Function analysis of the Arabidopsis epigenome through integrating genome-wide profiles and cell-type-specific approaches

The eukaryotic epigenome is a dynamic ensemble of chromatin modifications and chromatin structure variations. Its complexity demands experimental methods for the global profiling of epigenomic features as well as informatics tools that facilitate data analyses. I developed Chromatin Immunoprecipitation coupled with high-throughputsequencing (ChIP-seq) to interrogate histone modifications in the model plant Arabidopsis thaliana. Using the ChIP-seq and RNA-seq methods, I produced profiles of nine histone modifications and the transcriptome from aerial tissues of mature plants. With ANchored CORrelative Pattern (ANCORP) method, our analysis has delineated 42 chromatin states with distinct chromatin modification patterns. Selected states were tested by re-ChIP assay to validate the co-localization of histone modifications. I identified the enrichment of Gene Ontology, microRNAs and transposable elements in certain chromatin states, which suggests the regulation of these loci with chromatin-related mechanisms. To derive hypotheses regarding the interactions between the epigenome and transcriptome, State-Specific-Effects-Analysis (SSEA) was developed to incorporate quantitative information related to chromatin states to improve the sensitivity of correlative analyses. Combining ANCORP and SSEA, I identified correlations between the quantity of Natural Antisense Transcripts (NATs) and the enrichment of H3K36me2 and 5mC marks. Genetic analyses identified Polymerase Associated Factors as potential regulators for NAT abundance. I further observed evidence for both nuclear- and cytoplasmically-localized NATs. Although some nuclear-localized NATs are known to regulate chromatin-related functions, our results suggest that NATs may not commonly regulate the cognate locus in cis. Differentiation of distinct cell types is fundamental to the sophisticated body-plan and lifestyle of multicellular organisms. Therefore studying the epigenome in specific cell types is critical for elucidating the function of epigenetic regulation in plant development. I experimented with two different cell/nucleus isolation techniques –Fluorescence Activated Cell Sorting (FACS) and Isolation of Nuclei TAgged in specific Cell Types (INTACT) for performing epigenomic profiling in specific plant cell types. I showed that Arabidopsis root cells labeled with GFP can be effectively isolated with FACS. I initiated the effort to establish a high-throughput and Gateway-compatible INTACT platform for investigating interactions between key regulators and chromatin modifications in root cell layers. Our strategy and progress in this front will be describedPh. D.Includes bibliographical referencesby Chongyuan Lu