Project description:DNA methylation is involved in many biological processes during plant growth and development. Here, we report a novel annual growth rhythm that is found in cotton plants grown in different time-of-year. To further study this rhythm in other plants, we use Arabidopsis thaliana for genome-wide bisulfite sequencing. Two A. thaliana DNA samples were extracted from 20 days old whole plant in Feburary and August for bisulphite treatment and further Illumina sequencing.
Project description:Single-cell genomics provides unprecedented potential for research on plant development and environmental responses. Here, we introduce a generic procedure for plant nuclei isolation combined with nanowell-based library preparation. Our method enables the transcriptome analysis of thousands of individual plant nuclei. It serves as alternative to the use of protoplast isolation, which is currently the standard methodology for plant single-cell genomics, although it can be challenging for some plant tissues. We show the applicability of our nuclei isolation method by using different plant materials from different species. The potential of our snRNA-seq method is shown through the characterization of transcriptomes of seedlings and developing flowers from Arabidopsis thaliana. We evaluated the transcriptome dynamics during the early stages of anther development, identified stage-specific activities of transcription factors underlying this process and predicted potential target genes of these transcription factors. Our nuclei isolation procedure can be applied in different plant species and tissues, thus expanding the toolkit for plant single-cell genomics experiments.
Project description:We uncover distinct DNA methylation dynamics over genes and repeat sequences during early plant life in Arabidopsis. While gene body methylation, which is restricted to CG sites and concerns 20-30% of all genes is detected at all stages examined, it typically fluctuates over a few CGs, with no coherent pattern. In contrast, transposable elements and their relics as well as other repeat sequences, have consistently high methylation levels at CGs and show increasing CHG and especially CHH methylation during embryogenesis. Remarkably, methylation reaches 100% at many individual CHH sites in the mature embryo, compared to the 10-20% methylation usually observed at CHH sites in seedlings or adult plants. Moreover, the progressive increase in CHG and CHH methylation in embryos mirrors the loss of DNA methylation at CG and CHG sites in the endosperm, suggesting transfer of information from the endosperm to the embryo, presumably in the form of small RNAs. Finally, impairing the embryo to seedling transition through loss of PRC2 activity results in the persistence of high CHH methylation levels after germination and specifically over sequences that are targeted by the RNA-directed DNA methylation (RdDM) machinery. Collectively, our findings indicate a lack of extensive resetting of DNA methylation patterns during early plant life and point instead to an important role of RdDM in targeting DNA methylation to transposable element and other repeat sequences in all cells of the mature embryo.
Project description:Nucleosomes package eukaryotic DNA and are composed of four different histone proteins, H3, H4, H2A and H2B. Histone H3 has two main variants, H3.1 and H3.3, which show different genomic localization patterns in animals. We profiled H3.1 and H3.3 variants in the genome of the plant Arabidopsis thaliana and show that the localization of these variants shows broad similarity in plants and animals, in addition to some unique features. H3.1 was enriched in silent areas of the genome including regions containing the repressive chromatin modifications H3 lysine 27 methylation, H3 lysine 9 methylation, and DNA methylation. In contrast, H3.3 was enriched in actively transcribed genes, especially peaking at the 3’ end of genes, and correlated with histone modifications associated with gene activation such as histone H3 lysine 4 methylation, and H2B ubiquitylation, as well as by RNA Pol II occupancy. Surprisingly, both H3.1 and H3.3 were enriched on defined origins of replication, as was overall nucleosome density, suggesting a novel characteristic of plant origins. Our results are broadly consistent with the hypothesis that H3.1 acts as the canonical histone that is incorporated during DNA replication, whereas H3.3 acts as the replacement histone that can be incorporated outside of S-phase during chromatin disrupting processes like transcription.
Project description:DNA methylation is an epigenetic mark that silences transposable elements (TEs) and repeats. Whereas the establishment and maintenance of DNA methylation are relatively well understood, little is known on their dynamics and biological relevance in plant and animal innate immunity. Here, we show that some TEs are demethylated and transcriptionally reactivated during antibacterial defense in Arabidopsis. This effect is concomitant with the down-regulation of key transcriptional gene silencing factors as well as an active demethylation process. DNA demethylation restricts multiplication and vascular propagation of the bacterial pathogen Pseudomonas syringae in leaves and, accordingly, some immune-response genes, containing repeats in their promoters, are negatively regulated by DNA methylation. This study provides evidence that DNA demethylation is part of a plant-induced immune response, potentially acting to prime transcriptional activation of some defense genes linked to Tes/repeats. We have monitored the transcript changes in Arabidopsis plants treated with a flagellin-derived peptide.
Project description:DNA methylation in the promoters of plant genes sometimes leads to transcriptional repression, and the wholesale removal of DNA methylation as seen in methyltransferase mutants results in drastic changes in gene expression and severe developmental defects. However, many cases of naturally-occurring DNA methylation variations have been reported, whereby the altered expression of differentially methylated genes is responsible for agronomically important traits. The ability to manipulate plant methylomes to generate populations of epigenetically distinct individuals could provide invaluable resources for breeding and research purposes. Here we describe “epimutagenesis”, a novel method to rapidly generate variation of DNA methylation through random demethylation of the Arabidopsis thaliana genome. This method involves the expression of a human Ten-eleven translocation (TET) enzyme, and results in widespread hypomethylation that can be inherited to subsequent generations, mimicking mutants in the maintenance DNA methyltransferase met1. Application of TET-mediated epimutagenesis to agriculturally significant plants may result in differential expression of alleles normally silenced by DNA methylation, uncovering previously hidden phenotypic variations.
Project description:DNA methylation is involved in many biological processes during plant growth and development. Here, we report a novel annual growth rhythm that is found in cotton plants grown in different time-of-year. To further study this rhythm in other plants, we use Arabidopsis thaliana for genome-wide bisulfite sequencing.