Association of the human papillomavirus type 11 E1()E4 protein with cornified cell envelopes derived from infected genital epithelium

Abstract

The cornified cell envelope (CCE) is an insoluble matrix of covalently linked proteins assembled in differentiating keratinocytes, providing a barrier against external insults. CCEs derived from HPV 11-infected tissue are fragile compared to those derived from healthy epithelium. To study a possible role for the E1()E4 protein, HPV 11-infected epithelium was examined for the distribution of this protein and three CCE proteins. CCEs were then purified from genital epithelium, fragmented, washed to remove nonassociated proteins, and analyzed for E1()E4 protein. In HPV 11-infected tissue, the E1()E4 protein was detected in the region of the CCE in differentiated keratinocytes. Loricrin and cytokeratin 10 (K10) were absent in E1()E4-positive cells, and E1()E4 protein was not detected in cells containing these proteins. E1()E4 protein was detected in immunoblots as a 10- to 11-kDa doublet in extracts of intact CCEs from infected tissue and in extracts of CCE fragments prepared without using reducing agents. Extraction with reducing agents eliminated E1()E4 detection, suggesting that disulfide bonding was involved in the association with CCE fragments. In addition, cyanogen bromide degradation experiments, immunofluorescence, and immunoelectron microscopy provided evidence that E1()E4 protein was associated with CCE fragments by covalent bonds other than disulfide bonds. We conclude that E1()E4 protein expression is associated with profound alterations in detection of loricrin and K10 in HPV 11-infected genital epithelium. The E1()E4 protein copurified with CCEs derived from infected epithelium and could be identified in CCE fragments, suggesting a possible role for E1()E4 in the development of CCE abnormalities.