Project description:tri38-lar - hr - Analyse the transcriptome of Arabidopsis thaliana plants developing localized acquired resistance (LAR) and a hypersensitive response (HR). The goal is to identify genes inducing LAR and/or HR. Here, we want to analyse the transcriptome of Arabidopsis thaliana developing HR. To achieve this, we used Col0 leaf tissues developing an HR reaction after inoculation of the avirulent strain of PstDC3000 carrying the gene avrRpm1. Keywords: normal vs disease comparison
Project description:Arabidopsis genome sequencing has revealed the presence of at least three extensive gene families that may encode protein ligands. One of these, the SPH (S-protein homologue) family, was identified as a direct result of our studies on self-incompatibility in Papaver. The Arabidopsis SPH gene family consists of 81 members. We have initiated experimental work on a subset of these. RT-PCR studies indicate that many, if not all, SPH genes are expressed. Each SPH gene encodes an N-terminal signal peptide sequence and thus SPH proteins are likely to be secreted. Until recently none of the genes in this family had known function. However we have evidence that one member of the family, SPH1 is involved in leaf vascular development. In order to determine the function of SPH1, Arabidopsis plants were transformed with an SPH1 antisense construct. Analysis of the mutant phenotype shows that whilst plants appear as wt until principal growth stage 1.04, they subsequently show severe morphological defects. Plants are severely dwarfed with twisted rosette leaves at ~ 30% wt length and width as well as shortened inflorescence stem. Closer examination revealed aberrant leaf vasculature and severe reduction in expansion of parenchyma cells surrounding the primary leaf vein. We have conducted preliminary immunolocalisation studies with antibody raised to SPH1. These suggest that SPH1 protein is secreted by cells within the developing vasculature of the immature leaf (leaves<6mm in length). The data that we have so far obtained leads us to believe that SPH1 protein acts as a signalling molecule during early leaf development. As SPH1 is likely to be a signalling protein it is assumed that its interaction with a cognate receptor results in initiation of developmental processes within leaf tissue. The purpose of the experiment is to determine the network of genes within the normally developing rosette leaf whose expression is altered by SPH1. This will be accomplished by comparing transcriptional levels in rosette leaves of antisense-SPH1 plants with wt plants. We propose to make target RNA from rosette leaves taken from plants at principal stage 1.05 (immediately after initial appearance of developmental abnormality) and from plants at principal stage 1.14 (where gross changes are apparent). We propose to use replicate slides for each hybridisation, i.e. 2 hybridised to RNA from wt leaves at 1.05, 2 antisense at 1.05, 2 wt at 1.14 and 2 antisense at 1.14. Experiment Overall Design: Number of plants pooled:40
Project description:Plants grow continuously and undergo numerous changes in their vegetative morphology and physiology during their life span. The molecular basis of these changes is largely unknown. To provide a more comprehensive picture of shoot development in Arabidopsis, microarray analysis was used to profile the mRNA content of shoot apices of different ages, as well as leaf primordia and fully-expanded leaves from 6 different positions on the shoot, in early-flowering and late-flowering genotypes. This extensive dataset provides a new and unexpectedly complex picture of shoot development in Arabidopsis. At any given time, the pattern of gene expression is different in every leaf on the shoot, and reflects the activity at least 6 developmental programs. Three of these are specific to individual leaves (leaf maturation, leaf aging, leaf senescence), two occur at the level of the shoot apex (vegetative phase change, floral induction), and one involves the entire shoot (shoot aging). Our results demonstrate that vegetative development is a much more dynamic process that previously imagined, and provide new insights into the underlying mechanism of this process.
