Modulation of the epidermal growth factor receptor by the human papillomavirus type 16 E5 protein in raft cultures of human keratinocytes

It has been shown that the E5 protein of the human papillomavirus type 16 modulates epidermal growth factor receptor downregulation in monolayer cultures of human keratinocytes and mouse fibroblasts. We have now analysed the effect of this protein on the expression, the distribution and the activation of EGF receptors in raft cultures derived from an E5-transfected human keratinocyte cell line. The epithelia generated in these cultures were stratified and exhibited suprabasal expression of cytokeratins 1 and 10, which are known markers of early epidermal differentiation. In situ hybridization with an antisense riboprobe to the human papilloma virus type 16 E5 protein revealed a homogeneous gene expression within the entire epithelium of E5-transfected but not empty vector-transfected control cultures. Treatment of serum-starved rafts with EGF for 48 hours led to a strong decrease of suprabasal EGF receptors in control cultures, but not in rafts of E5-expressing cells. Under these conditions, no activated receptors were observed in control cultures, but activated receptors were still present in E5-raft cultures. Our results indicate that human papilloma virus type 16 E5-mediated modulation of EGF receptor expression occurs in a time- and structure-dependent manner in epithelial equivalents of human keratinocytes.