in plant response to nematode attack. Based on these results, we suggest that the downregulation of AtHRS1 expression by nematode is important for its successful development.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Cistos , Tylenchoidea , Animais , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Ciclopentanos , Cistos/metabolismo , Feminino , Regulação da Expressão Gênica de Plantas , Células Gigantes/metabolismo , Oxilipinas , Doenças das Plantas/genética , Doenças das Plantas/parasitologia , Raízes de Plantas/metabolismo , Fatores de Transcrição/metabolismo , Tylenchoidea/fisiologia
3.
Int J Mol Sci ; 22(22)2021 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-34830029

RESUMO

Cyst nematodes are important herbivorous pests in agriculture that obtain nutrients through specialized root structures termed syncytia. Syncytium initiation, development, and functioning are a research focus because syncytia are the primary interface for molecular interactions between the host plant and parasite. The small size and complex development (over approximately two weeks) of syncytia hinder precise analyses, therefore most studies have analyzed the transcriptome of infested whole-root systems or syncytia-containing root segments. Here, we describe an effective procedure to microdissect syncytia induced by Globodera rostochiensis from tomato roots and to analyze the syncytial proteome using mass spectrometry. As little as 15 mm2 of 10-µm-thick sections dissected from 30 syncytia enabled the identification of 100-200 proteins in each sample, indicating that mass-spectrometric methods currently in use achieved acceptable sensitivity for proteome profiling of microscopic samples of plant tissues (approximately 100 µg). Among the identified proteins, 48 were specifically detected in syncytia and 7 in uninfected roots. The occurrence of approximately 50% of these proteins in syncytia was not correlated with transcript abundance estimated by quantitative reverse-transcription PCR analysis. The functional categories of these proteins confirmed that protein turnover, stress responses, and intracellular trafficking are important components of the proteome dynamics of developing syncytia.


Assuntos
Cromadoria , Células Gigantes/metabolismo , Lycopersicon esculentum , Proteínas de Plantas/metabolismo , Raízes de Plantas , Proteoma/metabolismo , Animais , Lycopersicon esculentum/metabolismo , Lycopersicon esculentum/parasitologia , Raízes de Plantas/metabolismo , Raízes de Plantas/parasitologia
4.
Int J Mol Sci ; 22(16)2021 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-34445145

RESUMO

The main goal of growing plants under various photoperiods is to optimize photosynthesis for using the effect of day length that often acts on plants in combination with biotic and/or abiotic stresses. In this study, Brassica juncea plants were grown under four different day-length regimes, namely., 8 h day/16 h night, 12 h day/12 h night, 16 h day/8 h night, and continuous light, and were infected with a necrotrophic fungus Alternaria brassicicola. The development of necroses on B. juncea leaves was strongly influenced by leaf position and day length. The largest necroses were formed on plants grown under a 16 h day/8 h night photoperiod at 72 h post-inoculation (hpi). The implemented day-length regimes had a great impact on leaf morphology in response to A. brassicicola infection. They also influenced the chlorophyll and carotenoid contents and photosynthesis efficiency. Both the 1st (the oldest) and 3rd infected leaves showed significantly higher minimal fluorescence (F0) compared to the control leaves. Significantly lower values of other investigated chlorophyll a fluorescence parameters, e.g., maximum quantum yield of photosystem II (Fv/Fm) and non-photochemical quenching (NPQ), were observed in both infected leaves compared to the control, especially at 72 hpi. The oldest infected leaf, of approximately 30% of the B. juncea plants, grown under long-day and continuous light conditions showed a 'green island' phenotype in the form of a green ring surrounding an area of necrosis at 48 hpi. This phenomenon was also reflected in changes in the chloroplast's ultrastructure and accelerated senescence (yellowing) in the form of expanding chlorosis. Further research should investigate the mechanism and physiological aspects of 'green islands' formation in this pathosystem.


