Cell type identity determines transcriptomic immune responses in Arabidopsis thaliana roots
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ABSTRACT: Root pathogens are a major thread in global crop production and protection strategies are required to sustainably enhance the efficiency of root immunity. Our understanding of root immunity is still limited in comparison to the knowledge gained for the regulation of immune response in leaves. In an effort to reveal the organisation of root immunity in roots, we undertook a cell type-specific transcriptome analysis to identify gene networks in epidermis, cortex and pericycle cells of Arabidopsis roots upon treatment with two immunity elicitors, bacterial microbe-associated molecular pattern flagellin and the endogenous damage-associated molecular pattern Pep1. Our analyses revealed that both elicitors induced cell type-specific immunity gene networks. Interestingly, both elicitors did not alter cell identity determining gene networks. Using sophisticated paired motif promoter analyses, we identified key transcription factor pairs involved in the regulation of cell type-specific immunity networks. In addition, our data show that cell identity networks are liaised with cell immunity networks to activate cell type-specific immune response according to the functional capabilities of each cell type.
Project description:The apical root growth is immensely affected by pathogen-associated molecular patterns (PAMPs) or plant-derived damage-associated molecular pattern (DAMPs). Upon perceiving the danger signals, elevated defense machinery often compromises root apical growth, resulting in the meristem's aberrant proliferative and formative divisions. In our previous study, we reported that one of the DAMP signals, PEP1, reprograms developmental pathways in the root apical meristem resulting in the reduction of stele cell numbers, alteration of the xylem vessel organizations, and disruption of cell-to-cell symplastic connections. However, a comprehensive atlas of the gene regulatory networks associated with the root growth-defense tradeoffs is yet to be elucidated. To gain an insight into the dynamic molecular framework that modulates root development under elevated pathogenic signals, we performed a time course RNA-sequencing analysis in the root meristem after PEP1 treatment. Based on expression dynamics of the DEGs (fold change ≥ 1.5; p < 0.05), 12 clusters were obtained following K-means algorithm. Intriguingly, among the PEP1-induced gene clusters, we observed that a group of zinc finger proteins, comprising one of the most prominent transcription factor families, are highly enriched. On the contrary, the PEP1-repressed gene clusters included various developmental regulators expressed in the vascular tissues. Indeed, an in vitro network analysis using selected PEP1-induced and repressed clusters indicated STZ as a potential key mediator of the defense-induced developmental reprogramming pathway in the root stele. We also revealed the involvement of two related homologs of STZ, ZAT6, and AZF3, in this trade-off pathway. Our study provides a comprehensive overview of transcription factors that respond to the early hours of PEP1 induction and hijack cellular machinery, thereby contributing to cellular reorganization and growth in the root apical meristem.
Project description:Plant responses to drought stress require the regulation of transcriptional networks via drought responsive transcription factors, which mediate a range of morphological and physiological changes. AP2/ERF transcription factors are known to act as key regulators of drought resistance transcriptional networks; however, little is known about the associated molecular mechanisms that give rise to specific morphological and physiological adaptations. In this study, we functionally characterized the rice (Oryza sativa) drought responsive AP2/ERF transcription factor, OsERF71, which is predominantly expressed in the root meristem, pericycle, and endodermis. Overexpression of OsERF71 either throughout the entire plant or specifically in roots, resulted in a drought resistance phenotype at the vegetative growth stage, indicating that overexpression in roots was sufficient to confer drought resistance. The root specific overexpression was more effective in conferring drought resistance at the reproductive stage, such that grain yield was increased by 23-42% over wild type plants or whole-body overexpressing transgenic lines under drought conditions. OsERF71 overexpression in roots elevated the expression levels of genes related to cell wall loosening and lignin biosynthetic genes, which correlated with changes in root structure, the formation of enlarged aerenchyma and high lignification levels. Furthermore, OsERF71 was found to directly bind to the promoter of OsCCR1, a key gene in lignin biosynthesis. These results indicate that the OsERF71-mediated drought resistance pathway recruits factors involved in cell wall modification to enable root morphological adaptations, thereby providing a mechanism for enhancing drought resistance.
Project description:Plants adjust their growth in response to environmental cues by forming new organs in different development contexts. Underground lateral roots initiate from prepatterned cells in the main root, but cells can also bypass the root/shoot trajectory separation and generate shoot-borne-roots through an unknown mechanism. Here, we mapped tomato (Solanum lycoperiscum) shoot-borne-roots development at single-cell resolution and show that they initiate from differentiated phloem-associated cells via a unique transitional stem-cell-like state. This state required the activity of a transcription factor which we named SHOOTBORNE ROOTLESS (SBRL), a function that was deeply conserved in angiosperms. Phylogenetic analysis revealed that SBRL arose in angiosperms as an ancient duplicated superlocus with its paralogs showing root-type-specific transient expression in wound-induced and lateral root initiation. Mutants in all SBRL-like genes completely lost post-embryonic roots. We propose that the activation of a common transition state by context-specific regulators underlies the plasticity of plant root systems.
