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:We were interested in changes in small RNA abundance changes in response to developmental transitions in Arabidopsis thaliana shoots, with special focus on vegetative phase change. We specifically wanted to separate the temporal changes in gene expression that result from vegetative phase change and those from flowering. Because of the close timing between the juvenile-to-adult and adult-to-reproductive developmental transitions in Arabidopsis grown under long day conditions, we used the late-flowering genotype FRI;FLC developed by the lab of Richard Amasino by introgressing the FRI allele from Sf-2 into the Col-0 genetic background, which is fri;FLC. For the early flowering genotype, we used FRI;flc-3, also developed by the Amasino lab by EMS-mutagenizing FRI;FLC, identifying early flowering mutants, and backcrossing multiple times to eliminate other EMS-induced mutations. The onset of vegetative phase change in FRI;FLC and FRI;flc-3 under our growth conditions was identical, but the progression was slower in FRI;FLC. By sequencing small RNAs from shoot apices at different time points and fully-expanded leaves at different positions on the shoot and comparing the results between the two genotypes, we were able to obtain a clear picture of changes in small RNA abundance in response to vegetative phase change and flowering in Arabidopsis. For the small RNA samples, we performed two replicates using two different indices in the 5'-adapter and ran each replicate pair on the same sequencing lane. For the cotyledon and leaf samples we only performed one replicate using the same index for all samples because we obtained significantly different results with the two adapters used for the shoot apices, preventing us from using them as true replicates.
Project description:Two Arabidopsis thaliana splicing factor [AtU2AF65 isoforms (AtU2AF65a and AtU2AF65b)] mutants displayed the opposite flowering phenotypes. To assay the RNA processing including alternative splicing and pre-mRNA splicing of target genes of this protein in Arabidopsis, the 7-day seedlings (shoot apices) of wild type atu2af65a and atu2af65b mutants were used for RNA-Seq.
Project description:Microarray analysis of wild type plants and plants with reduced (ago1-27 and se-1) or increased miR156 levels (se-1 p35S:MIR156). Shoot apices were dissected from 20-day-old, short-day grown plants.
Project description:We used LM-RNAseq to compare the molecular fingerprints of cells enriched for subdomains within Selaginella, Equisetum, Arabidopsis and maize shoot apices. Three apical domains were isolated from the Selaginella and Equisetum SAMs: the AC domain, comprising the lone AC; the core domain, comprising the cells below the AC and above the initiating leaf primordium; and the initiating leaf primordium. LM-RNAseq analyses of these shoot apical subdomains generated hundreds of significantly DEGs for each cell type relative to whole-plant transcriptomes based on an FDR ⤠0.05. These data were analyzed for the presence of homologous developmental genetic programs across these three species, and for the identification of unique developmental programs operating within each species.
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:To determine whether IYO is not only necessary but also sufficient to activate transcription of developmental programs, we compared the transcriptome of shoot apices from 35S::IYO-GFP plants to that of 35S::GFP plants at the time of inflorescence emergence. Our results strongly suggest that IYO activates the transcription of key developmental regulators driving differentiation. Shoot apices RNA sample is a pool from RNAs from four independent experiments, and the RNA from each experiment was a pool of RNAs extracted from 12 individuals Arabidopsis 35S:IYO-GFP or 35S:GFP plants.