Assuntos
Alternaria/patogenicidade , Mostardeira/microbiologia , Mostardeira/fisiologia , Necrose/microbiologia , Necrose/patologia , Fotossíntese/fisiologia , Doenças das Plantas/microbiologia , Carotenoides/metabolismo , Clorofila/metabolismo , Clorofila A/metabolismo , Fluorescência , Mostardeira/metabolismo , Necrose/metabolismo , Fotoperíodo , Complexo de Proteína do Fotossistema II/metabolismo , Folhas de Planta/metabolismo , Folhas de Planta/microbiologia
5.
Cells ; 10(6)2021 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-34199921

RESUMO

Somatic embryogenesis is the formation of a plant embryo from a cell other than the product of gametic fusion. The need to recognize the determinants of somatic cell fate has prompted investigations on how endogenous factors of donor tissues can determine the pattern of somatic embryo origin. The undertaking of this study was enabled by the newly developed experimental system of somatic embryogenesis of the tree fern Cyathea delgadii Sternb., in which the embryos are produced in hormone-free medium. The contents of 89 endogenous compounds (such as sugars, auxins, cytokinins, gibberellins, stress-related hormones, phenolic acids, polyamines, and amino acids) and cytomorphological features were compared between two types of explants giving rise to somatic embryos of unicellular or multicellular origin. We found that a large content of maltose, 1-kestose, abscisic acid, biologically active gibberellins, and phenolic acids was characteristic for single-cell somatic embryo formation pattern. In contrast, high levels of starch, callose, kinetin riboside, arginine, and ethylene promoted their multicellular origin. Networks for visualization of the relations between studied compounds were constructed based on the data obtained from analyses of a Pearson correlation coefficient heatmap. Our findings present for the first time detailed features of donor tissue that can play an important role in the somatic-to-embryogenic transition and the somatic embryo origin.


Assuntos
Citocininas/farmacologia , Gleiquênias/metabolismo , Técnicas de Embriogênese Somática de Plantas , Gleiquênias/citologia
6.
Int J Mol Sci ; 22(12)2021 Jun 16.
Artigo em Inglês | MEDLINE | ID: mdl-34208611

RESUMO

Transcription factors are proteins that directly bind to regulatory sequences of genes to modulate and adjust plants' responses to different stimuli including biotic and abiotic stresses. Sedentary plant parasitic nematodes, such as beet cyst nematode, Heterodera schachtii, have developed molecular tools to reprogram plant cell metabolism via the sophisticated manipulation of genes expression, to allow root invasion and the induction of a sequence of structural and physiological changes in plant tissues, leading to the formation of permanent feeding sites composed of modified plant cells (commonly called a syncytium). Here, we report on the AtMYB59 gene encoding putative MYB transcription factor that is downregulated in syncytia, as confirmed by RT-PCR and a promoter pMyb59::GUS activity assays. The constitutive overexpression of AtMYB59 led to the reduction in A. thaliana susceptibility, as indicated by decreased numbers of developed females, and to the disturbed development of nematode-induced syncytia. In contrast, mutant lines with a silenced expression of AtMYB59 were more susceptible to this parasite. The involvement of ABA in the modulation of AtMYB59 gene transcription appears feasible by several ABA-responsive cis regulatory elements, which were identified in silico in the gene promoter sequence, and experimental assays showed the induction of AtMYB59 transcription after ABA treatment. Based on these results, we suggest that AtMYB59 plays an important role in the successful parasitism of H. schachtii on A. thaliana roots.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Arabidopsis/parasitologia , Regulação da Expressão Gênica de Plantas , Doenças das Plantas/genética , Doenças das Plantas/parasitologia , Fatores de Transcrição/genética , Tylenchoidea/fisiologia , Animais , Arabidopsis/ultraestrutura , Resistência à Doença/genética , Interações Hospedeiro-Parasita , Fenótipo , Reguladores de Crescimento de Plantas/metabolismo , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Raízes de Plantas/parasitologia , Raízes de Plantas/ultraestrutura , Regiões Promotoras Genéticas
7.
New Phytol ; 232(1): 318-331, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34133755

