EXTRA-seq: a genome-integrated extended massively parallel reporter assay to quantify enhancer-promoter communication DOI Creative Commons
Judith F. Kribelbauer, Vincent Gardeux, Gerard Llimos

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Dec. 11, 2024

Precise control of gene expression is essential for cellular function, but the mechanisms by which enhancers communicate with promoters to coordinate this process are not fully understood. While sequence-based deep learning models show promise in predicting enhancer-driven expression, experimental validation and human-interpretable mechanistic insights lag behind. Here, we present EXTRA-seq , a novel EXT ended R eporter A ssay followed seq uencing designed quantify enhancer activity endogenous contexts over kilobase-scale distances. We demonstrate that can be targeted disease-relevant loci captures changes at resolution individual transcription factor binding sites, enabling discovery. Using engineered synthetic enhancer-promoter combinations, reveal TATA-box acts as dynamic range amplifier, modulating levels function strength. Importantly, find integrating state-of-the-art plasmid-based assays improves prediction measured EXTRA-seq. These findings open new avenues predictive modeling therapeutic applications. Overall, our work provides powerful platform interrogate complex interplay between promoters, bridging gap silico predictions biological mechanisms.

Language: Английский

Multiplex profiling of developmental cis-regulatory elements with quantitative single-cell expression reporters DOI Creative Commons
Jean‐Benoît Lalanne, Samuel G. Regalado, Silvia Domcke

et al.

Nature Methods, Journal Year: 2024, Volume and Issue: 21(6), P. 983 - 993

Published: May 9, 2024

Abstract The inability to scalably and precisely measure the activity of developmental cis -regulatory elements (CREs) in multicellular systems is a bottleneck genomics. Here we develop dual RNA cassette that decouples detection quantification tasks inherent multiplex single-cell reporter assays. resulting measurement expression accurate over multiple orders magnitude, with precision approaching limit set by Poisson counting noise. Together barcode stabilization via circularization, these scalable quantitative reporters provide high-contrast readouts, analogous classic situ assays but entirely from sequencing. Screening >200 regions accessible chromatin vitro model early mammalian development, identify 13 (8 previously uncharacterized) autonomous cell-type-specific CREs. We further demonstrate chimeric CRE pairs generate cognate two-cell-type profiles assess gain- loss-of-function phenotypes variants perturbed transcription factor binding sites. Single-cell can be applied quantitatively characterize native, synthetic CREs at scale, high sensitivity resolution.

Language: Английский

Citations

18
Large-scale analysis of the integration of enhancer-enhancer signals by promoters DOI Creative Commons
Miguel Martinez-Ara, Federico Comoglio, Bas van Steensel

et al.

eLife, Journal Year: 2023, Volume and Issue: 12

Published: Nov. 3, 2023

Genes are often regulated by multiple enhancers. It is poorly understood how the individual enhancer activities combined to control promoter activity. Anecdotal evidence has shown that enhancers can combine sub-additively, additively, synergistically, or redundantly. However, it not clear which of these modes more frequent in mammalian genomes. Here, we systematically tested pairs activate promoters using a three-way combinatorial reporter assay mouse embryonic stem cells. By assaying about 69,000 enhancer-enhancer-promoter combinations found generally near-additively. This behaviour was conserved across seven developmental tested. Surprisingly, scale signals non-linear manner depends on strength. A housekeeping showed an overall different response pairs, and smaller dynamic range. Thus, our data indicate mostly act but transform their collective effect non-linearly.

Language: Английский

Citations

11
Mechanisms of enhancer‐driven oncogene activation DOI Creative Commons

Joyce Vriend,

Ruud Delwel, Dorien Pastoors

et al.

International Journal of Cancer, Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 24, 2025

Abstract An aggressive subtype of acute myeloid leukemia (AML) is caused by enhancer hijacking resulting in MECOM overexpression. Several chromosomal rearrangements can lead to this: the most common (inv(3)/t(3;3)) results a hijacked GATA2 enhancer, and there are several atypical involving enhancers from other hematopoietic genes. The set which be also BCL11B . Enhancer deregulation driver oncogenesis range malignancies. mechanisms observed cancer types, including TAD boundary disruptions creation de novo (super‐) enhancers, may explain overexpression or oncogenes AML without upon translocation. Gaining mechanistic insight both super‐enhancer activity critical pave way for new treatments cancers that result deregulation.

Language: Английский

Citations

0
Modelling and design of transcriptional enhancers DOI Creative Commons
Seppe De Winter, Vasileios Konstantakos, Stein Aerts

et al.

Nature Reviews Bioengineering, Journal Year: 2025, Volume and Issue: unknown

Published: Feb. 28, 2025

Language: Английский

Citations

0
The roles of enhancer, especially super-enhancer-driven genes in tumor metabolism and immunity DOI
Shouwu Guo, Lu Zhang,

Jiao Ren

et al.

International Journal of Biological Macromolecules, Journal Year: 2025, Volume and Issue: unknown, P. 142414 - 142414

Published: March 1, 2025

Language: Английский

Citations

0
A mathematical model clarifies the ABC Score formula used in enhancer-gene prediction DOI Open Access
Joseph Nasser, Kee-Myoung Nam, Jeremy Gunawardena

et al.

