Enhancer identification in mouse embryonic stem cells using integrative modeling of chromatin and genomic features
BMC Genomics
Enhancer identification in mouse embryonic stem cells using integrative modeling of chromatin and genomic features
Chih-yu Chen 0
Quaid Morris 1
Jennifer A Mitchell 0
0 Department of Cell and Systems Biology, University of Toronto , 25 Harbord Street, Toronto, ON, M5S 3G5 , Canada
1 Donnelly Centre for Cellular and Biomolecular Research, University of Toronto , Toronto , Canada
Background: Epigenetic modifications, transcription factor (TF) availability and differences in chromatin folding influence how the genome is interpreted by the transcriptional machinery responsible for gene expression. Enhancers buried in non-coding regions are found to be associated with significant differences in histone marks between different cell types. In contrast, gene promoters show more uniform modifications across cell types. Here we used histone modification and chromatin-associated protein ChIP-Seq data sets in mouse embryonic stem (ES) cells as well as genomic features to identify functional enhancer regions. Using co-bound sites of OCT4, SOX2 and NANOG (co-OSN, validated enhancers) and co-bound sites of MYC and MYCN (limited enhancer activity) as enhancer positive and negative training sets, we performed multinomial logistic regression with LASSO regularization to identify key features. Results: Cross validations reveal that a combination of p300, H3K4me1, MED12 and NIPBL features to be top signatures of co-OSN regions. Using a model from 10 signatures, 83% of top 1277 putative 1 kb enhancer regions (probability greater than or equal to 0.8) overlapped with at least one TF peak from 7 mouse ES cell ChIP-Seq data sets. These putative enhancers are associated with increased gene expression of neighbouring genes and significantly enriched in multiple TF bound loci in agreement with combinatorial models of TF binding. Furthermore, we identified several motifs of known TFs significantly enriched in putative enhancer regions compared to random promoter regions and background. Comparison with an active H3K27ac mark in various cell types confirmed cell type-specificity of these enhancers. Conclusions: The top enhancer signatures we identified (p300, H3K4me1, MED12 and NIPBL) will allow for the identification of cell type-specific enhancer regions in diverse cell types.
Enhancer; Embryonic stem cells; Transcription factor; ChIP-Seq; Histone methylation; Regulation of gene expression
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Background
Chromatin immunoprecipitation followed by massively
parallel sequencing (ChIP-Seq) has enabled
genomewide investigation of chromatin features and epigenetic
modifications within the non-coding regions of
mammalian genomes in high resolution [1]. ChIP-Seq provides the
opportunity to characterise and begin to understand on
a genome-wide scale how genes are regulated in a
celltype specific manner by sequence-specific DNA-binding
transcription factors (TFs). However, identifying regulatory
regions within the genome and linking these regions to the
regulation of specific genes remains a challenge.
Distal regulatory elements have been identified which
regulate gene transcription from several kilobases (kb)
away and have even been found to regulate genes located
on separate chromosomes [2-4]. Functional
characterisation of these regulatory elements can be done by
identifying bound TFs and investigating whether or not they act as
enhancers, increasing transcription of a gene in a position
and orientation independent manner. ChIP-Seq analysis for
several TFs has revealed a significant fraction (4060%) of
the binding sites for most TFs are located in intergenic
regions >10 kb from transcription start sites (TSSs) of
annotated genes [5-7]. In addition, enhancer regions are
associated with significant epigenetic differences between
cell types, while gene promoters show more uniform
modifications across different cell types [8,9]. These
findings suggest that enhancers, which can be located at great
distances from the genes they regulate, play a larger
role in regulating tissue-specific gene expression than
the sequences proximal to gene promoters. Moreover,
mutations in DNA sequences of distant-acting
enhancers contribute to various diseases [10], further stressing
their importance in regulating gene expression.
Prior to the availability of ChIP-Seq and ChIP-chip data,
computational approaches based solely on genomic
sequences were used to identify enhancer regions. Initially
these approaches compared the genomic sequence with
TF binding motifs represented by position specific scoring
matrices (PSSM) from TRANSFAC [11] and JASPAR [12].
TF motif clustering and comparative genomics improved
the predictive power of these approaches [13-16]. In
addition, intergenic regions with high sequence
conservation between human and Fugu or ultra-conserved regions
between human-mouse-rat (>200 bp of 100% identity) are
predictive of regulatory regions involved in conserved
processes such as embryonic development [17,18]. As
many enhancer regio (...truncated)