CRISPR-Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genome
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ABSTRACT:
PROVIDER: PRJNA379971 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR5364850_1.fastq.gz | Fastqsanger.gz | |||
| SRR5364850_2.fastq.gz | Fastqsanger.gz | |||
| SRR5364860_1.fastq.gz | Fastqsanger.gz | |||
| SRR5364860_2.fastq.gz | Fastqsanger.gz | |||
| SRR5364870_1.fastq.gz | Fastqsanger.gz |
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Project description:This SuperSeries is composed of the SubSeries listed below. Large genome mapping consortia and thousands of genome-wide association studies have identified non-protein coding elements in the genome as a having a central role in tissue development, cell-type specification, response to environmental or pharmacologic signals, and susceptibility to most common diseases. However, decoding the function of the millions of putative regulatory elements discovered in these studies remains a primary challenge. New CRISPR/Cas9-based epigenome editing technologies have enabled the precise perturbation of the activity of specific regulatory elements. Here we describe CRISPR/Cas9-based Epigenomic Regulatory Element Screening (CERES) for high-throughput screening of regulatory element activity within the native genomic context. We perform both loss- and gain-of-function screens with complementary epigenome editing tools to identify known and unknown regulatory elements of medically relevant genes in human cells. The high-throughput functional annotation of putative regulatory elements by CERES constitutes a new platform for screening biological mechanisms that cannot be perturbed by traditional methods.
2017-03-23 | GSE96876 | GEO
Project description:Large genome mapping consortia and thousands of genome-wide association studies have identified non-protein coding elements in the genome as a having a central role in tissue development, cell-type specification, response to environmental or pharmacologic signals, and susceptibility to most common diseases. However, decoding the function of the millions of putative regulatory elements discovered in these studies remains a primary challenge. New CRISPR/Cas9-based epigenome editing technologies have enabled the precise perturbation of the activity of specific regulatory elements. Here we describe CRISPR/Cas9-based Epigenomic Regulatory Element Screening (CERES) for high-throughput screening of regulatory element activity within the native genomic context. We perform both loss- and gain-of-function screens with complementary epigenome editing tools to identify known and unknown regulatory elements of medically relevant genes in human cells. The high-throughput functional annotation of putative regulatory elements by CERES constitutes a new platform for screening biological mechanisms that cannot be perturbed by traditional methods.
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Project description:Large genome mapping consortia and thousands of genome-wide association studies have identified non-protein coding elements in the genome as a having a central role in tissue development, cell-type specification, response to environmental or pharmacologic signals, and susceptibility to most common diseases. However, decoding the function of the millions of putative regulatory elements discovered in these studies remains a primary challenge. New CRISPR/Cas9-based epigenome editing technologies have enabled the precise perturbation of the activity of specific regulatory elements. Here we describe CRISPR/Cas9-based Epigenomic Regulatory Element Screening (CERES) for high-throughput screening of regulatory element activity within the native genomic context. We perform both loss- and gain-of-function screens with complementary epigenome editing tools to identify known and unknown regulatory elements of medically relevant genes in human cells. The high-throughput functional annotation of putative regulatory elements by CERES constitutes a new platform for screening biological mechanisms that cannot be perturbed by traditional methods.
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Project description:Large genome-mapping consortia and thousands of genome-wide association studies have identified non-protein-coding elements in the genome as having a central role in various biological processes. However, decoding the functions of the millions of putative regulatory elements discovered in these studies remains challenging. CRISPR-Cas9-based epigenome editing technologies have enabled precise perturbation of the activity of specific regulatory elements. Here we describe CRISPR-Cas9-based epigenomic regulatory element screening (CERES) for improved high-throughput screening of regulatory element activity in the native genomic context. Using dCas9KRAB repressor and dCas9p300 activator constructs and lentiviral single guide RNA libraries to target DNase I hypersensitive sites surrounding a gene of interest, we carried out both loss- and gain-of-function screens to identify regulatory elements for the ?-globin and HER2 loci in human cells. CERES readily identified known and previously unidentified regulatory elements, some of which were dependent on cell type or direction of perturbation. This technology allows the high-throughput functional annotation of putative regulatory elements in their native chromosomal context.
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Project description:Epigenome editing with the CRISPR/Cas9 platform is a promising technology to modulate gene expression to direct cell phenotype and to dissect the causal epigenetic mechanisms that direct gene regulation. Fusions of the nuclease-inactive dCas9 to the KRAB repressor domain (dCas9-KRAB) can effectively silence target gene expression. We targeted dCas9-KRAB to the HS2 enhancer, a distal regulatory element that orchestrates the expression of multiple globin genes. Genome-wide analyses demonstrated that localization of dCas9-KRAB to HS2 specifically induced H3K9 tri-methylation (H3K9me3) at that enhancer and reduced the chromatin accessibility of both the enhancer and its promoter targets. Targeted epigenetic modification of HS2 silenced the expression of multiple globin genes, with minimal off-target changes in gene expression. These results demonstrate that repression mediated by dCas9-KRAB is sufficiently specific to disrupt the activity of individual enhancers via local modification of the epigenome. This approach enables precise modulation of epigenetic function without modifying the underlying genome sequence.
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Project description:Epigenetic modifications determine the structure and regulation of eukaryotic genomes and define key signatures of cell lineage specification. Technologies that facilitate the targeted manipulation of epigenetic marks could be used to precisely control cell phenotype or interrogate the relationship between the epigenome and transcriptional control. Here we have generated a programmable acetyltransferase based on the CRISPR/Cas9 gene regulation system, consisting of the nuclease-null dCas9 protein fused to the catalytic core of the human acetyltransferase p300. This fusion protein catalyzes acetylation of histone H3 lysine 27 (H3K27) at its target sites, leading to robust transcriptional activation of target genes from promoters, proximal enhancers, and distal enhancers. In contrast to conventional dCas9-based activators, the acetyltransferase fusion effectively activated genes from enhancer regions and with individual guide RNAs. The core p300 domain was also portable to other programmable DNA-binding proteins. This technology enables the targeted perturbation of native epigenetic architecture and will be useful for reprogramming the epigenome for applications in genomics, genetics, disease modeling, and manipulating cell fate.
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