Expression profiling by high throughput sequencing
Summary
Activation domains (ADs) within transcription factors (TFs) induce gene expression by recruiting coactivators to specific regulatory regions. Within the prevailing model, TF-coactivator recruitment is independent of DNA binding, which is consistent with direct AD-coactivator interactions seen outside cells. However, this independence was not yet tested within the genomic context. Here, we targeted two Med15-interacting ADs to hundreds of budding yeast promoters through fusions with multiple DNA binding domains (DBDs), gradually controlling their abundances using libraries of synthetic promoters. Genomic profiling revealed that AD identity influences DNA binding locations and that transcription induction and Med15 recruitment are restricted to a subset of DBD-bound promoters displaying flexible expression, multiple-TFs binding, and fuzzy nucleosome architecture. Further, when fused to a DBD, Med15 redirected binding towards promoters of fuzzy nucleosomes, overcoming DBD-based preferences. Our results demonstrate that ADs and their recruited coactivators posses an inherent preference for genomic localization and, therefore, define the subset of induced promoters.
Overall design
The experiments here are RNA-seq samples, collection of samples was performed at OD600 of 0.4, for controls we used the expression profiling of non-genetically modified yeast strain (BY4741).
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