Project description:Arabidopsis genome sequencing has revealed the presence of at least three extensive gene families that may encode protein ligands. One of these, the SPH (S-protein homologue) family, was identified as a direct result of our studies on self-incompatibility in Papaver. The Arabidopsis SPH gene family consists of 81 members. We have initiated experimental work on a subset of these. RT-PCR studies indicate that many, if not all, SPH genes are expressed. Each SPH gene encodes an N-terminal signal peptide sequence and thus SPH proteins are likely to be secreted. Until recently none of the genes in this family had known function. However we have evidence that one member of the family, SPH1 is involved in leaf vascular development. In order to determine the function of SPH1, Arabidopsis plants were transformed with an SPH1 antisense construct. Analysis of the mutant phenotype shows that whilst plants appear as wt until principal growth stage 1.04, they subsequently show severe morphological defects. Plants are severely dwarfed with twisted rosette leaves at ~ 30% wt length and width as well as shortened inflorescence stem. Closer examination revealed aberrant leaf vasculature and severe reduction in expansion of parenchyma cells surrounding the primary leaf vein. We have conducted preliminary immunolocalisation studies with antibody raised to SPH1. These suggest that SPH1 protein is secreted by cells within the developing vasculature of the immature leaf (leaves<6mm in length). The data that we have so far obtained leads us to believe that SPH1 protein acts as a signalling molecule during early leaf development. As SPH1 is likely to be a signalling protein it is assumed that its interaction with a cognate receptor results in initiation of developmental processes within leaf tissue. The purpose of the experiment is to determine the network of genes within the normally developing rosette leaf whose expression is altered by SPH1. This will be accomplished by comparing transcriptional levels in rosette leaves of antisense-SPH1 plants with wt plants. We propose to make target RNA from rosette leaves taken from plants at principal stage 1.05 (immediately after initial appearance of developmental abnormality) and from plants at principal stage 1.14 (where gross changes are apparent). We propose to use replicate slides for each hybridisation, i.e. 2 hybridised to RNA from wt leaves at 1.05, 2 antisense at 1.05, 2 wt at 1.14 and 2 antisense at 1.14. Keywords: genetic_modification_design
Project description:tri38-lar - hr - Analyse the transcriptome of Arabidopsis thaliana plants developing localized acquired resistance (LAR) and a hypersensitive response (HR). The goal is to identify genes inducing LAR and/or HR. Here, we want to analyse the transcriptome of Arabidopsis thaliana developing HR. To achieve this, we used Col0 leaf tissues developing an HR reaction after inoculation of the avirulent strain of PstDC3000 carrying the gene avrRpm1. Keywords: normal vs disease comparison 1 dye-swap - CATMA arrays
Project description:Expression of the F-Box protein Leaf Curling Responsiveness (LCR) is regulated by microRNA, miR394, and alterations to this interplay in Arabidopsis thaliana produce defects in leaf polarity and shoot apical meristem (SAM) organisation. Although the miR394-LCR node has been documented in Arabidopsis, the identification of proteins targeted by LCR F-box itself has proven problematic. Here, a proteomic analysis of shoot apices from plants with altered LCR levels identified a member of the Major Latex Protein (MLP) family gene as a potential LCR F-box target. Bioinformatic and molecular analyses also suggested that other MLP family members are likely to be targets for this post-translational regulation. Direct interaction between LCR F-Box and MLP423 was validated. Additional MLP members had reduction in protein accumulation, in varying degrees, mediated by LCR F-Box. Transgenic Arabidopsis lines, in which MLP28 expression was reduced through an artificial miRNA technology, displayed severe developmental defects, including changes in leaf patterning and morphology, shoot apex defects, and eventual premature death. These phenotypic characteristics resemble those of Arabidopsis plants modified to over-express LCR. Taken together, the results demonstrate that MLPs are driven to degradation by LCR, and indicate that MLP gene family is target of miR394-LCR regulatory node, representing potential targets for directly post-translational regulation mediated by LCR F-Box. In addition, MLP28 family member is associated with the LCR regulation that is critical for normal Arabidopsis development.
Project description:Analyses of expression differences in flower bud and leaf of scion and rootstock, in homografts of Arabidopsis Gene expression data of unopened developing flower buds and leaves, newly emerged after homografting, from the scion and the rootstock
Project description:Leaf senescence is the final developmental process that includes the mobilization of nutrients from old leaves to newly growing tissues. The progression of leaf senescence requires dynamic but coordinated changes of gene expression. Although several transcription factors (TFs) are known to be involved in both negative and positive modes of regulation of leaf senescence, detailed mechanisms that underlie the progression of leaf senescence are largely unknown. We report here that the class II ERF transcriptional repressors are controlled by proteasome and regulate the progression of leaf senescence in Arabidopsis. Since we had previously demonstrated that NtERF3, a model of tobacco class II ERFs, specifically interacts with a ubiquitin-conjugating enzyme, we examined the stability of NtERF3 and found that bacterially produced NtERF3 was rapidly degraded by plant protein extracts in vitro. Whereas NtERF3 accumulation was low in plants, it was increased by treatment with a proteasome inhibitor. Arabidopsis class II ERFs, namely, AtERF4 and AtERF8, were also controlled by proteasome and stabilized by aging of plants. The transgenic plants in which NtERF3, AtERF4, and AtERF8 were individually expressed under the control of the 35S promoter exhibited the precocious leaf senescence. Our microarray and RT-PCR analyses revealed that AtERF4 regulated expression of genes involving in various stress responses and leaf senescence. In contrast, aterf4 aterf8 mutant exhibited delayed leaf senescence. Taken together, we present the important role of class II ERFs in the regulation of leaf senescence.