Project description:We report the application of laser capture microdissection (LCM) for high resolution transcriptome profiling of the second internode of the Arabidopsis thaliana inflorescence stem. In this series, we used LCM to determine and compare the transcriptome profiles of the phloem cap, the pith, and the remaining vascular bundle area.
Project description:The first described feedback loop of the Arabidopsis circadian clock is based on reciprocal regulation between TOC1 and CCA1/LHY. CCA1 and LHY are MYB transcription factors that bind directly to the TOC1 promoter to negatively regulate its expression. Conversely, the activity of TOC1 has remained less well characterized. Genetic data supports that TOC1 is necessary for the reactivation of CCA1/LHY, but there is little description of its biochemical function. Here we show that TOC1 occupies specific genomic regions in the CCA1 and LHY promoters. Purified TOC1 binds directly to DNA through its CCT domain, which is similar to known DNA binding domains. Chemical induction and transient overexpression of TOC1 in Arabidopsis seedlings cause repression of CCA1/LHY expression demonstrating that TOC1 can repress direct targets, and mutation or deletion of the CCT domain prevents this repression showing that DNA binding is necessary for TOC1 action. Furthermore, we use the Gal4/UAS system in Arabidopsis to show that TOC1 acts as a general transcriptional repressor, and that repression activity is in the Pseudoreceiver (PR) domain of the protein. To identify the genes regulated by TOC1 on a genomic scale, we couple TOC1 chemical induction with microarray analysis and identify new potential TOC1 targets and output pathways. Together these results define the biochemical action of the core clock protein TOC1 and refine our perspective on how plant clocks function. Keywords: Expression profiling by array
Project description:The intention of these gene expression analysis was to study host responses to an infection with Agrobacterium tumefaciens at different stages of crown gall development. Therefore the transcriptome of infected inflorescence stalk tissue and mature crown galls of Arabidopsis thaliana (WS-2) was determined of three different time points. These were compared with the transcriptome of mock-infected inflorescence stalk tissue (reference) of the same age. The following time points were analyzed: (i) three hours post inoculation, before the T-DNA is integrated into the host genome (ii) six days after inoculation when the T-DNA is present in the nucleus and the oncogenes are expressed in the host cell, and (iii) 35 days after inoculation when a mature tumors has developed. For the three-hour- (3hpi) and six-day- time point (6dpi) plants were infected with the virulent strain C58, harboring a T-DNA, or with strain GV3101, containing a disarmed Ti-plasmid. This allows discrimination between signals which derive from the bacterial pathogen and the T-DNA encoded oncogenes. This SuperSeries is composed of the following subset Series:; GSE13929: Arabidopsis thaliana three hours after infection with Agrobacterium tumefaciens; GSE13930: Arabidopsis thaliana six days after infection with Agrobacterium tumefaciens; GSE13927: Transcriptome of mature A. thaliana crown galls. Experiment Overall Design: Refer to individual Series
Project description:The first described feedback loop of the Arabidopsis circadian clock is based on reciprocal regulation between TOC1 and CCA1/LHY. CCA1 and LHY are MYB transcription factors that bind directly to the TOC1 promoter to negatively regulate its expression. Conversely, the activity of TOC1 has remained less well characterized. Genetic data supports that TOC1 is necessary for the reactivation of CCA1/LHY, but there is little description of its biochemical function. Here we show that TOC1 occupies specific genomic regions in the CCA1 and LHY promoters. Purified TOC1 binds directly to DNA through its CCT domain, which is similar to known DNA binding domains. Chemical induction and transient overexpression of TOC1 in Arabidopsis seedlings cause repression of CCA1/LHY expression demonstrating that TOC1 can repress direct targets, and mutation or deletion of the CCT domain prevents this repression showing that DNA binding is necessary for TOC1 action. Furthermore, we use the Gal4/UAS system in Arabidopsis to show that TOC1 acts as a general transcriptional repressor, and that repression activity is in the Pseudoreceiver (PR) domain of the protein. To identify the genes regulated by TOC1 on a genomic scale, we couple TOC1 chemical induction with microarray analysis and identify new potential TOC1 targets and output pathways. Together these results define the biochemical action of the core clock protein TOC1 and refine our perspective on how plant clocks function. Keywords: Expression profiling by array wild type (Col-0) and ALC::TOC1 were sown on Murashige-Skoog with 0.8% agar, stratified for 48 hours and grown in12:12 light:dark (LD) for 12 days and either left in LD or transferred to constant light (LL) and then grown for one more day before the start of the experiment. Tissue was submerged in Murashige-Skoog media supplemented with 2.5% ethanol or no ethanol (mock) and with 20mM MG132 for 3 hours and harvested at ZT1. Three replicates each of the seedlings were collected and frozen in liquid nitrogen.
Project description:We established a novel in vitro culture system, in which vascular development can be induced in Arabidopsis thaliana leaf-disks using the specific GSK3s inhibitor. To elucidate gene expression profiles during vascular development, we performed GeneChip analysis using our newly established culture system.