RESUMO

Reactive oxygen species (ROS) generated in response to infections often activate immune responses in eukaryotes including plants. In plants, ROS are primarily produced by plasma membrane-bound NADPH oxidases called respiratory burst oxidase homologue (Rboh). Surprisingly, Rbohs can also promote the infection of plants by certain pathogens, including plant parasitic cyst nematodes. The Arabidopsis genome contains 10 Rboh genes (RbohA-RbohJ). Previously, we showed that cyst nematode infection causes a localised ROS burst in roots, mediated primarily by RbohD and RbohF. We also found that plants deficient in RbohD and RbohF (rbohD/F) exhibit strongly decreased susceptibility to cyst nematodes, suggesting that Rboh-mediated ROS plays a role in promoting infection. However, little information is known of the mechanism by which Rbohs promote cyst nematode infection. Here, using detailed genetic and biochemical analyses, we identified WALLS ARE THIN1 (WAT1), an auxin transporter, as a downstream target of Rboh-mediated ROS during parasitic infections. We found that WAT1 is required to modulate the host's indole metabolism, including indole-3-acetic acid levels, in infected cells and that this reprogramming is necessary for successful establishment of the parasite. In conclusion, this work clarifies a unique mechanism that enables cyst nematodes to use the host's ROS for their own benefit.


Assuntos
Proteínas de Arabidopsis , Cistos , Nematoides , Animais , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Indóis , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Nematoides/metabolismo , Espécies Reativas de Oxigênio/metabolismo
8.
Int J Mol Sci ; 22(8)2021 Apr 18.
Artigo em Inglês | MEDLINE | ID: mdl-33919557

RESUMO

Bacterial angular leaf spot disease (ALS) caused by Pseudomonas syringae pv. lachrymans (Psl) is one of the biological factors limiting cucumber open-field production. The goal of this study was to characterize cytological and transcriptomic response of cucumber to this pathogen. Plants of two inbred lines, B10 (susceptible) and Gy14 (resistant), were grown, and leaves were inoculated with highly virulent Psl strain 814/98 under growth chamber conditions. Microscopic and transcriptional evaluations were performed at three time points: before, 1 and 3 days post inoculation (dpi). Investigated lines showed distinct response to Psl. At 1 dpi bacterial colonies were surrounded by necrotized mesophyll cells. At 3 dpi, in the susceptible B10 line bacteria were in contact with degraded cells, whereas cells next to bacteria in the resistant Gy14 line were plasmolyzed, but apparently still alive and functional. Additionally, the level of H2O2 production was higher in resistant Gy14 plants than in B10 at both examined time points. In RNA sequencing more than 18,800 transcripts were detected in each sample. As many as 1648 and 2755 differentially expressed genes (DEGs) at 1 dpi as well as 2992 and 3141 DEGs at 3 dpi were identified in B10 and Gy14, respectively. DEGs were characterized in terms of functional categories. Resistant line Gy14 showed massive transcriptomic response to Psl at 1 dpi compared to susceptible line B10, while a similar number of DEGs was detected for both lines at 3 dpi. This suggests that dynamic transcriptomic response to the invading pathogen may be related with host resistance. This manuscript provides the first transcriptomic data on cucumber infected with the pathovar lachrymans and helps to elucidate resistance mechanism against ALS disease.


Assuntos
Cucumis sativus/genética , Cucumis sativus/microbiologia , Perfilação da Expressão Gênica/métodos , Pseudomonas syringae/patogenicidade , Regulação da Expressão Gênica de Plantas , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
9.
Genes (Basel) ; 11(12)2020 11 28.
Artigo em Inglês | MEDLINE | ID: mdl-33260722

RESUMO

Although the use of natural resistance is the most effective management approach against the potato cyst nematode (PCN) Globodera pallida, the existence of pathotypes with different virulence characteristics constitutes a constraint towards this goal. Two resistance sources, GpaV (from Solanum vernei) and H3 from S. tuberosum ssp. andigena CPC2802 (from the Commonwealth Potato Collection) are widely used in potato breeding programmes in European potato industry. However, the use of resistant cultivars may drive strong selection towards virulence, which allows the increase in frequency of virulent alleles in the population and therefore, the emergence of highly virulent nematode lineages. This study aimed to identify Avirulence (Avr) genes in G. pallida populations selected for virulence on the above resistance sources, and the genomic impact of selection processes on the nematode. The selection drive in the populations was found to be specific to their genetic background. At the genomic level, 11 genes were found that represent candidate Avr genes. Most of the variant calls determining selection were associated with H3-selected populations, while many of them seem to be organised in genomic islands facilitating selection evolution. These phenotypic and genomic findings combined with histological studies performed revealed potential mechanisms underlying selection in G. pallida.


