Hyunjin Shin

Nucleosome Dynamics Define Transcriptional Enhancers

Chromatin plays a central role in eukaryotic gene regulation. We performed genome-wide mapping of epigenetically marked nucleosomes to determine their position both near transcription start sites and at distal regulatory elements, including enhancers. In prostate cancer cells, where androgen receptor binds primarily to enhancers, we found that androgen treatment dismisses a central nucleosome present at androgen receptor binding sites that is flanked by a pair of marked nucleosomes. A new quantitative model built on the behavior of such nucleosome pairs correctly identified regions bound by the regulators of the immediate androgen response, including androgen receptor and FOXA1. More importantly, this model also correctly predicted previously unidentified binding sites for other transcription factors present after prolonged androgen stimulation, including OCT1 and NKX3-1. Therefore, quantitative modeling of enhancer structure provides a powerful predictive method to infer the identity of transcription factors involved in cellular responses to specific stimuli.

Cistrome: an integrative platform for transcriptional regulation studies

The increasing volume of ChIP-chip and ChIP-seq data being generated creates a challenge for standard, integrative and reproducible bioinformatics data analysis platforms. We developed a web-based application called Cistrome, based on the Galaxy open source framework. In addition to the standard Galaxy functions, Cistrome has 29 ChIP-chip- and ChIP-seq-specific tools in three major categories, from preliminary peak calling and correlation analyses to downstream genome feature association, gene expression analyses, and motif discovery. Cistrome is available at http://cistrome.org/ap/.

CEAS: cis-regulatory element annotation system

UNLABELLEDnWe present a tool designed to characterize genome-wide protein-DNA interaction patterns from ChIP-chip and ChIP-Seq data. This stand-alone extension of our web application CEAS (cis-regulatory element annotation system) provides summary statistics on ChIP enrichment in important genomic regions such as individual chromosomes, promoters, gene bodies or exons, and infers the genes most likely to be regulated by the binding factor under study. CEAS also enables biologists to visualize the average ChIP enrichment signals over specific genomic regions, particularly allowing observation of continuous and broad ChIP enrichment that might be too subtle to detect from ChIP peaks alone.nnnAVAILABILITYnThe CEAS Python package is publicly available at http://liulab.dfci.harvard.edu/CEAS.

Broad chromosomal domains of histone modification patterns in C. elegans

Chromatin immunoprecipitation identifies specific interactions between genomic DNA and proteins, advancing our understanding of gene-level and chromosome-level regulation. Based on chromatin immunoprecipitation experiments using validated antibodies, we define the genome-wide distributions of 19 histone modifications, one histone variant, and eight chromatin-associated proteins in Caenorhabditis elegans embryos and L3 larvae. Cluster analysis identified five groups of chromatin marks with shared features: Two groups correlate with gene repression, two with gene activation, and one with the X chromosome. The X chromosome displays numerous unique properties, including enrichment of monomethylated H4K20 and H3K27, which correlate with the different repressive mechanisms that operate in somatic tissues and germ cells, respectively. The data also revealed striking differences in chromatin composition between the autosomes and between chromosome arms and centers. Chromosomes I and III are globally enriched for marks of active genes, consistent with containing more highly expressed genes, compared to chromosomes II, IV, and especially V. Consistent with the absence of cytological heterochromatin and the holocentric nature of C. elegans chromosomes, markers of heterochromatin such as H3K9 methylation are not concentrated at a single region on each chromosome. Instead, H3K9 methylation is enriched on chromosome arms, coincident with zones of elevated meiotic recombination. Active genes in chromosome arms and centers have very similar histone mark distributions, suggesting that active domains in the arms are interspersed with heterochromatin-like structure. These data, which confirm and extend previous studies, allow for in-depth analysis of the organization and deployment of the C. elegans genome during development.

Differentiation-Specific Histone Modifications Reveal Dynamic Chromatin Interactions and Partners for the Intestinal Transcription Factor CDX2

VIDEO ABSTRACTnCell differentiation requires remodeling of tissue-specific gene loci and activities of key transcriptional regulators, which are recognized for their dominant control over cellular programs. Using epigenomic methods, we characterized enhancer elements specifically modified in differentiating intestinal epithelial cells and found enrichment of transcription factor-binding motifs corresponding to CDX2, a critical regulator of the intestine. Directed investigation revealed surprising lability in CDX2 occupancy of the genome, with redistribution from hundreds of sites occupied only in proliferating cells to thousands of new sites in differentiated cells. Knockout mice confirmed distinct Cdx2 requirements in dividing and mature adult intestinal cells, including responsibility for the active enhancer configuration associated with maturity. Dynamic CDX2 occupancy corresponds with condition-specific gene expression and, importantly, to differential co-occupancy with other tissue-restricted transcription factors, such as GATA6 and HNF4A. These results reveal dynamic, context-specific functions and mechanisms of a prominent transcriptional regulator within a cell lineage.

