Figure 1: HPV16 impairs IFN-γ and TNF-α-induced cytokine production and RelA K310 acetylation in KCs.

(a) RT–qPCR of CCL2, RANTES, IL-8 and CXCL9 expression by 24-h control, IFN-γ- and/or TNF-α-stimulated undifferentiated KCs or HPV16+KCs. Gene expression was normalized using GAPDH as the calibrator gene. Fold changes over control-stimulated undifferentiated KCs were calculated and depicted. (b) Enzyme-linked immunosorbent assay for CCL2, RANTES, IL-8 and CXCL9 in cleared supernatants of 24-h control, IFN-γ- and/or TNF-α-stimulated undifferentiated KCs or HPV16+KCs. (c) PBMCs migration towards cleared supernatants of 24-h control, IFN-γ- and/or TNF-α-stimulated KCs or HPV16+KCs. A representative example of three different donors is shown. (d) RelA phosphorylation, acetylation and total levels in KCs and HPV16+KCs stimulated with TNF-α for 0, 5, 15 and 30 min. (e) RelA acetylation and total levels at steady state in three human primary KC donor pools originating from human foreskin keratinocytes (HFKs), human vaginal keratinocytes (HVKs) or human cervical keratinocytes (HCKs) and two HPV16+genome-transfected primary KC pools of foreskin (HFK16) or vaginal (HVK16) origin. (f) RT–qPCR of CCL2, RANTES, IL-8 and CXCL9 in HPV16+KCs and KCs. Gene expression was normalized using GAPDH as the calibrator gene. Gene expression in HPV16+KCs was standardized over KCs. All data are representative for at least three independent experiments. Error bars indicate s.d. P values were determined using Welch-corrected unpaired t-tests. *P<0.05, **P<0.01 and ***P<0.001.