Biology subjects

Chang, H. Y.

Publications and source records attributed to Chang, H. Y..

8 recordsLinked to original sources

bioRxiv2018

Atlas of Subcellular RNA Localization Revealed by APEX-seq

We introduce APEX-seq, a method for RNA sequencing based on spatial proximity to the peroxidase enzyme APEX2. APEX-seq in nine distinct subcellular locales produced a nanometer-resolution spatial map of the human transcriptome, revealing extensive and exquisite patterns of localization for diverse RNA classes and transcript isoforms. We uncover a radial organization of the nuclear transcriptome, which is gated at the inner surface of the nuclear pore for cytoplasmic export of processed transcripts. We identify two distinct pathways of messenger RNA localization to mitochondria, each associated with specific sets of transcripts for building complementary macromolecular machines within the organelle. APEX-seq should be widely applicable to many systems, enabling comprehensive investigations of the spatial transcriptome.

cell biology↗

bioRxiv2018

Coupled single-cell CRISPR screening and epigenomic profiling reveals causal gene regulatory networks

Here we present Perturb-ATAC, a method which combines multiplexed CRISPR interference or knockout with genome-wide chromatin accessibility profiling in single cells, based on the simultaneous detection of CRISPR guide RNAs and open chromatin sites by assay of transposase-accessible chromatin with sequencing (ATAC-seq). We applied Perturb-ATAC to transcription factors (TFs), chromatin-modifying factors, and noncoding RNAs (ncRNAs) in [~]4,300 single cells, encompassing more than 63 unique genotype-phenotype relationships. Perturb-ATAC in human B lymphocytes uncovered regulators of chromatin accessibility, TF occupancy, and nucleosome positioning, and identified a hierarchical organization of TFs that govern B cell state, variation, and disease-associated cis-regulatory elements. Perturb-ATAC in primary human epidermal cells revealed three sequential modules of cis-elements that specify keratinocyte fate, orchestrated by the TFs JUNB, KLF4, ZNF750, CEBPA, and EHF. Combinatorial deletion of all pairs of these TFs uncovered their epistatic relationships and highlighted genomic co-localization as a basis for synergistic interactions. Thus, Perturb-ATAC is a powerful and general strategy to dissect gene regulatory networks in development and disease.\n\nHighlightsO_LIA new method for simultaneous measurement of CRISPR perturbations and chromatin state in single cells.\nC_LIO_LIPerturb-ATAC reveals regulatory factors that control cis-element accessibility, trans-factor occupancy, and nucleosome positioning.\nC_LIO_LIPerturb-ATAC reveals regulatory modules of coordinated trans-factor activity in B lymphoblasts.\nC_LIO_LIKeratinocyte differentiation is orchestrated by synergistic activities of co-binding TFs on cis-elements.\nC_LI

genomics↗

bioRxiv2018

CRISPRpic: Fast and precise analysis for CRISPR-induced mutations via prefixed index counting

Analysis of CRISPR-induced mutations at targeted loci can be achieved by PCR amplification followed by massively parallel sequencing. We developed a novel algorithm, called CRISPRpic, to analyze sequencing reads from CRISPR experiments via counting exact-matches and pattern-searching. Compared to other methods that are based on sequence alignment, CRISPRpic provides precise mutation calling and ultrafast analysis of sequencing results. The Python script for CRISPRpic is available at https://github.com/compbio/CRISPRpic.

bioinformatics↗

bioRxiv2018

Feedback Regulation between Initiation and Maturation Networks Orchestrates the Chromatin Dynamics of Epidermal Lineage Commitment

Tissue development results from lineage-specific transcription factors (TF) programming a dynamic chromatin landscape through progressive cell fate transitions. Here, we interrogate the epigenomic landscape during epidermal differentiation and create an inference network that ranks the coordinate effects of TF-accessible regulatory element-target gene expression triplets on lineage commitment. We discover two critical transition periods: surface ectoderm initiation and keratinocyte maturation, and identify TFAP2C and p63 as lineage initiation and maturation factors, respectively. Surprisingly, we find that TFAP2C, and not p63, is sufficient to initiate surface ectoderm differentiation, with TFAP2C-initiated progenitor cells capable of maturing into functional keratinocytes. Mechanistically, TFAP2C primes the surface ectoderm chromatin landscape and induces p63 expression and binding sites, thus allowing maturation factor p63 to positively auto-regulate its expression and close a subset of the TFAP2C-initiated early program. Our work provides a general framework to infer TF networks controlling chromatin transitions that will facilitate future regenerative medicine advances.