Project description:Cell-type specific transcriptional profiles were generated by FACS (Fluorescence Activated Cell Sorting) sorting of roots that express cell-type specific GFP-reporters. Five different GFP-reporter lines were utilized allowing us to obtain transcriptional profiles for cells in all radial zones of the root. FACS cell populations were isolated from roots grown under standard conditions or roots that had been transfered to -Fe media for 24 hours. Little is known about how developmental cues affect the way cells interpret their environment. Here we characterize the transcriptional response of different cell layers and developmental stages of the Arabidopsis root to high salinity and find that transcriptional responses are highly constrained by developmental parameters. These transcriptional changes lead to the differential regulation of specific biological functions in subsets of cell-layers, several of which correspond to observable physiological changes. We show that known stress pathways primarily control semi-ubiquitous responses and use mutants that disrupt epidermal patterning to reveal cell-layer specific and inter-cell-layer effects. By performing a similar analysis using iron-deprivation we identify common cell-type specific stress responses and environment-independent biological functions that define each cell type. Keywords: Cell-type specific analysis using FACS
Project description:Arbuscular mycorrhiza (AM) interactions between plants and Glomeromycota fungi primarily support phosphate aquisition of most terrestrial plant species. To unravel cell-type specific gene expression during late stages of Medicago truncatula root colonization by AM fungi, we used genome-wide transcriptome profiling based on laser-microdissected cells. We used Medicago GeneChips to detail the cell-type specific programme of gene expression in late stages of colonization by arbuscular mycorrhizal fungi and identified genes differentially expressed during these stages. Medicago truncatula Gaertn M-bM-^@M-^XJemalongM-bM-^@M-^Y genotype A17 plantlets were grown in the climate chamber. Plants grown for the collection of root cortical cells containing arbuscules (ARB), root cortical cells from mycorrhizal roots (CMR), and root epidermal cells from mycorrhizal roots (EPI) were mycorrhized after 2 weeks with Glomus intraradices and mycorrhizal roots were harvested at around 21 days post inoculation (dpi).
Project description:The basidiomycete Ustilago maydis causes smut disease in maize. Colonization of the host plant is initiated by direct penetration of cuticle and cell wall of maize epidermis cells. The invading hyphae are surrounded by the plant plasma membrane and proliferate within the plant tissue. We identified a novel secreted protein, termed Pep1. Disruption mutants of pep1 are not affected in saprophytic growth and develop normal infection structures. However, Îpep1 mutants fail to penetrate the epidermal cell wall and elicit a strong plant defense response. Using Affymetrix maize arrays we identified about 110 plant genes which are differentially regulated in Îpep1 and wild type infections during the penetration stage. Experiment Overall Design: In three independent experiments plants were infected with the strain SG200Dpep1 which is derived from the solopathogenic U. maydis strain SG200. Samples from infected leaves were taken at 24 hours post infection. Samples were treated under the same conditions as described previosly (Doehlemann et al. (2008) Plant J, in press).
Project description:Symbiotic legume nodules and lateral roots arise away from the root meristem via dedifferentiation events. While these organs share some morphological and developmental similarities, whether legume nodules are modified lateral roots is an open question. We dissected emerging nodules (EN), mature nodules (MN), emerging lateral roots (ELR) and young lateral roots (YLR), and constructed strand-specific RNAseq libraries using polyA-enriched RNA preparations. Root sections above and below these organs devoid of any lateral organs were used to construct respective control tissue libraries (ABEN, ABMN, ABELR, ABYLR respectively). High sequence quality, predominant mapping to coding sequences, and consistency between replicates indicated that the RNAseq libraries were of very high quality. We identified genes enriched in emerging nodules, mature nodules, emerging lateral roots and young lateral roots in soybean by comparing global gene expression profiles between each of these organs and adjacent root segments. Potential uses for this high quality transcriptome data set include generation of global gene regulatory networks to identify key regulators; metabolic pathway analyses and comparative analysis of key gene families to discover organ-specific biological processes; and identification of organ-specific alternate spliced transcripts. When combined with other similar datasets especially from leguminous plants these analyses can help answer questions on the evolutionary origins of root nodules and relationships between the development of different plant lateral organs.
Project description:Detached Arabidopsis leaves can regenerate adventitious roots, providing a platform to study de novo root regeneration (DNRR). We performed time-lapse RNA-seq within 5 d revealed activation of gene networks in cell fate transition.
Project description:Despite the broad use of single-cell and single-nucleus RNA sequencing in plant research, accurate cluster annotation in less studied plant species remains a major challenge due to the lack of validated marker genes. Here, using soil-grown wheat roots as a model, we generated a single-cell RNA-sequencing (scRNA-seq) atlas and annotated cluster identities in an unbiased way by transferring existing annotations from publicly available datasets in wheat, rice, maize and Arabidopsis. These cross-species orthology-based predictions were next validated using untargeted spatial transcriptomics. This information refined existing cluster annotations for different datasets across key plant model species. We then used the validated clusters to generate cell type-specific gene regulatory networks for root tissues of wheat and two other monocot crop species. By integrating all available data, including homeolog expression in wheat, we predicted reliable tissue-specific markers which are conserved across different species. In summary, we provided an annotated and validated single cell transcriptomic resource for soil-grown wheat root apical meristems and revealed conserved cell type-specific regulators and markers across species. These data expand upon previous root single cell atlas resources in crops, and will facilitate cell type annotation in non-model plant species in the future.
Project description:Despite the broad use of single-cell and single-nucleus RNA sequencing in plant research, accurate cluster annotation in less studied plant species remains a major challenge due to the lack of validated marker genes. Here, using soil-grown wheat roots as a model, we generated a single-cell RNA-sequencing (scRNA-seq) atlas and annotated cluster identities in an unbiased way by transferring existing annotations from publicly available datasets in wheat, rice, maize and Arabidopsis. These cross-species orthology-based predictions were next validated using untargeted spatial transcriptomics. This information refined existing cluster annotations for different datasets across key plant model species. We then used the validated clusters to generate cell type-specific gene regulatory networks for root tissues of wheat and two other monocot crop species. By integrating all available data, including homeolog expression in wheat, we predicted reliable tissue-specific markers which are conserved across different species. In summary, we provided an annotated and validated single cell transcriptomic resource for soil-grown wheat root apical meristems and revealed conserved cell type-specific regulators and markers across species. These data expand upon previous root single cell atlas resources in crops, and will facilitate cell type annotation in non-model plant species in the future.