Systematic comparison of estimates of transcription factor activity by ATAC-seq and multiplexed reporter assays

Transcription factors (TFs) are central to gene regulation and play critical roles in development, cellular homeostasis and disease. The ability to accurately measure TF activity is essential to understanding how TFs respond to signals and regulate target genes. In one commonly used approach, activities of TFs are computationally inferred from genome-wide chromatin accessibility data (ATAC-seq). However, it has remained unclear how well these inferences reflect actual regulatory activity of TFs. An alternative approach employs a collection of synthetic reporters that are designed to each probe the regulatory activity of a single TF; when used in a multiplexed assay, the activity of many TFs can be probed in parallel In this study, we systematically compared TF activities as inferred by ATAC-seq with those measured by multiplexed reporters across diverse perturbations known to alter specific TFs. We observed considerable overlap between the two methods, but also notable discrepancies. Our findings suggest that reporter assays and chromatin-based inference capture distinct aspects of TF function: reporter assays are more sensitive to signal-responsive TFs, while ATAC-seq better detects chromatin-modifying TFs.

转录因子 (Transcription factors, TFs) 是基因调控的核心，在个体发育、细胞稳态维持以及疾病发生过程中发挥关键作用。精准测定转录因子的活性，对于解析其如何响应上游信号并调控靶基因至关重要。目前常用的一种方法是基于全基因组染色质开放程度数据（ATAC-seq），通过计算手段推断转录因子的活性。然而，此类推断结果能否准确反映转录因子的真实调控活性，迄今仍未明确。另一种方法则采用一系列合成报告元件，每个元件均设计用于特异性探测单个转录因子的调控活性；当采用多重检测实验时，可同时并行测定多种转录因子的活性。本研究系统对比了通过ATAC-seq推断得到的转录因子活性，与通过多重报告元件测得的转录因子活性，对比场景涵盖多种已知会改变特定转录因子活性的扰动条件。研究结果显示，两种方法的检测结果存在显著重叠，但也存在明显差异。本研究结果表明，报告元件检测与基于染色质的推断方法分别捕捉到了转录因子功能的不同维度：报告元件检测对信号响应型转录因子更为敏感，而ATAC-seq则更擅长检测染色质修饰型转录因子。