Assuntos
Nematoides , Doenças das Plantas/genética , Doenças das Plantas/parasitologia , Solanum tuberosum/parasitologia , Animais , Resistência à Doença , Nematoides/genética , Nematoides/patogenicidade , Virulência
10.
Cells ; 9(10)2020 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-33092216

RESUMO

Black spot disease, caused by Alternaria brassicicola in Brassica species, is one of the most devastating diseases all over the world, especially since there is no known fully resistant Brassica cultivar. In this study, the visualization of black spot disease development on Brassica oleracea var. capitata f. alba (white cabbage) leaves and subsequent ultrastructural, molecular and physiological investigations were conducted. Inter- and intracellular hyphae growth within leaf tissues led to the loss of host cell integrity and various levels of organelle disintegration. Severe symptoms of chloroplast damage included the degeneration of chloroplast envelope and grana, and the loss of electron denseness by stroma at the advanced stage of infection. Transcriptional profiling of infected leaves revealed that photosynthesis was the most negatively regulated biological process. However, in infected leaves, chlorophyll and carotenoid content did not decrease until 48 hpi, and several chlorophyll a fluorescence parameters, such as photosystem II quantum yield (Fv/Fm), non-photochemical quenching (NPQ), or plant vitality parameter (Rdf) decreased significantly at 24 and 48 hpi compared to control leaves. Our results indicate that the initial stages of interaction between B. oleracea and A. brassicicola are not uniform within an inoculation site and show a complexity of host responses and fungal attempts to overcome host cell defense mechanisms. The downregulation of photosynthesis at the early stage of this susceptible interaction suggests that it may be a part of a host defense strategy, or, alternatively, that chloroplasts are targets for the unknown virulence factor(s) of A. brassicicola. However, the observed decrease of photosynthetic efficiency at the later stages of infection is a result of the fungus-induced necrotic lesion expansion.


Assuntos
Alternaria/ultraestrutura , Brassica/genética , Brassica/microbiologia , Regulação para Baixo , Interações Hospedeiro-Patógeno/genética , Fotossíntese , Doenças das Plantas/microbiologia , Transcrição Genética , Alternaria/fisiologia , Brassica/fisiologia , Brassica/ultraestrutura , Clorofila A/metabolismo , Cloroplastos/metabolismo , Cloroplastos/ultraestrutura , Suscetibilidade a Doenças , Regulação da Expressão Gênica de Plantas , Ontologia Genética , Células do Mesofilo/microbiologia , Células do Mesofilo/ultraestrutura , Fotossíntese/genética , Folhas de Planta/microbiologia , Folhas de Planta/ultraestrutura , Fatores de Tempo
11.
Plant Cell Physiol ; 61(7): 1273-1284, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32374847

RESUMO

In this report, we describe studies on symplasmic communication and cellular rearrangement during direct somatic embryogenesis (SE) in the tree fern Cyathea delgadii. We analyzed changes in the symplasmic transport of low-molecular-weight fluorochromes, such as 8-hydroxypyrene-1,3,6-trisulfonic acid, trisodium salt (HPTS) and fluorescein (delivered to cells as fluorescein diacetate, FDA), within stipe explants and somatic embryos originating from single epidermal cells and developing during 16-d long culture. Induction of SE is preceded by a restriction in fluorochrome distribution between certain explant cells. Microscopic analysis showed a series of cellular changes like a decrease in vacuole size, increase in vacuole numbers, and increased density of cytoplasm and deposition of electron-dense material in cell walls that may be related with embryogenic transition. In somatic embryos, the limited symplasmic communication between cells was observed first in linear tri-cellular embryos. Further development of the fern embryo was associated with the formation of symplasmic domains corresponding to the four segments of the plant body. Using symplasmic tracers, we provided evidence that the changes in plasmodesmata permeability are corelated with somatic-to-embryogenic transition and somatic embryo development.


