IER5 is required for Notch-mediated induction of squamous cell differentiation [RNA-seq]

Conditional activation of Notch in squamous cells activates a context-specific gene expression program through lineage-specific regulatory elements. IER5 is a direct Notch-regulated gene and required for Notch-induced differentiation of squamous carcinoma cells. Overall design: To create a squamous cell carcinoma (SCC) model in which GSI (?-secretase inhibitor) washout activates NOTCH1, we engineered a cDNA encoding a mutated truncated form of NOTCH1, ?EGF-L1596H, that cannot respond to ligand and that has a point substitution in its negative regulatory region that produces ligand-independent, ?-secretase-dependent Notch activation. ?EGF-L1596H was transduced into human SCC cell line IC8 that have biallelic inactivating mutation in NOTCH1. SC2 cells were isolated as a single clone from IC8/?EGF-L1596H cells. GSI washout was performed for 4, 24 and 72 hours with SC2, I5 (SC2 cells with IER5 deleted) and I5/IER5 (I5 cells with IER5 addback) cells. Biological triplicates were analyzed for each of 12 conditions. Two runs for all 36 RNA-seq library samples