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Genome-wide functional assessment of enhancer activities in the human genome

ABSTRACT: We illustrate that modifications to the previously published STARR-seq protocols are capable of assessing enhancer activity of whole genomes on mammalian systems using the human prostate cancer cell line, LNCaP. Besides detecting accessible enhancers located in open chromatin regions, we are also able to discover active, inaccessible enhancers in closed chromatin regions. By applying histone deacetylase inhibitor, trichostatin A, genes nearby the previously inaccessible enhancers are up-regulated, indicating the potential for endogenous functionality of these regulatory elements. These results exemplify how whole genome STARR-seq provides an improved approach to gain a better understanding of the intricacies of transcriptional regulation of high complexity mammalian genomes.

ORGANISM(S): Homo sapiens

PROVIDER: GSE82204 | GEO | 2017/03/02

SECONDARY ACCESSION(S): PRJNA324294

REPOSITORIES: GEO

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Genome-wide quantitative enhancer activity maps identified by STARR-seq

Project description:Genomic enhancers are important regulators of gene expression, but their identification is a challenge and methods depend on indirect measures of activity. We developed a method termed STARR-seq to directly and quantitatively assess enhancer activity for millions of candidates from arbitrary sources of DNA, enabling screens across entire genomes. When applied to the Drosophila genome, STARR-seq identifies thousands of cell type-specific enhancers across a broad continuum of strengths, linking differential gene expression to differences in enhancer activity and creating a genome-wide quantitative enhancer map. This map reveals the highly complex regulation of transcription, with several independent enhancers for both developmental regulators and ubiquitously expressed genes. STARR-seq can be used to identify and quantitate enhancer activity in other eukaryotes, including human. STARR-seq was performed in S2 and OSC cells with paired-end sequencing in two replicates and respective inputs. DHS-seq was done with single-end sequencing in two replicates for S2 and OSC cells. RNA-seq was performed with a strand-specific protocol using single-end sequencing in two replicates within S2 and OSC cells. STARR-seq was also performed in HeLa cells with single-end sequencing with a respective input.

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Global Quantitative Mapping of Enhancer Activity in Rice Genome by STARR-seq

Project description:Enhancers are important regulators of gene expression, but their identification is a challenge in plants. STARR-seq is a method measuring directly the enhancer activity of millions fragments in parallel, which had been successfully used to identify enhancers in Drosophila and human genomes. Here we present a global map of rice enhancers whose activities are quantitatively determined by STARR-seq.We also predicted intergenic enhancers based on DNase I hypersensitivity as described in a previously published work. Predicted enhancers overlap poorly with STARR-seq enhancers, only about 400 sites accounting for 3-4% of total enhancers identified by these two different methods. In summary, our results of STARR-seq reveal that enhancers in a plant genome differ from animal enhancers in several aspects and provide a regulatory element resource for further functional and mechanistic studies in different contexts

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Genome-wide quantitative enhancer activity maps identified by STARR-seq

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Resolving systematic errors in widely-used enhancer activity assays in human cells enables genome-wide functional enhancer characterization.

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Project description:Phenotypic differences between closely related species are thought to arise primarily from changes in gene expression due to mutations in cis-regulatory sequences (enhancers). However, it has remained unclear, how frequently mutations alter enhancer activity or create functional enhancers de novo. Here, we use STARR-seq, a recently developed quantitative enhancer assay, to determine genome-wide enhancer activity profiles for five Drosophila species in the constant trans-regulatory environment of D. melanogaster S2 cells. We find that the function of a large fraction of D. melanogaster enhancers is conserved in their orthologous sequences due to selection and stabilizing turnover of transcription factor motifs. Moreover, hundreds of enhancers have been gained since the D. melanogaster M-bM-^@M-^S D. yakuba split about 11 million years ago without apparent adaptive selection and can contribute to gene expression changes in vivo. Our finding that enhancer activity is often deeply conserved and frequently gained provides important functional insights into regulatory evolution. STARR-seq was performed in S2 cells with paired-end sequencing in two replicates and respective inputs using genomic DNA from different Drosophila species. RNA-seq was performed in a non-stranded manner without replicates for two Drosophila species.

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Cis-regulatory atlas of primary human CD4+ T cells

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CRISPR Inversion of CTCF Sites Alters Genome Topology and Enhancer/Promoter Function

Project description:CTCF/cohesin play a central role in insulator function and higher-order chromatin organization of mammalian genomes. Recent studies identified a correlation between the orientation of CTCF-binding sites (CBSs) and chromatin loops. To test the functional significance of this observation, we combined CRISPR/Cas9-based genomic-DNA-fragment editing with chromosome-conformation-capture experiments to show that the location and relative orientations of CBSs determine the specificity of long-range chromatin looping in mammalian genomes, using protocadherin (Pcdh) and β-globin as model genes. Inversion of CBS elements within the Pcdh enhancer reconfigures the topology of chromatin loops between the distal enhancer and target promoters, and alters gene-expression patterns. Thus, although enhancers can function in an orientation-independent manner in reporter assays, in the native chromosome context the orientation of at least some enhancers carrying CBSs can determine both the architecture of topological chromatin domains and enhancer/promoter specificity. The findings reveal how 3D chromosome architecture can be encoded by genome sequence.

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