Effect of IRF3 knockdown on gene expression in an HPV16-containing epithelium

The stromal compartment is critical for regulating the growth and differentiation of the epithelium. The life cycle of HPV is tightly linked to epithelial differentiation, and so we investigated the influence of the interferon-related transcription factor IRF3 (expressed in the stroma) on the gene expression in an HPV-containing epithelium. Organotypic cultures were created using shRNAs to knock down IRF3 in the stromal fibroblasts (or non-target controls). HPV16-containing keratinocytes were seeded on the surface and the epithelium was allowed to develop for 14 days. RNA-seq was performed on RNAs from both the epithelial and stromal compartments. We found that knockdown of IRF3 in the stroma did not affet the interferon system at all; rather, gene expression related to epithelial-mesenchymal transition was decreased, and epithelial differentiation markers were increased. These data suggest that IRF3 in stromal cells promotes EMT. Organotypic cultures were created from HPV16-containing keratinocytes, using human foreskin fibroblasts either containing IRF3 knockdown shRNAs or non-target controls. Total RNAs were harvested from both the keratinocytes and fibroblasts (stroma) 14 days following lifting to the air-liquid interface.