Transcriptional enhancers: from prediction to functional assessment on a genome-wide scale

Abstract

Enhancers are cis-regulatory sequences located distally to target genes. These sequences consolidate developmental and environmental cues to coordinate gene expression in a tissue-specific manner. Enhancer function and tissue specificity depend on the expressed set of transcription factors, which recognize binding sites and recruit cofactors that regulate local chromatin organization and gene transcription. Unlike other genomic elements, enhancers are challenging to identify because they function independently of orientation, are often distant from their promoters, have poorly defined boundaries and display no reading frame. In addition, there are no defined genetic or epigenetic features that are unambiguously associated with enhancer activity. Over recent years there have been developments in both empirical assays and computational methods for enhancer prediction. We review genome-wide tools, CRISPR advancements and high-throughput screening approaches that have improved our ability to both observe and manipulate enhancers in vitro at the level of primary genetic sequences, chromatin states and spatial interactions. We also highlight contemporary animal models and their importance to enhancer validation. Together, these experimental systems and techniques complement one another and broaden our understanding of enhancer function in development, evolution, and disease.

Regulatory SNPs: Altered Transcription Factor Binding Sites Implicated in Complex Traits and Diseases

“…The corresponding information encoded in regulatory DNA is actuated via the combinatorial binding of sequence-specific TFs to regulatory regions (cis-regulatory modules, CRMs). CRMs switch on promoters and enhancers and are actually the assemblies of TFBSs arranged to provide particular functions [ 10 , 11 , 14 , 176 , 177 , 178 ].…”

“…The two main methods mentioned above dating back to the beginning of the history of rSNP research—EMSA and reporter gene assay—still remain a golden standard in this area and are widely used in the state-of-the-art studies at the first stage of analysis because they allow the presence of a regulatory potential of a nucleotide substitution to be asserted. However, the expressed sets of TFs in different tissues are significantly different [ 11 ]; correspondingly, it is most desirable in such experiments to use several cell lines [ 3 , 23 , 51 ]. It is especially important that the EMSA with specific antibodies or purified TFs is able to reliably identify the TF with its binding site affected by a nucleotide substitution [ 23 , 28 , 30 , 40 , 41 ] and numerous other papers ( Table 1 ).…”

“…As is known, the regulatory regions of the genome represent clusters of the binding sites for sequence-specific transcription factors (TFs). There, the interplay between these TFs and their binding sites ( cis -regulatory elements) as well as the interaction of TFs with one another and the coactivator and chromatin remodeling complexes orchestrate the dynamic and diverse genetic programs, thereby determining the tissue-specific gene expression, spatiotemporal specificity of gene activities during development, and the ability of genes to respond to different external signals [ 7 , 8 , 9 , 10 , 11 , 12 ]. Thus, thanks to the binding to their specific sites on DNA (transcription factor binding sites, TFBSs), TFs directly interpret the regulatory part of the genome, performing the first step in deciphering the DNA sequence [ 13 , 14 , 15 ].…”

Epigenetic reprogramming of a distal developmental enhancer cluster drivesSOX2overexpression in breast and lung cancer

“…Although reporter assays can be used to assess enhancer activity, enhancer knockout approaches remain the current gold standard method for enhancer validation 95,96 . To investigate whether the SRR124-134 cluster drives SOX2 expression in cancer cells, we used CRISPR/Cas9 to delete this cluster from the H520, MCF-7, PC-9, and (which was not certified by peer review) is the author/funder.…”

Quo vadis PGRMC? Grand-Scale Biology in Human Health and Disease

“…Therefore, histone methylases and deacetylases play critical roles in transcriptional regulation [518]. Active enhancers bind transcription factors with transactivation domains which recruit co-activators such as transcriptional activator CREB binding protein (CBP) or P300 to in turn recruit RNA polymerase II [519,520]. In this respect, the PGRMC1-modulated NAD/1-MNA pathway (Fig.…”