Dynamic network-guided CRISPRi screen identifies CTCF-loop-constrained nonlinear enhancer gene regulatory activity during cell state transitions

Luo, Renhe; Yan, Jielin; Engreitz, J; Wang, Xi; Shigaki, Dustin; Wong, Wilfred Hing Sang; Cho, H S; Murphy, Dylan; Cutler, Ronald R.; Rosen, B.P.; Pulecio, Julián; Yang, Dapeng; Glenn, Rachel A.; Chen, Tingxu; Li, Qing V.; Vierbuchen, Thomas; Apostolou, Effie; Huangfu, Danwei; Beer, M

doi:10.1038/s41588-023-01450-7

Cited by 13 publications

(14 citation statements)

References 67 publications

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“…Taken together, this data confirms that in isolation, priming elements may have little enhancer activity by themselves, but can in some cases acquire high stimulatory activity at later developmental stages by binding additional TFs or by being repressed, indicating that a larger network of factors is modulating enhancer activity. Our data therefore provide a molecular explanation for the cell-stage specific sensitivity of gene expression to perturbation of indivdual enhancers within individual gene loci (Luo et al, 2023).…”

Section: Resultsmentioning

confidence: 80%

Chromatin priming elements direct tissue-specific gene activity prior to hematopoietic specification

Maytum,

Edginton-White,

Keane

et al. 2023

Preprint

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Tissue-specific gene regulation during development involves the interplay between transcription factors and epigenetic regulators binding to enhancer and promoter elements. The pattern of active enhancers defines the cellular differentiation state. However, developmental gene activation involves a previous step called chromatin priming which is not fully understood. We recently developed a genome-wide functional assay that allowed us to functionally identify enhancer elements integrated in chromatin regulating each of five stages spanning the in vitro differentiation of embryonic stem cells to blood. We also measured global chromatin accessibility, histone modifications and transcription factor binding. The integration of these data identified and characterised cis-regulatory elements which become activated prior to the onset of gene expression, some of which are primed in a signalling-dependent fashion. Deletion of such a priming element leads to a delay in the upregulation of its associated gene in development. Our work uncovers the details of a complex network of regulatory interactions with the dynamics of early chromatin opening being at the heart of dynamic tissue-specific gene expression control.

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Section: Resultsmentioning

confidence: 80%

Chromatin priming elements direct tissue-specific gene activity prior to hematopoietic specification

Maytum,

Edginton-White,

Keane

et al. 2023

Preprint

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“…Promoters showed consistently high conservation rate across the 45 tissues (mean 67%) while enhancers showed highly variable conservation rate, ranging from 6.7% (embryonic liver) to 31% (embryonic brain) ( Fig 2A ). Such lack of conservation is not observed in CTCF ChIP-seq peaks, which are also often distal to TSSs (conservation rate for brain, heart, B-cell, ESC = 82%, 79%, 72%, 76%; Fig 2B ), suggesting that many CTCF loops and topologically associated domains are conserved 14,40 . The strong and somewhat counter-intuitive tissue specificity of enhancer conservation will be explored extensively below.…”

Section: Resultsmentioning

confidence: 92%

“…Among these top orthologous enhancers, we selected a subset of enhancers in the vicinity of orthologous genes with cell/tissue specific expression 55 ( Fig 5D ; Supp Fig 13 ). These genes include KCND3 (brain; voltage-gated potassium channel subunit), PAX2 (kidney; TF associated with renal malformation 59 ), GATA6 (stomach; definite endoderm TF 14 ), MYOD1(muscle; TF associated with myopathy 60 ), and IKZF3 (B-cell; TF mutated in leukemia 61,62 ). For each of the 45 cell/tissue pairs, we generated a table of ranked orthologous human-to-mouse enhancer pairs.…”

Section: Resultsmentioning

confidence: 99%

“…OTUD7A knockout leads to morphological deformation of cortical neurons and frequent seizure-like events in mice 57,58 . We identified orthologous pairs of putative OTUD7A enhancers in intron 1 of human and (stomach; definite endoderm TF 14 ), MYOD1(muscle; TF associated with myopathy 60 ), and IKZF3 (Bcell; TF mutated in leukemia 61,62 ). For each of the 45 cell/tissue pairs, we generated a table of ranked orthologous human-to-mouse enhancer pairs.…”

Section: F; Methods)mentioning

confidence: 99%

“…Further, duplication of redundant TFBS and accumulation of enhancer mutations can lead to turnover of TFBS through stabilizing selection 11 . This flexibility also applies on a larger scale to combinations of enhancers within intergenic loci, as enhancer function is weakly constrained by position relative to the target promoter, and enhancers are often accompanied by redundant shadow enhancers that regulate the same target genes 12–14 . As a result, the functional and mechanistic properties of enhancers likely have facilitated their rapid turnover throughout evolutionary history while maintaining DNA binding specificities of TFs 8,15 .…”

Section: Introductionmentioning

confidence: 99%

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Gapped-kmer sequence modeling robustly identifies regulatory vocabularies and distal enhancers conserved between evolutionarily distant mammals

Oh,

Beer

2023

Preprint

Self Cite

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Gene regulatory elements drive many complex biological phenomena such as fetal development, and their mutations are linked to a multitude of common human diseases. The phenotypic impacts of regulatory variants are often tested using their conserved orthologous counterparts in model organisms such as mice. However, mapping human enhancers to conserved elements in mice remains a challenge, due to both rapid evolution of enhancers and limitations of current computational methods to detect conserved regulatory sequences. To improve upon existing computational methods and to better understand the sources of this apparent regulatory divergence, we comprehensively measured the evolutionary dynamics of distal enhancers across 45 matched human/mouse cell/tissue pairs from more than 1,000 DNase-seq experiments. Using this expansive dataset, we show that while cell-specific regulatory vocabulary is conserved, enhancers evolve more rapidly than other genomic elements such as promoters and CTCF binding sites. We observed surprisingly high levels of cell-specific variability in enhancer conservation rates, in part explainable by tissue specific transposable element activity. To improve orthologous enhancer mapping, we developed an improved genome alignment algorithm using gapped-kmer sequence features, and using the matched cell/tissue pairs, we show that this novel computational method,gkm-align, discovers 23,660 novel human/mouse conserved enhancers missed by standard alignment algorithms.

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CRISPR screening uncovers a long-range enhancer for ONECUT1 in pancreatic differentiation and links a diabetes risk variant

Kaplan,

Wong,

Yan

et al. 2024

Cell Reports

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Dynamic network-guided CRISPRi screen identifies CTCF-loop-constrained nonlinear enhancer gene regulatory activity during cell state transitions

Cited by 13 publications

References 67 publications

Chromatin priming elements direct tissue-specific gene activity prior to hematopoietic specification

Chromatin priming elements direct tissue-specific gene activity prior to hematopoietic specification

Gapped-kmer sequence modeling robustly identifies regulatory vocabularies and distal enhancers conserved between evolutionarily distant mammals

CRISPR screening uncovers a long-range enhancer for ONECUT1 in pancreatic differentiation and links a diabetes risk variant

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