Project description:To gain insight into patterns of chromatin accessibility in distinct cell subtypes of mouse pituitary, pituitaries from two adult mice were collected and subjected to single-cell ATAC-seq analysis using 10X Genomics platform
Project description:Cell-to-cell variation is a universal feature of life that impacts a wide range of biological phenomena, from developmental plasticity to tumor heterogeneity. While recent advances have improved our ability to document cellular phenotypic variation the fundamental mechanisms that generate variability from identical DNA sequences remain elusive. Here we reveal the landscape and principles of cellular DNA regulatory variation by developing a robust method for mapping the accessible genome of individual cells via assay of transposase accessible chromatin sequencing (ATAC-seq). Single-cell ATAC-seq (scATAC-seq) maps from hundreds of single-cells in aggregate closely resemble accessibility profiles from tens of millions of cells and provides insights into cell-to-cell variation. Accessibility variance is systematically associated with specific trans-factors and cis-elements, and we discover combinations of trans-factors associated with either induction or suppression of cell-to-cell variability. We further identify sets of trans-factors associated with cell-type specific accessibility variance across 6 cell types. Targeted perturbations of cell cycle or transcription factor signaling evoke stimulus-specific changes in this observed variability. The pattern of accessibility variation in cis across the genome recapitulates chromosome topological domains de novo, linking single-cell accessibility variation to three-dimensional genome organization. All together, single-cell analysis of DNA accessibility provides new insight into cellular variation of the “regulome.” Profiles of single cell epigenomes, assayed using scATAC-seq, across 8 cell types and 4 targeted cell manipulations. The complete data set contains a total of 1,632 assayed wells.
Project description:A single hematopoietic stem cell can give rise to all blood cells with remarkable fidelity. Here, we define the chromatin accessibility and transcriptional landscape controlling this process in thirteen primary cell types that traverse the hematopoietic hierarchy. Exploiting the finding that enhancer landscapes better reflect cell identity than mRNA levels, we enable "enhancer cytometry" for accurate enumeration of pure cell types from complex populations. We further reveal the lineage ontogeny of genetic elements linked to diverse human diseases. In acute myeloid leukemia, chromatin accessibility reveals distinctive regulatory evolution in pre-leukemic HSCs (pHSCs), leukemia stem cells, and leukemic blasts. These leukemic cells demonstrate unique lineage infidelity, confirmed by single cell regulomes. We further show that pHSCs have a competitive advantage that is conferred by reduced chromatin accessibility at HOXA9 targets and is associated with adverse patient outcomes. Thus, regulome dynamics can provide diverse insights into human hematopoietic development and disease. Single-cell ATAC-seq of LMPPs, Monocytes, LSCs and Luekemic blast cells.
Project description:Open or accessible regions of the genome are the primary positions of binding sites for transcription factors and chromatin regulators. Transposase-accessible chromatin sequencing (ATAC-seq) can probe chromatin accessibility in the intact nucleus. Here, we describe a protocol to generate ATAC-seq libraries from fresh Arabidopsis thaliana tissues and establish an easy-to-use bioinformatic analysis pipeline. Our method could be applied to other plants and other tissues and allows for the reliable detection of changes in chromatin accessibility throughout plant growth and development. For complete details on the use and execution of this protocol, please refer to Wang et al. (2020).
Project description:The assay for transposase-accessible chromatin using sequencing (ATAC-seq) provides a simple and scalable way to detect the unique chromatin landscape associated with a cell type and how it may be altered by perturbation or disease. ATAC-seq requires a relatively small number of input cells and does not require a priori knowledge of the epigenetic marks or transcription factors governing the dynamics of the system. Here we describe an updated and optimized protocol for ATAC-seq, called Omni-ATAC, that is applicable across a broad range of cell and tissue types. The ATAC-seq workflow has five main steps: sample preparation, transposition, library preparation, sequencing and data analysis. This protocol details the steps to generate and sequence ATAC-seq libraries, with recommendations for sample preparation and downstream bioinformatic analysis. ATAC-seq libraries for roughly 12 samples can be generated in 10 h by someone familiar with basic molecular biology, and downstream sequencing analysis can be implemented using benchmarked pipelines by someone with basic bioinformatics skills and with access to a high-performance computing environment.
Project description:Multimodal measurements of single-cell profiles are proving increasingly useful for characterizing cell states and regulatory mechanisms. In the present study, we developed PHAGE-ATAC (Assay for Transposase-Accessible Chromatin), a massively parallel droplet-based method that uses phage displaying, engineered, camelid single-domain antibodies ('nanobodies') for simultaneous single-cell measurements of protein levels and chromatin accessibility profiles, and mitochondrial DNA-based clonal tracing. We use PHAGE-ATAC for multimodal analysis in primary human immune cells, sample multiplexing, intracellular protein analysis and the detection of SARS-CoV-2 spike protein in human cell populations. Finally, we construct a synthetic high-complexity phage library for selection of antigen-specific nanobodies that bind cells of particular molecular profiles, opening an avenue for protein detection, cell characterization and screening with single-cell genomics.