Paramo Mauricio I, Leung Alden King-Yung, Shah Sagar R, Zhang Junke, Tippens Nathaniel D, Liang Jin, Yao Li, Jin Yiyang, Pan Xiuqi, Ozer Abdullah, Lis John T, Yu Haiyuan
Department of Molecular Biology and Genetics, Cornell University; Ithaca, NY, USA.
Weill Institute for Cell and Molecular Biology, Cornell University; Ithaca, NY, USA.
Res Sq. 2025 May 13:rs.3.rs-6363545. doi: 10.21203/rs.3.rs-6363545/v1.
Growing evidence indicates that transcriptional regulatory elements can exert both promoter and enhancer activity; however, the relationship and determinants of this dual functionality remain poorly understood. We developed a massively parallel dual reporter assay that enables simultaneous assessment of the intrinsic promoter and enhancer potential exerted by the same sequence. Parallel quantification for thousands of elements reveals that canonical human promoters and enhancers can act as both promoters and enhancers under the same contexts, and that promoter activity may be necessary but not sufficient for enhancer function. We find that regulatory potential is intrinsic to element sequences, irrespective of downstream features typically associated with distinct element classes. Perturbations to element transcription factor binding motifs lead to disruptions in both activities, implicating a shared syntax for the two regulatory functions. Combinations of elements with different minimal promoters reveal reciprocal activity modulation between associated elements and a strong positive correlation between promoter and enhancer functions imply a bidirectional feedback loop used to maintain environments of high transcriptional activity. Finally, our results indicate that the magnitude and balance between promoter and enhancer functions are shaped by both intrinsic sequence properties and contextual regulatory influences, suggesting a degree of plasticity in regulatory action. Our approach provides a new lens for understanding fundamental principles of regulatory element biology.
越来越多的证据表明，转录调控元件可同时发挥启动子和增强子的活性；然而，这种双重功能的关系和决定因素仍知之甚少。我们开发了一种大规模平行双报告基因检测方法，能够同时评估同一序列发挥的内在启动子和增强子潜能。对数千个元件进行平行定量分析发现，典型的人类启动子和增强子在相同背景下可同时作为启动子和增强子发挥作用，且启动子活性对于增强子功能可能是必要的，但并不充分。我们发现调控潜能是元件序列所固有的，与通常与不同元件类别相关的下游特征无关。对元件转录因子结合基序的扰动会导致两种活性都受到破坏，这意味着两种调控功能存在共同的语法规则。具有不同最小启动子的元件组合揭示了相关元件之间的相互活性调节，并且启动子和增强子功能之间的强正相关意味着存在一个双向反馈环，用于维持高转录活性环境。最后，我们的结果表明，启动子和增强子功能之间的强度和平衡受到内在序列特性和背景调控影响的共同塑造，这表明调控作用具有一定程度的可塑性。我们的方法为理解调控元件生物学的基本原理提供了一个新的视角。
