Profiling transcription factor binding affinities across the genome using BANC-seq
Transcription factor (TF) binding across the genome regulates gene expression and cell fate. Furthermore, TF-mediated gene expression is frequently deregulated in diseases such as cancer. In recent years, various sequencing-based methods have been developed to profile the epigenetic landscape and transcription factor binding specificity across the genome. However, to biochemically understand transcription factor binding, the binding affinity – i.e. the TF concentration required for specific binding – of a transcription factor for DNA sequences in the genome must also be considered. However, no existing technology is capable of quantifying absolute transcription factor binding affinities to native, chromatinized DNA.
Hannah Neikes, Rik Lindeboom, Michiel Vermeulen and their colleagues addressed this major shortcoming in the field by developing a method to determine Binding Affinities to Native Chromatin by sequencing or BANC-seq. In BANC-seq, a concentration range of a tagged transcription factor is added to isolated nucleic from mammalian cells. Concentration dependent binding for each titration point is then measured to quantify binding affinities across the genome. BANC-seq revealed that, by and large, accessible chromatin is a pre-requisite for high and low affinity transcription factor binding to occur in a specific epigenetic state. Furthermore, changes in chromatin context during cellular differentiation result in cell type-specific transcription factor binding affinities. Interestingly however, chromatin context is interpreted differently by the pioneering transcription factor FOXA1 compared to YY1, SP1 or MYC. Notably, whereas consensus DNA binding motifs for TFs are important to establish high-affinity binding sites in the genome, these motifs are not always strictly required to generate nanomolar affinity interactions in the genome.
In summary, BANC-seq adds a much needed quantitative dimension to transcription factor biology and enables stratification of transcription factor binding sites in pathological conditions, such as over-expression of oncogenes in cancer. These breakthrough results were recently published in Nature Biotechnology.