Detection by antibody probes of human papillomavirus type 6 E5 proteins in respiratory papillomata

Abstract

We have demonstrated the expression of proteins arising from the E5a and E5b open reading frames (ORFs) of human papillomavirus type 6c (HPV-6c) in respiratory tract papillomata. Recombinant plasmids were constructed to express the ORFs in the bacterial vectors pATH and pRIT2T. Fusion proteins were purified and injected into rabbits to produce polyclonal antibodies. Characterized antibodies generated against these fusion proteins were used in immunoperoxidase assays to identify the presence and distribution of HPV-6 E5 proteins in biopsy specimens of respiratory tract papillomata. The results showed that the E5a and E5b proteins were distributed throughout the thickness of the epithelium in the papillomata but not in the basal layer. The proteins were found in nuclei and in the cytoplasm of koilocytotic cells. Positive reactivity with a similar distribution in the epithelium and subcellular location was obtained in papillomata induced by other HPV-6 subtypes. This cross-reactivity was not unexpected, since nucleotide and amino acid (aa) sequence comparisons between HPV-6c and -6e demonstrated 79% sequence identity with 15 aa substitutions in the 91 aa of E5a. The E5b ORF of HPV-6c has the potential to encode a protein of 74 aa that differed at 28 positions compared with the 72 aa of HPV-6e.