Published: April 11, 2025

Enhancers are discrete DNA elements that regulate the expression of eukaryotic genes. They important not only for their regulatory function, but also as loci frequently associated with disease traits. Despite significance, our conceptual understanding how enhancers work remains limited. CRISPR-interference methods have recently provided means to systematically screen in cell culture, from which a formula predicting whether an enhancer regulates gene, Activity-by-Contact (ABC) Score, has emerged and been widely adopted. While useful binary classifier, it is less effective at quantitative effect on gene expression. It unclear algebraic form ABC Score arises underlying molecular mechanisms what assumptions needed hold. Here, we use graph-theoretic linear framework, previously introduced analyze regulation, formulate default model , mathematical multiple independently gene. We show this model. However, imply act additively steady-state This known be false certain genes modifying can accommodate discrepancy. Overall, approach lays rigorous, biophysical foundation future studies enhancer-gene regulation.

Language: Английский

Citations

0
A mathematical model clarifies the ABC Score formula used in enhancer-gene prediction DOI Open Access
Joseph Nasser, Kee-Myoung Nam, Jeremy Gunawardena

et al.

Published: April 11, 2025

Enhancers are discrete DNA elements that regulate the expression of eukaryotic genes. They important not only for their regulatory function, but also as loci frequently associated with disease traits. Despite significance, our conceptual understanding how enhancers work remains limited. CRISPR-interference methods have recently provided means to systematically screen in cell culture, from which a formula predicting whether an enhancer regulates gene, Activity-by-Contact (ABC) Score, has emerged and been widely adopted. While useful binary classifier, it is less effective at quantitative effect on gene expression. It unclear algebraic form ABC Score arises underlying molecular mechanisms what assumptions needed hold. Here, we use graph-theoretic linear framework, previously introduced analyze regulation, formulate default model , mathematical multiple independently gene. We show this model. However, imply act additively steady-state This known be false certain genes modifying can accommodate discrepancy. Overall, approach lays rigorous, biophysical foundation future studies enhancer-gene regulation.

Language: Английский

Citations

0
Cis-Regulation of the CFTR Gene in Pancreatic Cells DOI Open Access

Clara Blotas,

Anaïs Le Nabec,

Mégane Collobert

et al.

International Journal of Molecular Sciences, Journal Year: 2025, Volume and Issue: 26(8), P. 3788 - 3788

Published: April 17, 2025

Genome organization is essential for precise spatial and temporal gene expression relies on interactions between promoters distal cis-regulatory elements (CREs), which constitute ~8% of the human genome. For cystic fibrosis transmembrane conductance regulator (CFTR) gene, tissue-specific expression, especially in pancreas, remains poorly understood. Unraveling its regulation could clarify clinical heterogeneity observed CFTR-related disorders. To understand role 3D chromatin architecture establishing CFTR we mapped epigenomic Capan-1 pancreatic cells. Candidate CREs are validated by luciferase reporter assay CRISPR knock-out. We identified active not only around but also outside topologically associating domain (TAD). demonstrate involvement multiple upstream downstream reveal a cooperative effect −44 kb, −35 +15.6 +37.7 kb regions, share common predicted transcription factor (TF) motifs. extend our analysis to compare conformation intestinal cells, providing valuable insights into tissue specificity regulating expression.

Language: Английский

Citations

0
Bridging bioengineering and epigenetics: from technical innovations to clinical applications DOI Creative Commons

Jim Jacob,

Quint van Loosen,

Anna C. H. van den Berg van Saparoea

et al.

Epigenetics Communications, Journal Year: 2024, Volume and Issue: 4(1)

Published: Dec. 24, 2024

The 7th International Conference on Epigenetics & Bioengineering held in Amsterdam, Netherlands was a successful event covering cutting-edge research utilizing innovative technologies from multidisciplinary international scientists the fields of epigenetics and bioengineering, with an emphasis development disease. This conference report highlights outstanding presented engaging discussions that took place. Throughout sessions, leading experts demonstrated novel to explore epigenetic mechanisms, including advanced data analysis pipelines bioengineered systems. Several speakers uncovered emerging fundamental principles how these insights are being applied address (bio)medical challenges, underscoring field is progressing toward (pre)clinical applications targeted therapies. featured stimulating causal relationship between marks transcription, emphasizing importance standardizing editing methodologies.

Language: Английский

Citations

1
A mathematical model clarifies the ABC Score formula used in enhancer-gene prediction DOI Creative Commons
Joseph Nasser, Kee-Myoung Nam, Jeremy Gunawardena

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: Dec. 3, 2024

Enhancers are discrete DNA elements that regulate the expression of eukaryotic genes. They important not only for their regulatory function, but also as loci frequently associated with disease traits. Despite significance, our conceptual understanding how enhancers work remains limited. CRISPR-interference methods have recently provided means to systematically screen in cell culture, from which a formula predicting whether an enhancer regulates gene, Activity-by-Contact (ABC) Score, has emerged and been widely adopted. While useful binary classifier, it is less effective at quantitative effect on gene expression. It unclear algebraic form ABC Score arises underlying molecular mechanisms what assumptions needed hold. Here, we use graph-theoretic linear framework, previously introduced analyze regulation, formulate

Language: Английский

Citations

0