Identifying Positioned Nucleosomes with Epigenetic Marks in Human from ChIP-Seq

BackgroundIn vivo positioning and covalent modifications of nucleosomes play an important role in epigenetic regulation, but genome-wide studies of positioned nucleosomes and their modifications in human still remain limited.ResultsThis paper describes a novel computational framework to efficiently identify positioned nucleosomes and their histone modification profiles from nucleosome-resolution histone modification ChIP-Seq data. We applied the algorithm to histone methylation ChIP-Seq data in human CD4+ T cells and identified over 438,000 positioned nucleosomes, which appear predominantly at functionally important regions such as genes, promoters, DNase I hypersensitive regions, and transcription factor binding sites. Our analysis shows the identified nucleosomes play a key role in epigenetic gene regulation within those functionally important regions via their positioning and histone modifications.ConclusionOur method provides an effective framework for studying nucleosome positioning and epigenetic marks in mammalian genomes. The algorithm is open source and available at http://liulab.dfci.harvard.edu/NPS/.

Estrogen receptor prevents p53-dependent apoptosis in breast cancer

More than two-thirds of breast cancers express the estrogen receptor (ER) and depend on estrogen for growth and survival. Therapies targeting ER function, including aromatase inhibitors that block the production of estrogens and ER antagonists that alter ER transcriptional activity, play a central role in the treatment of ER+ breast cancers of all stages. In contrast to ER− breast cancers, which frequently harbor mutations in the p53 tumor suppressor, ER+ breast cancers are predominantly wild type for p53. Despite harboring wild-type p53, ER+ breast cancer cells are resistant to chemotherapy-induced apoptosis in the presence of estrogen. Using genome-wide approaches, we have addressed the mechanism by which ER antagonizes the proapoptotic function of p53. Interestingly, both ER agonists such as estradiol and the selective ER modulator (SERM) tamoxifen promote p53 antagonism. In contrast, the full ER antagonist fulvestrant blocks the ability of ER to inhibit p53-mediated cell death. This inhibition works through a mechanism involving the modulation of a subset of p53 and ER target genes that can predict the relapse-free survival of patients with ER+ breast cancer. These findings suggest an improved strategy for the treatment of ER+ breast cancer using antagonists that completely block ER action together with drugs that activate p53-mediated cell death.

TCF4 and CDX2, major transcription factors for intestinal function, converge on the same cis-regulatory regions

Surprisingly few pathways signal between cells, raising questions about mechanisms for tissue-specific responses. In particular, Wnt ligands signal in many mammalian tissues, including the intestinal epithelium, where constitutive signaling causes cancer. Genome-wide analysis of DNA cis-regulatory regions bound by the intestine-restricted transcription factor CDX2 in colonic cells uncovered highly significant overrepresentation of sequences that bind TCF4, a transcriptional effector of intestinal Wnt signaling. Chromatin immunoprecipitation confirmed TCF4 occupancy at most such sites and co-occupancy of CDX2 and TCF4 across short distances. A region spanning the single nucleotide polymorphism rs6983267, which lies within a MYC enhancer and confers colorectal cancer risk in humans, represented one of many co-occupied sites. Co-occupancy correlated with intestine-specific gene expression and CDX2 loss reduced TCF4 binding. These results implicate CDX2 in directing TCF4 binding in intestinal cells. Co-occupancy of regulatory regions by signal-effector and tissue-restricted transcription factors may represent a general mechanism for ubiquitous signaling pathways to achieve tissue-specific outcomes.

Essential and Redundant Functions of Caudal Family Proteins in Activating Adult Intestinal Genes

ABSTRACT Transcription factors that potently induce cell fate often remain expressed in the induced organ throughout life, but their requirements in adults are uncertain and varied. Mechanistically, it is unclear if they activate only tissue-specific genes or also directly repress heterologous genes. We conditionally inactivated mouse Cdx2, a dominant regulator of intestinal development, and mapped its genome occupancy in adult intestinal villi. Although homeotic transformation, observed in Cdx2-null embryos, was absent in mutant adults, gene expression and cell morphology were vitally compromised. Lethality was significantly accelerated in mice lacking both Cdx2 and its homolog Cdx1, with particular exaggeration of defects in villus enterocyte differentiation. Importantly, Cdx2 occupancy correlated with hundreds of transcripts that fell but not with equal numbers that rose with Cdx loss, indicating a predominantly activating role at intestinal cis-regulatory regions. Integrated consideration of a transcription factors mutant phenotype and cistrome hence reveals the continued and distinct requirement in adults of a critical developmental regulator that activates tissue-specific genes.

Intestinal Master Transcription Factor CDX2 Controls Chromatin Access for Partner Transcription Factor Binding

ABSTRACT Tissue-specific gene expression requires modulation of nucleosomes, allowing transcription factors to occupy cis elements that are accessible only in selected tissues. Master transcription factors control cell-specific genes and define cellular identities, but it is unclear if they possess special abilities to regulate cell-specific chromatin and if such abilities might underlie lineage determination and maintenance. One prevailing view is that several transcription factors enable chromatin access in combination. The homeodomain protein CDX2 specifies the embryonic intestinal epithelium, through unknown mechanisms, and partners with transcription factors such as HNF4A in the adult intestine. We examined enhancer chromatin and gene expression following Cdx2 or Hnf4a excision in mouse intestines. HNF4A loss did not affect CDX2 binding or chromatin, whereas CDX2 depletion modified chromatin significantly at CDX2-bound enhancers, disrupted HNF4A occupancy, and abrogated expression of neighboring genes. Thus, CDX2 maintains transcription-permissive chromatin, illustrating a powerful and dominant effect on enhancer configuration in an adult tissue. Similar, hierarchical control of cell-specific chromatin states is probably a general property of master transcription factors.