The nucleosome-binding affinities of p53, p63, and p73 are collectively determined by the composition, accessibility, and helical orientation of their binding sites

The p53 family of transcription factors plays key roles in driving development and combating cancer by regulating gene expression. p53, p63, and p73, which are the three members of the p53 family, regulate gene expression changes by binding to their DNA binding sites, many of which are positioned within nucleosomes. To comprehensively examine the nucleosome binding abilities of the p53 family, we developed Pioneer seq. Pioneer seq is a novel technique that enables the comprehensive assessment of the binding affinity of a transcription factor to its DNA binding sites at all 147 base pair positions of the nucleosome core particle. Using Pioneer seq, we analyzed the binding affinity of p53, p63, and p73 to 10 p53 family binding sites across the nucleosome core particle. The nucleosomal positioning of p53 family binding sites had the largest impact on p53 family binding affinity. p53 family members bound strongly near the more accessible edges of nucleosomes but weakly near the less accessible centers of nucleosomes. The nucleosome binding affinity of each p53 family member was also impacted by the composition of their binding sites. That is, each p53 family member was able to bind nucleosomal DNA more strongly in the presence of high affinity p53 family binding sites than in the presence of low affinity p53 family binding sites at the same nucleosomal position. We also found that the DNA helical orientation of p53 family binding sites within nucleosomal DNA impacted the nucleosome binding affinity of p53 family members. Taken together, our results help explain the rules underlying p53 family nucleosome binding and thus provide requisite insight into how we may better control the expression of genes involved in development and tumor suppression.