Functional dissection of the enhancer repertoire in human embryonic stem cells

Enhancers are genetic elements that regulate spatiotemporal gene expression. Enhancer function requires transcription factor (TF) binding and correlates with histone modifications. However, the extent to which TF binding and histone modifications functionally define active enhancers remains unclear. Here, we combine chromatin immunoprecipitation with a massively parallel reporter assay (ChIP-STARR-seq) to identify functional enhancers in human embryonic stem cells (ESCs) genome-wide in a quantitative unbiased manner. Although active enhancers associate with TFs, only a minority of regions marked by NANOG, OCT4, H3K27ac, and H3K4me1 function as enhancers, with activity markedly changing under naive versus primed culture conditions. We identify an enhancer set associated with functions extending to non-ESC-specific processes. Moreover, although transposable elements associate with putative enhancers, only some exhibit activity. Similarly, within super-enhancers, large tracts are non-functional, with activity restricted to small sub-domains. This catalog of validated enhancers provides a valuable resource for further functional dissection of the regulatory genome.Barakat et al. use a combination of chromatin immunoprecipitation and a massively parallel reporter assay to identify functional enhancers in primed and naive human embryonic stem cells. This genome-wide catalog of validated enhancers provides a valuable resource for the further dissection of the regulatory genome

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

Abstract Background Non-coding regulatory elements (NCREs), such as enhancers, play a crucial role in gene regulation, and genetic aberrations in NCREs can lead to human disease, including brain disorders. The human brain is a complex organ that is susceptible to numerous disorders; many of these are caused by genetic changes, but a multitude remain currently unexplained. Understanding NCREs acting during brain development has the potential to shed light on previously unrecognized genetic causes of human brain disease. Despite immense community-wide efforts to understand the role of the non-coding genome and NCREs, annotating functional NCREs remains challenging. Methods Here we performed an integrative computational analysis of virtually all currently available epigenome data sets related to human fetal brain. Results Our in-depth analysis unravels 39,709 differentially active enhancers (DAEs) that show dynamic epigenomic rearrangement during early stages of human brain development, indicating likely biological function. Many of these DAEs are linked to clinically relevant genes, and functional validation of selected DAEs in cell models and zebrafish confirms their role in gene regulation. Compared to enhancers without dynamic epigenomic rearrangement, DAEs are subjected to higher sequence constraints in humans, have distinct sequence characteristics and are bound by a distinct transcription factor landscape. DAEs are enriched for GWAS loci for brain-related traits and for genetic variation found in individuals with neurodevelopmental disorders, including autism. Conclusion This compendium of high-confidence enhancers will assist in deciphering the mechanism behind developmental genetics of human brain and will be relevant to uncover missing heritability in human genetic brain disorders

Temporal and tissue-specific variability of SMN protein levels in mouse models of spinal muscular atrophy

textabstractSpinal muscular atrophy (SMA) is a progressive motor neuron disease caused by deleterious variants in SMN1 that lead to a marked decrease in survival motor neuron (SMN) protein expression. Humans have a second SMN gene (SMN2) that is almost identical to SMN1. However, due to alternative splicing the majority of SMN2 messenger ribonucleic acid (mRNA) is translated into a truncated, unstable protein that is quickly degraded. Because the presence of SMN2 provides a unique opportunity for therapy development in SMA patients, the mechanisms that regulate SMN2 splicing and mRNA expression have been elucidated in great detail. In contrast, how much SMN protein is produced at different developmental time points and in different tissues remains under-characterized. In this study, we addressed this issue by determining SMN protein expression levels at three developmental time points across six different mouse tissues and in two distinct mouse models of SMA ('severe' Taiwanese and 'intermediate' Smn2B/mice). We found that, in healthy control mice, SMN protein expression was significantly influenced by both age and tissue type.When comparing mouse models of SMA, we found that, despite being transcribed from genetically different alleles, control SMN levels were relatively similar. In contrast, the degree of SMN depletion between tissues in SMA varied substantially over time and between the two models. These findings offer an explanation for the differential vulnerability of tissues and organs observed in SMA and further our understanding of the systemic and temporal requirements for SMN with direct relevance for developing effective therapies for SMA

The functional enhancer repertoire of human embryonic stem cells

Genome binding/occupancy profiling by high throughput sequencingWe combined Self-Transcribing Active Regulatory Region Sequencing (STARR-seq) with an enrichment step using chromatin immunoprecipitation in a massively parallel reporter assay. We applied this assay, termed ChIP-STARR-seq, to normal (primed) and naive human embryonic stem cells, building up a comprehensive catalogue of functional enhancers. For further details, please refer to the sub-series.Barakat TS, Halbritter F, Zhang M, Rendeiro AF et al. Functional Dissection of the Enhancer Repertoire in Human Embryonic Stem Cells. National Center for Biotechnology Information (Gene Expression Omnibus). https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE9963

The functional enhancer repertoire of human embryonic stem cells

Genome binding/occupancy profiling by high throughput sequencin