Assuntos
Gleiquênias/crescimento & desenvolvimento , Sementes/crescimento & desenvolvimento , Gleiquênias/ultraestrutura , Corantes Fluorescentes , Microscopia Confocal , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Epiderme Vegetal/crescimento & desenvolvimento , Sementes/ultraestrutura
12.
Plant Cell Environ ; 43(5): 1160-1174, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32103526

RESUMO

Plant-parasitic cyst nematodes induce hypermetabolic syncytial nurse cells in the roots of their host plants. Syncytia are their only food source. Cyst nematodes are sexually dimorphic, with their differentiation into male or female strongly influenced by host environmental conditions. Under favourable conditions with plenty of nutrients, more females develop, whereas mainly male nematodes develop under adverse conditions such as in resistant plants. Here, we developed and validated a method to predict the sex of beet cyst nematode (Heterodera schachtii) during the early stages of its parasitism in the host plant Arabidopsis thaliana. We collected root segments containing male-associated syncytia (MAS) or female-associated syncytia (FAS), isolated syncytial cells by laser microdissection, and performed a comparative transcriptome analysis. Genes belonging to categories of defence, nutrient deficiency, and nutrient starvation were over-represented in MAS as compared with FAS. Conversely, gene categories related to metabolism, modification, and biosynthesis of cell walls were over-represented in FAS. We used ß-glucuronidase analysis, qRT-PCR, and loss-of-function mutants to characterize FAS- and MAS-specific candidate genes. Our results demonstrate that various plant-based factors, including immune response, nutrient availability, and structural modifications, influence the sexual fate of the cyst nematodes.


Assuntos
Arabidopsis/parasitologia , Interações Hospedeiro-Parasita , Doenças das Plantas/parasitologia , Raízes de Plantas/parasitologia , Processos de Determinação Sexual , Tylenchoidea/fisiologia , Animais , Feminino , Regulação da Expressão Gênica , Genes de Helmintos , Masculino , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase Via Transcriptase Reversa
13.
Mol Plant Pathol ; 21(1): 38-52, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31605455

RESUMO

Vacuolar processing enzymes (VPEs) play an important role during regular growth and development and defence responses. Despite substantial attempts to understand the molecular basis of plant-cyst nematode interaction, the mechanism of VPEs functioning during this interaction remains unknown. The second-stage Heterodera filipjevi juvenile penetrates host roots and induces the formation of a permanent feeding site called a syncytium. To investigate whether infection with H. filipjevi alters plant host VPEs, the studies were performed in Hordeum vulgare roots and leaves on the day of inoculation and at 7, 14 and 21 days post-inoculation (dpi). Implementing molecular, biochemical and microscopic methods we identified reasons for modulation of barley VPE activity during interaction with H. filipjevi. Heterodera filipjevi parasitism caused a general decrease of VPE activity in infected roots, but live imaging of VPEs showed that their activity is up-regulated in syncytia at 7 and 14 dpi and down-regulated at 21 dpi. These findings were accompanied by tissue-specific VPE gene expression patterns. Expression of the barley cystatin HvCPI-4 gene was stimulated in leaves but diminished in roots upon infestation. External application of cyclotides that can be produced naturally by VPEs elicits in pre-parasitic juveniles vesiculation of their body, enhanced formation of granules, induction of exploratory behaviour (stylet thrusts and head movements), production of reactive oxygen species (ROS) and final death by methuosis. Taken together, down-regulation of VPE activity through nematode effectors promotes the nematode invasion rates and leads to avoidance of the induction of the plant proteolytic response and death of the invading juveniles.


Assuntos
Cisteína Endopeptidases/metabolismo , Hordeum/enzimologia , Hordeum/parasitologia , Doenças das Plantas/parasitologia , Tylenchoidea/fisiologia , Animais , Clorofila/metabolismo , Ciclotídeos/farmacologia , Cistatinas/genética , Perfilação da Expressão Gênica , Hordeum/genética , Interações Hospedeiro-Parasita , Raízes de Plantas/parasitologia
14.
Plant J ; 100(2): 221-236, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31322300

