Microfluidic concatemer sequencing unveils cooperative and competitive enhancer clusters in gene regulation

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ID: 339308
2026
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Abstract
Chromatin looping plays a pivotal role in eukaryotic gene regulation; however, technical limitations have hindered the comprehensive characterization of higher-order interactions among genomic loci. As a result, key questions, such as whether enhancers function cooperatively or competitively at the single-molecule and ensemble levels, remain largely unresolved. Here, we present Drop-t, a novel microfluidics-based method that enables systematic profiling of higher-order chromatin architecture at both the single-molecule and population scales. We demonstrate that Drop-t is not only resistant to common ligation-induced DNA entanglements, which frequently cause nanopore clogging, but also provides greater relevance for elucidating regulatory mechanisms compared to existing ligation-free approaches. Using Drop-t, we uncovered extensive, previously underappreciated non-specific enhancer-promoter interactions (EPIs) and showed that these may influence transcriptional variability by competing with specific EPIs. Notably, we identified two major gene classes, dictator genes and leader genes, based on whether their promoters are predominantly regulated by competitive or cooperative specific EPIs, respectively. Together, our findings established Drop-t as a powerful tool for decoding the higher-order organizational principles of chromatin architecture and their impact on transcriptional regulation and variability.
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Authors Qiwen Chen, Shiqi Zheng, Feifei Li, Yue Shi, Zijian Li, Xiaolu Li, Junmei Wang, Bingxiang Xu, Xiaoli Li, Qing Zhang, Meizhen Zheng
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Journal Nucleic Acids Research
Year 2026
DOI
10.1093/nar/gkag1146
URL
https://doi.org/10.1093/nar/gkag1146
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