cell biology↗

bioRxiv2018

Morphogen-Lineage Selector Interactions During Surface Epithelial Commitment

Human embryonic stem cell (hESC) differentiation promises advances in regenerative medicine1-3, yet conversion of hESCs into tissues such as keratinocytes requires a better understanding of epigenetic interactions between the inductive morphogens retinoic acid (RA) and bone morphogenetic protein 4 (BMP), and the master regulator p634,5. Here we develop a robust, defined, keratinocyte differentiation system, and use a multi-dimensional genomics approach to interrogate the contributions of the morphogens and lineage selector to chromatin dynamics during early surface ectoderm commitment. In stark contrast to other master regulators6-9, we find using p63 gain and loss of function hESC lines, that p63 effects major transcriptional changes only after morphogenetic action. Morphogens alter chromatin accessibility and histone modifications, establishing an epigenetic landscape for p63 to modify. In turn, p63 closes chromatin accessibility and promotes the accumulation of repressive H3K27me3 histone modifications at sites distal to where it binds. Surprisingly, cohesin HiChIP10 visualization of genome-wide chromosome conformation reveals that both p63 and the morphogens contribute to dynamic long-range genomic interactions that increase the probability of negative transcriptional regulation at p63 target loci. p63-regulated accessibility, not H3K27me3 deposition, appears to drive early transcriptional changes. We illustrate morphogen-selector interactions by studying p63 negative feedback regulation of TFAP2Ci11, whereby disruption of the single p63 binding site results in a loss of p63-mediated transcriptional control and dramatic increases in TFAP2C and p63 expression. Our study reveals the unexpected dependency of p63 on morphogenetic signaling to control long-range chromatin interactions during tissue specification and provides novel insights into how master regulators specify diverse morphological outcomes.

genetics↗

bioRxiv2017

Enhancer connectome in primary human cells reveals target genes of disease-associated DNA elements

The challenge of linking intergenic mutations to target genes has limited molecular understanding of diverse human diseases. Here, we show H3K27ac HiChIP generates high-resolution contact maps of active enhancers and target genes in rare primary human T cell subtypes and coronary artery smooth muscle cells. Differentiation of naive T cells to either T helper 17 cells or regulatory T cells create subtype-specific enhancer-promoter interactions, specifically at regions of shared DNA accessibility. These data provide a principled means of assigning molecular functions to autoimmune and cardiovascular disease risk variants, linking hundreds of noncoding variants to putative gene targets. Target genes identified with HiChIP are further supported by CRISPR interference and activation at linked enhancers, by the presence of expression quantitative trait loci, and by allele-specific enhancer loops in patient-derived primary cells. The majority of disease-associated enhancers contact genes beyond the nearest gene in the linear genome, leading to a four-fold increase of potential target genes for autoimmune and cardiovascular diseases.

genomics↗

bioRxiv2017

An improved ATAC-seq protocol reduces background and enables interrogation of frozen tissues

We present Omni-ATAC, an improved ATAC-seq protocol for chromatin accessibility profiling that works across multiple applications with substantial improvement of signal-to-background ratio and information content. The Omni-ATAC protocol enables chromatin accessibility profiling from archival frozen tissue samples and 50 m sections, revealing the activities of disease-associated DNA elements in distinct human brain structures. The Omni-ATAC protocol enables the interrogation of personal regulomes in tissue context and translational studies.

genomics↗

bioRxiv2016

Genetic determinants of chromatin accessibility and gene regulation in T cell activation across human individuals

Over 90% of genetic variants associated with complex human traits map to non-coding regions, but little is understood about how they modulate gene regulation in health and disease. One possible mechanism is that genetic variants affect the activity of one or more cis-regulatory elements leading to gene expression variation in specific cell types. To identify such cases, we analyzed Assay for Transposase-Accessible Chromatin sequencing (ATAC-seq) and RNA-seq profiles from activated CD4+ T cells of up to 105 healthy donors. We found that regions of accessible chromatin (ATAC-peaks) are co-accessible at kilobase and megabase resolution, in patterns consistent with the 3D organization of chromosomes measured by in situ Hi-C in T cells. 15% of genetic variants located within ATAC-peaks affected the accessibility of the corresponding peak through disrupting binding sites for transcription factors important for T cell differentiation and activation. These ATAC quantitative trait nucleotides (ATAC-QTNs) have the largest effects on co-accessible peaks, are associated with gene expression from the same aliquot of cells, are rarely affecting core binding motifs, and are enriched for autoimmune disease variants. Our results provide insights into how natural genetic variants modulate cis- regulatory elements, in isolation or in concert, to influence gene expression in primary immune cells that play a key role in many human diseases.

genomics↗