RESUMO

Plant-parasitic nematodes (PPNs) cause tremendous yield losses worldwide in almost all economically important crops. The agriculturally most important PPNs belong to a small group of root-infecting sedentary endoparasites that includes cyst and root-knot nematodes. Both cyst and root-knot nematodes induce specialized long-term feeding structures in root vasculature from which they obtain their nutrients. A specialized cell layer in roots called the endodermis, which has cell walls reinforced with suberin deposits and a lignin-based Casparian strip (CS), protects the vascular cylinder against abiotic and biotic threats. To date, the role of the endodermis, and especially of suberin and the CS, during plant-nematode interactions was largely unknown. Here, we analyzed the role of suberin and CS during interaction between Arabidopsis plants and two sedentary root-parasitic nematode species, the cyst nematode Heterodera schachtii and the root-knot nematode Meloidogyne incognita. We found that nematode infection damages the endodermis leading to the activation of suberin biosynthesis genes at nematode infection sites. Although feeding sites induced by both cyst and root-knot nematodes are surrounded by endodermis during early stages of infection, the endodermis is degraded during later stages of feeding site development, indicating periderm formation or ectopic suberization of adjacent tissue. Chemical suberin analysis showed a characteristic suberin composition resembling peridermal suberin in nematode-infected tissue. Notably, infection assays using Arabidopsis lines with CS defects and impaired compensatory suberization, revealed that the CS and suberization impact nematode infectivity and feeding site size. Taken together, our work establishes the role of the endodermal barrier system in defence against a soil-borne pathogen.


Assuntos
Doenças das Plantas/parasitologia , Raízes de Plantas/citologia , Raízes de Plantas/parasitologia , Tylenchoidea/patogenicidade , Animais , Arabidopsis/citologia , Arabidopsis/metabolismo , Arabidopsis/parasitologia , Parede Celular/metabolismo , Parede Celular/parasitologia , Interações Hospedeiro-Parasita , Lipídeos/fisiologia , Raízes de Plantas/metabolismo
15.
Micron ; 116: 36-45, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30292168

RESUMO

Berry Shrivel (BS) is a post-veraison physiological ripening disorder of grapevine berries. Its symptoms encompass low pH, reduced content of sugars and anthocyanins, and loss of turgor leading to berries shriveling. Evidence for the primary causes of BS is still speculative and anatomical studies are scarce. So far, anatomical studies have determined necrotic cells, degraded primary phloem cells and hardening of secondary phloem cells in the rachis of BS affected grapes. The picture is far from being complete. Herein we report in-depth analyses of the ultrastructure, anatomy and spatial elementary analysis of rachis and pedicel tissues of BS symptomatic grape clusters with different symptom severity. We hypothesize that structural changes in the vascular system of BS affected grape clusters could alter transport functions of the phloem tissue and contribute to the appearance of BS symptoms. By applying different microscopic techniques (LM, SEM, TEM and EDS) we found a number of anatomical differences in both, rachis and pedicels, between H and BS symptomatic grapes, which include: (i) extended areas of collapsed cells and cell wall thickenings in the secondary phloem in BS samples; (ii) reduced number of cell layers in the cambium in BS samples; (iii) higher rate of callose deposition on sieve plates that are additionally covered with a carbohydrate-like material in BS samples; and (iv) reduced (up to 60%) estimated sieve tube conductivity in BS samples.

16.
Protoplasma ; 256(2): 419-429, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30187342

RESUMO

Plant parasitic cyst nematodes induce specific hypermetabolic syncytial nurse cell structures in host roots. A characteristic feature of syncytia is the lack of the central vacuole and the formation of numerous small and larger vesicles. We show that these structures are formed de novo via widening of ER cisternae during the entire development of syncytium, whereas in advanced stages of syncytium development, larger vacuoles are also formed via fusion of vesicles/tubules surrounding organelle-free pre-vacuole regions. Immunogold transmission electron microscopy of syncytia localised the vacuolar markers E subunit of vacuolar H+-adenosinetriphosphatase (V-ATPase) complex and tonoplast intrinsic protein (γ-TIP1;1) mostly in membranes surrounding syncytial vesicles, thus indicating that these structures are vacuoles and that some of them have a lytic character. To study the function of syncytial vacuoles, changes in expression of AtVHA-B1, AtVHA-B2 and AtVHA-B3 (coding for isoforms of subunit B of V-ATPase), and TIP1;1 and TIP1;2 (coding for γ-TIP proteins) genes were analysed. RT-qPCR revealed significant downregulation of AtVHA-B2, TIP1;1 and TIP1;2 at the examined stages of syncytium development compared to uninfected roots. Expression of VHA-B1 and VHA-B3 decreased at 3 dpi but reached the level of control at 7 dpi. These results were confirmed for TIP1;1 by monitoring At-γ-TIP-YFP reporter construct expression. Infection test conducted on tip1;1 mutant plants showed formation of larger syncytia and higher numbers of females in comparison to wild-type plants indicating that reduced levels or lack of TIP1;1 protein promote nematode development.


