Project description:This study investigates extent and functional significance of alternative splicing in Arabidopsis thaliana defense against the bacterial pathogen Pseudomonas syringae pv tomato (Pst). We have provided a detailed characterization of the Arabidopsis thaliana transcriptional response to Pseudomonas syringae infection in both susceptible and resistant hosts. We carried out two independent inoculation experiments (biological replicates) for each treatment. Col-0 is susceptible to virulent Pst DC3000 but has a functional RPS4 resistance gene effective against DC3000 expressing AvrRps4
Project description:Plant cell walls are complex structures that contain a matrix of cellulose, lignin and hemicellulose. The regulation of the biosynthesis of these components has been well-studied in the eudicot plant Arabidopsis thaliana, and a transcriptional network has been elucidated. Several NAC and MYB family transcription factors are key regulators of secondary cell wall biosynthesis, and their functional characterization provides significant insight into the complex underlying transcriptional network. Genetic and structural evidence suggests that genes controlling this process might be different between eudicots and monocots. Here, the model grass Brachypodium distachyon has been selected to characterize the function of GNRF (GRASS NAC REPRESSOR OF FLOWERING), SWAM1 (SECONDARY WALL ASSOCIATED MYB1), and SWAM4 in the regulation of secondary cell wall biosynthesis. Functional characterization was performed by using the overexpression plants GNRF-OE and SWAM4-OE; sodium azide mutant plants from a TILLING (Targeting Induced Local Lesion IN Genome) collection for gnrf-1, gnrf-2, gnrf-3, gnrf-4, gnrf-5, swam4-1, and swam4-2; a T-DNA insertional mutant plant, gnrf-6; and a dominant repressor plant, SWAM4-DR. GNRF-OE plants remained at juvenile stage and exhibited persistent vegetative growth, and some gnrf mutant plants were late flowering. SWAM4-DR plants were severely dwarfed. Stems of all genotypes mentioned above were subjected to RNA-seq analysis.
Project description:Despite great advances in sequencing capacity, generating functional information for non-model organisms remains a challenge. One solution lies in an improved ability to predict genetic circuits based on primary DNA sequence combined with the characterization of regulatory molecules from model species. Here, we focus on the LEAFY (LFY) transcription factor, a conserved master regulator of floral development. Starting with biochemical and structural information, we built a biophysical model describing LFY DNA binding specificity in vitro that accurately predicts in vivo LFY binding sites in the Arabidopsis thaliana genome. Extending the model to other species, we show that it can correctly identify functional homologs of known LFY targets from Arabidopsis thaliana in other angiosperms, even if a functional shift between orthologs and paralogs has occurred. Moreover, this model demonstrates the evolutionary fluidity of the link between LFY and one of its target genes, underlining how this regulatory interaction can be conserved despite changes in position, sequence and affinity of the LFY binding sites. Our study shows that the cis-element fluidity recently illustrated in animals also exists in plants, and that it can be detected without any experimental work in each individual species, using a biophysical transcription factor model. A. thaliana LEAFY ChIP-seq w control, 2 replicates
Project description:rs04-03_myb - myb - Xylogenesis is a fundamental developmental process that is specific of vascular plants. It allows the formation of xylem, also called wood in trees, a complex tridimensional tissue composed of different cell types. This process occurs through the control of fundamental cell mechanisms like cell division and differentiation, secondary cell wall synthesis, lignin deposition and programmed cell death. Xylogenesis is controlled spatially and temporally by specific genetic programs that involve hundreds of genes. For instance, lignin biosynthetic genes, CAD and CCR, are specifically expressed during xylogenesis through MYB transcription factor binding sites, a process that seems to be common to all vascular plants. We have cloned two xylem specific MYB transcription factors, EgMYB1 et EgMYB2, in Eucalyptus. Interestingly, they are able to bind MYB consensus sequences of CAD and CCR promoters in vitro and to modulate CAD and CCR expression in vivo. When overexpressed in Arabidopsis or tobacco, they affect xylem structure by changing cell wall structure and quality. To follow expression changes of Arabidopsis genes in transgenic plants overexpressing Eg MYB1 and EgMYB2 should help us to find out which genes might be target of those transcription factors. This should help us to decipher the actual role of those two MYBs in xylogenesis, two new members of a large family of transcription factors in plants. - Xylogenesis is a fundamental developmental process that is specific of vascular plants. It allows the formation of xylem, also called wood in trees, a complex tridimensional tissue composed of different cell types. This process occurs through the control of fundamental cell mechanisms like cell division and differentiation, secondary cell wall synthesis, lignin deposition and programmed cell death. Xylogenesis is controlled spatially and temporally by specific genetic programs that involve hundreds of genes. For instance, lignin biosynthetic genes, CAD and CCR, are specifically expressed during xylogenesis through MYB transcription factor binding sites, a process that seems to be common to all vascular plants. We have cloned two xylem specific MYB transcription factors, EgMYB1 et EgMYB2, in Eucalyptus. Interestingly, they are able to bind MYB consensus sequences of CAD and CCR promoters in vitro and to modulate CAD and CCR expression in vivo. When overexpressed in Arabidopsis or tobacco, they affect xylem structure by changing cell wall structure and quality. To follow expression changes of Arabidopsis genes in transgenic plants overexpressing Eg MYB1 and EgMYB2 should help us to find out which genes might be target of those transcription factors. This should help us to decipher the actual role of those two MYBs in xylogenesis, two new members of a large family of transcription factors in plants. Keywords: gene knock in (transgenic)
Project description:MicroRNAs are a class of endogeneously expressed non-coding small, ~21nt RNAs involved in the negative regulation of gene expression. In plants, miRNAs are known to play a critical role in developmental and metabolic pathways, as they predominantly target transcription factors. Studies in Arabidopsis and apple have shown that few microRNAs and small interfering (si) RNAs target MYB transcription factors, which are key regulators of phenylpropanoid pathway. However, it is not well-understood how miRNAs mediate regulation of MYBs to produce secondary metabolites such as anthocyanins and flavonoids. Here we show that, a cluster of abundant miRNAs target MYB transcription factors in anthocyanin rich fruits such as grapes. Using deep small RNA-sequencing we establish that grape varieties with high anthocyanin content express abundant MYB-targeting miRNAs resulting in differential expression of MYB proteins among grape varieties, thereby regulating the phenylpropanoid pathway.