Context dependent activity of p63-bound gene regulatory elements

Assessing the contribution of the p63 binding motif on p63-dependent enhancer activity in 5 different cellular contexts. Implemented STARR-seq MPRA method (Arnold et al., 2013) to assay 17,310 unique enhancers. The library pool included 5 variants of each enhancer: wild-type, p63RE motif mutant, p63RE shuffle, p63RE flanking region shuffle, and a fully shuffled enhancer sequences. Each fragment tested included 119-120 nt of the enhancer centered around p63RE. All enhancers were assayed in several epithelial cell lines or their derivatives: MCF10A, MCF10A p53KO, HaCaT, SCC25, MCF10A GUS overexpression, MCF10A TAp63β overexpression. Plasmid pool 1 was transfected into all cell types except for HaCaT replicates 1 and 2 where plasmid pool 2 was transfected.