Trauernicht Max, Filipovska Teodora, Rastogi Chaitanya, van Steensel Bas
Oncode Institute, Division of Gene regulation and Division of Molecular Genetics, Netherlands Cancer Institute, 1066 CX Amsterdam, the Netherlands.
Department of Biological Sciences, Columbia University, New York, NY, USA.
bioRxiv. 2024 Jul 26:2024.07.26.605239. doi: 10.1101/2024.07.26.605239.
In any given cell type, dozens of transcription factors (TFs) act in concert to control the activity of the genome by binding to specific DNA sequences in regulatory elements. Despite their considerable importance in determining cell identity and their pivotal role in numerous disorders, we currently lack simple tools to directly measure the activity of many TFs in parallel. Massively parallel reporter assays (MPRAs) allow the detection of TF activities in a multiplexed fashion; however, we lack basic understanding to rationally design sensitive reporters for many TFs. Here, we use an MPRA to systematically optimize transcriptional reporters for 86 TFs and evaluate the specificity of all reporters across a wide array of TF perturbation conditions. We thus identified critical TF reporter design features and obtained highly sensitive and specific reporters for 60 TFs, many of which outperform available reporters. The resulting collection of "prime" TF reporters can be used to uncover TF regulatory networks and to illuminate signaling pathways.
在任何给定的细胞类型中，数十种转录因子（TFs）协同作用，通过与调控元件中的特定DNA序列结合来控制基因组的活性。尽管它们在决定细胞身份方面具有相当重要的意义，并且在众多疾病中起着关键作用，但目前我们缺乏能够直接并行测量多种TF活性的简单工具。大规模平行报告基因检测（MPRAs）允许以多重方式检测TF活性；然而，对于许多TF，我们缺乏合理设计敏感报告基因的基本认识。在这里，我们使用MPRA系统地优化了86种TF的转录报告基因，并在广泛的TF干扰条件下评估了所有报告基因的特异性。因此，我们确定了关键的TF报告基因设计特征，并获得了60种TF的高灵敏度和特异性报告基因，其中许多报告基因的性能优于现有报告基因。由此产生的“优质”TF报告基因集合可用于揭示TF调控网络并阐明信号通路。