Assuntos
Proteínas de Arabidopsis/química , Arabidopsis/genética , Beta vulgaris/parasitologia , Dracunculus/patogenicidade , Regulação da Expressão Gênica de Plantas/genética , Vacúolos/química , Animais , Células Gigantes
17.
Plant Cell ; 30(12): 3058-3073, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30413655

RESUMO

Successful biotrophic plant pathogens can divert host nutrition toward infection sites. Here we describe how the protist Plasmodiophora brassicae establishes a long-term feeding relationship with its host by stimulating phloem differentiation and phloem-specific expression of sugar transporters within developing galls. Development of galls in infected Arabidopsis (Arabidopsis thaliana) plants is accompanied by stimulation of host BREVIS RADIX, COTYLEDON VASCULAR PATTERN, and OCTOPUS gene expression leading to an increase in phloem complexity. We characterized how the arrest of this developmental reprogramming influences both the host and the invading pathogen. Furthermore, we found that infection leads to phloem-specific accumulation of SUGARS WILL EVENTUALLY BE EXPORTED TRANSPORTERS11 and 12 facilitating local distribution of sugars toward the pathogen. Utilizing Fourier-transform infrared microspectroscopy to monitor spatial distribution of carbohydrates, we found that infection leads to the formation of a strong physiological sink at the site of infection. High resolution metabolic and structural imaging of sucrose distributions revealed that sweet11 sweet12 double mutants are impaired in sugar transport toward the pathogen, delaying disease progression. This work highlights the importance of precise regulation of sugar partitioning for plant-pathogen interactions and the dependence of P brassicae's performance on its capacity to induce a phloem sink at the feeding site.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Floema/citologia , Floema/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Doenças das Plantas , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Espectroscopia de Infravermelho com Transformada de Fourier
18.
Mol Breed ; 38(9): 114, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30237748

RESUMO

The high content of carotenoids, sugars, dry matter, vitamins and minerals makes the fruit of winter squash (Cucurbita maxima Duchesne) a valuable fresh-market vegetable and an interesting material for the food industry. Due to their nutritional value, long shelf-life and health protective properties, winter squash fruits have gained increased interest from researchers in recent years. Despite these advantages, the genetic and genomic resources available for C. maxima are still limited. The aim of this study was to use the genetic mapping approach to map the ovary colour locus and to identify the quantitative trait loci (QTLs) for high carotenoid content and flesh colour. An F6 recombinant inbred line (RIL) mapping population was developed and used for evaluations of ovary colour, carotenoid content and fruit flesh colour. SSR markers and DArTseq genotyping-by-sequencing were used to construct an advanced genetic map that consisted of 1824 molecular markers distributed across linkage groups corresponding to 20 chromosomes of C. maxima. Total map length was 2208 cM and the average distance between markers was 1.21 cM. The locus affecting ovary colour was mapped at the end of chromosome 14. The identified QTLs for carotenoid content in the fruit and fruit flesh colour shared locations on chromosomes 2, 4 and 14. QTLs on chromosomes 2 and 4 were the most meaningful. A correlation was clearly confirmed between fruit flesh colour as described by the chroma value and carotenoid content in the fruit. A high-density genetic map of C. maxima with mapped loci for important fruit quality traits is a valuable resource for winter squash improvement programmes.

