Functional genomic approaches to elucidate the role of enhancers during development

Abstract

Successful development depends on the precise tissue-specific regulation of genes by enhancers, genetic elements that act as switches to control when and where genes are expressed. Because enhancers are critical for development, and the majority of disease-associated mutations reside within enhancers, it is essential to understand which sequences within enhancers are important for function. Advances in sequencing technology have enabled the rapid generation of genomic data that predict putative active enhancers, but functionally validating these sequences at scale remains a fundamental challenge. Herein, we discuss the power of genome-wide strategies used to identify candidate enhancers, and also highlight limitations and misconceptions that have arisen from these data. We discuss the use of massively parallel reporter assays to test enhancers for function at scale. We also review recent advances in our ability to study gene regulation during development, including CRISPR-based tools to manipulate genomes and single-cell transcriptomics to finely map gene expression. Finally, we look ahead to a synthesis of complementary genomic approaches that will advance our understanding of enhancer function during development.

In search of lost time: Enhancers as modulators of timing in lymphocyte development and differentiation

“…Such studies have mostly involved analysis of individual candidate elements through generation of mice with targeted enhancer deletions. Cas9‐based approaches, 136,137 by enabling disruption of genomic elements in primary cells, 138,139 could enable a more high‐throughput analysis. Ultimately, to conclusively demonstrate that an enhancer acts in cis to generate an activation delay, it will be necessary to make time course measurements of gene activation at the level of single alleles in single cells.…”

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

“…ChIP-seq for the proteins or modifications exhibiting the strongest association with enhancers are RNAPII, the histone acetyltransferase EP300, H3K4me1, H3K27ac, and lineage-specific transcription factors (Myers et al 2011;Ryan and Farley 2020). RNAPII ChIP-seq primarily identifies poised or transcribed genes, but active enhancers are also marked by RNAPII peaks (Li et al 2012;Mitchell et al 2012).…”

Comprehensive multi-omics integration identifies differentially active enhancers during human brain development with clinical relevance

“…Large scale community driven efforts [7][8][9][10][11] and a plethora of individual studies have produced a vast amount of epigenome data sets, such as profiles of histone modifications, chromatin accessibility and chromatin interactions for different human tissues and cell types, that can be used to predict putative enhancers at large scale. More recently, new technologies such as massively parallel reporter assays and CRISPR-Cas9 based screens have entered the stage [12][13][14], providing novel means to directly test the functionality of non-coding regions. In addition, computational prediction algorithms [15,16], trained on epigenome and experimental data are improving the capability to predict functional sequences and the effects of variants in these regions.…”