Essential genes are pioneered and activated by master transcription factors
Cell viability depends on the expression of common essential genes (CEGs) that encode for fundamental cellular processes such as metabolism. The level of CEG expression is tightly controlled across cell types and developmental stages, with dysregulation linked to disease, yet, the mechanisms governing CEG regulation remain unclear. CpG island (CGI) promoters, the most prevalent cis-regulatory element in the mammalian genome, drive CEG expression. They contain assemblies of transcription factor (TF) binding sites and the highest TF binding density among promoters, leading to a model where CGIs activity is regulated by the combinatorial action of multiple TFs. This makes CGIs an intriguing model to study how TFs cooperate to open chromatin and activate genes, including CEGs.
To address this, we investigated five essential TFs that co-bind extensively at CEG promoters in murine stem cells. To dissect their regulatory roles, we used individual and combinatorial inducible degron systems, enabling to map the primary effects on gene expression and chromatin accessibility following rapid TF depletion and recovery. This identified TFs that can drive chromatin opening and transcriptional activation, while others regulate gene expression without altering chromatin state. The ability to open chromatin was promoter-specific and could not be compensated for by additional co-bound TFs. Furthermore, this pioneer activity facilitated the recruitment of co-bound TFs, allowing us to reconstruct the hierarchical binding events necessary for CGI promoter function. We found some genes where TF cooperativity tuned gene expression, both enhancing and repressing activity, however, the majority of genes were predominantly controlled by individual TFs. Our study provides new insights into CEG regulation, revealing that CGI promoters are continually pioneered by master TFs that organize chromatin, define binding hierarchies, and drive gene activation.