19.
BMC Plant Biol ; 18(1): 183, 2018 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-30189843

RESUMO

BACKGROUND: Pollen development is a strictly controlled post-meiotic process during which microspores differentiate into microgametophytes and profound structural and functional changes occur in organelles. Annexin 5 is a calcium- and lipid-binding protein that is highly expressed in pollen grains and regulates pollen development and physiology. To gain further insights into the role of ANN5 in Arabidopsis development, we performed detailed phenotypic characterization of Arabidopsis plants with modified ANN5 levels. In addition, interaction partners and subcellular localization of ANN5 were analyzed to investigate potential functions of ANN5 at cellular level. RESULTS: Here, we report that RNAi-mediated suppression of ANN5 results in formation of smaller pollen grains, enhanced pollen lethality, and delayed pollen tube growth. ANN5 RNAi knockdown plants also displayed aberrant development during the transition from the vegetative to generative phase and during embryogenesis, reflected by delayed bolting time and reduced embryo size, respectively. At the subcellular level, ANN5 was delivered to the nucleus, nucleolus, and cytoplasm, and was frequently localized in plastid nucleoids, suggesting a likely role in interorganellar communication. Furthermore, ANN5-YFP co-immunoprecipitated with RABE1b, a putative GTPase, and interaction in planta was confirmed in plastidial nucleoids using FLIM-FRET analysis. CONCLUSIONS: Our findings let us to propose that ANN5 influences basal cell homeostasis via modulation of plastid activity during pollen maturation. We hypothesize that the role of ANN5 is to orchestrate the plastidial and nuclear genome activities via protein-protein interactions however not only in maturing pollen but also during the transition from the vegetative to the generative growth and seed development.


Assuntos
Anexina A5/fisiologia , Proteínas de Arabidopsis/fisiologia , Arabidopsis/crescimento & desenvolvimento , Núcleo Celular/metabolismo , Proteínas de Cloroplastos/farmacologia , Plastídeos/fisiologia , Pólen/crescimento & desenvolvimento , Proteínas rab1 de Ligação ao GTP/farmacologia , Anexina A5/genética , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/farmacologia , Clorofila/metabolismo , Proteínas de Cloroplastos/genética , Técnicas de Silenciamento de Genes , Genes de Plantas , Homeostase , Pólen/anatomia & histologia , Pólen/genética , Tubo Polínico/crescimento & desenvolvimento , Plântula/metabolismo , Tabaco/genética , Tabaco/fisiologia , Transcriptoma , Proteínas rab1 de Ligação ao GTP/genética
20.
Plant Cell Rep ; 37(9): 1279-1292, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29947953

RESUMO

KEY MESSAGE: After initial up-regulation, expression of TUBG1 and TUBG2 is significantly down-regulated in mature syncytia, but lack of expression of either of γ-tubulin genes reduces numbers of nematode infections and developing females. Infective second stage juveniles of sedentary plant parasitic nematode Heterodera schachtii invade the root vascular tissue and induce a feeding site, named syncytium, formed as a result of cell hypertrophy and partial cell wall dissolution leading to a multinucleate state. Syncytium formation and maintenance involves a molecular interplay between the plant host and the developing juveniles leading to rearrangements and fragmentation of the plant cytoskeleton. In this study, we investigated the role of two Arabidopsis γ-tubulin genes (TUBG1 and TUBG2), involved in MTs nucleation during syncytium development. Expression analysis revealed that both γ-tubulin's transcript levels changed during syncytium development and after initial up-regulation (1-3 dpi) they were significantly down-regulated in 7, 10 and 15 dpi syncytia. Moreover, TUBG1 and TUBG2 showed distinct immunolocalization patterns in uninfected roots and syncytia. Although no severe changes in syncytium anatomy and ultrastructure in tubg1-1 and tubg2-1 mutants were observed compared to syncytia induced in wild-type plants, nematode infection assays revealed reduced numbers of infecting juveniles and developed female nematodes in mutant lines. Our results indicate that the expression of both TUBG1 and TUBG2 genes, although generally down-regulated in mature syncytia, is essential for successful root infection, development of functional syncytium and nematode maturation.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Arabidopsis/parasitologia , Células Gigantes/citologia , Células Gigantes/metabolismo , Tubulina (Proteína)/metabolismo , Tylenchoidea/patogenicidade , Animais , Arabidopsis/citologia , Proteínas de Arabidopsis/genética , Feminino , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Tubulina (Proteína)/genética
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