Mechanisms of transcription factor binding and cofactor activity in chromatin

Organisms rely on the establishment of specific gene expression patterns to ensure proper development and response to external cues. Transcription factors (TFs) and cofactors are key players in transcriptional regulation and they exert their function in the context of chromatin.
Here, DNA is wrapped around histone octamers forming structures called nucleosomes, which drive its compaction and are thought to regulate access of TFs to DNA. Histones can be post-translationally modified adding yet another layer of regulation. In the context of this thesis, I have studied two aspects of this process.
First, I tested the model that nucleosome position influences TF binding and separates TFs into different classes. Towards this goal, we developed a novel approach to systematically measure in the cellular context the ability of individual motifs to recruit TFs and to evaluate how motif position along nucleosomal DNA affects TF binding and nucleosome remodeling. This revealed that only few motifs allow stable recruitment of their cognate TF and that nucleosome phasing over the motif affects binding to different degrees in a highly TF-specific fashion. In contrast, all tested TFs, including the canonical pioneer factors OCT4-SOX2, displayed strong preference for motifs residing in linker regions and at the entry/exit sites of the nucleosome. This suggests that nucleosomes are negative regulator of TF binding in vivo and that chromatin sensitivity is a continuous- rather than binary feature.
Secondly, I have investigated how a cofactor that leads to modification of histones is recruited to chromatin and how it functions. More specifically, I focused on the essential corepressor SIN3A and dissected its activity by exploiting an inducible rapid degradation system. We generated a high-quality genome-wide binding profile of SIN3A and followed the effects of its rapid depletion over time. This enabled us to define its primary targets and revealed that SIN3A, despite being a corepressor, is present at almost all active promoters and only at a small fraction of silent genes. We found that SIN3A can be recruited through two independent mechanisms and its binding is required for constant deacetylation and transcriptional repression. We suggest that SIN3A buffers transcription and acetylation at active genes and is selectively recruited at silent promoters to maintain repression.