ChIP-seq accurately predicts tissue-specific activity of enhancers

Abstract

A major yet unresolved quest in decoding the human genome is the identification of the regulatory sequences that control the spatial and temporal expression of genes. Distant-acting transcriptional enhancers are particularly challenging to uncover because they are scattered among the vast noncoding portion of the genome. Evolutionary sequence constraint can facilitate the discovery of enhancers, but fails to predict when and where they are active in vivo. Here we present the results of chromatin immunoprecipitation with the enhancer-associated protein p300 followed by massively parallel sequencing, and map several thousand in vivo binding sites of p300 in mouse embryonic forebrain, midbrain and limb tissue. We tested 86 of these sequences in a transgenic mouse assay, which in nearly all cases demonstrated reproducible enhancer activity in the tissues that were predicted by p300 binding. Our results indicate that in vivo mapping of p300 binding is a highly accurate means for identifying enhancers and their associated activities, and suggest that such data sets will be useful to study the role of tissue-specific enhancers in human biology and disease on a genome-wide scale.The initial sequencing of the human genome 1,2 , complemented by effective computational and experimental strategies for mammalian gene discovery 3,4 , has resulted in a virtually complete list of protein-coding sequences. In contrast, the genomic location and function of regulatory elements that orchestrate gene expression in the developing and adult body remain more obscure, hindering studies of their contribution to developmental processes and human disease. Evolutionary constraint of non-coding sequences can predict the location of enhancers in the genome 5-12 , but does not reveal when and where these enhancers are active in vivo. Furthermore, it has been suggested that a substantial proportion of regulatory elements is not sufficiently conserved to be detectable by comparative genomic methods [13][14][15][16] .

Epigenetic Mechanisms in Inflammation

“…Genes with bivalent modification (H3K4me3 and H3K27me3) are typically important developmental regulators in pluripotent embryonic stem (ES) cells or multipotent progenitor cells which are silenced, but are poised for activation, as differentiation proceeds (Bernstein et al, 2006;Mikkelsen et al, 2007). Enhancers, which determine tissue-specific gene expression, are marked by histone H3 lysine 4 mono-methylation (H3K4me1) and by the histone acetyltransferase co-factors CBP/p300 binding (Heintzman et al, 2007;Visel et al, 2009). Methylation of histones is carried out by histone methyltransferases (HMT) and demethylation by histone demethylases (HDM) such as members of the Jumonji protein family (Cheng and Blumenthal, 2010).…”

Application of a systems approach to study developmental gene regulation

“…ChIP-seq is a powerful technology to discover an important class of physical interactions at a genome-wide scale: protein-DNA interactions between regulatory factors and DNA. These regulatory proteins can include master regulators of embryonic stem cells (Chen et al 2008), lineagespecific factors (Nielsen et al 2008), and general enhancer-associated proteins such as p300 (Visel et al 2009). The genome-wide binding profiles of these transcription factors are usually highly specific to a tissue or a developmental stage.…”

Transcriptional control of macrophage diversity and specialization

“…Accordingly, when a putative enhancer acting in a given tissue is identified, a standard assay for the functional evaluation of its activity is to clone it into a transgenic mouse reporter system and to observe that the pattern of reporter gene expression recapitulates the anatomical location where the enhancer was identified [24]. Bringing these observations to the ultimate conclusion, not only do they imply that enhancers largely account for the expression of tissue-restricted genes, but they may also indicate that even the expression of ubiquitous or broadly expressed genes in different anatomical regions may be regulated by non-overlapping sets of enhancers selectively active in specific tissues.…”

“…An increasing number of reports have allowed the definition of a chromatin ''signature'' for enhancers, which can be located according to a high level of H3K4me1 together with a low enrichment of H3K4me3 (H3K4me1 hi /H3K4me3 lo ) [25]. Additional marks associated with active enhancers include p300 binding, acetylation of H3K27 (H3K27ac), hypersensitivity to DNAse and nucleosomal depletion [21,24,26]. Importantly, histone modifications can also discriminate between different activity states, with poised enhancers being marked by the absence of H3K27ac and inactive/ repressed enhancers by the presence of H3K27me3 [27].…”