UNIFORMITY OF THE SPLICING PATTERN OF THE E6 E7 TRANSCRIPTS IN HUMAN PAPILLOMAVIRUS TYPE-16-TRANSFORMED HUMAN FIBROBLASTS, HUMAN CERVICAL PREMALIGNANT LESIONS AND CARCINOMAS

We utilized the RNA polymerase chain reaction (PCR) to analyse the transcripts of the E6/E7 open reading frames of human papillomavirus type 16 (HPV-16). Total RNA was isolated from 14 cervical squamous carcinomas, nine cervical intraepithelial neoplasias and from human fibroblasts transformed with different HPV-16 constructs. In all specimens two spliced transcripts were detected. Sequence analysis of the cloned PCR products showed that both transcripts were generated by splicing out an intron in E6, from nucleotides (nt) 226 to 409 in one transcript and from nt 226 to 526 in the other. The major transcript present in all RNA specimens had the smallest intron in E6. The RNA PCR described here is the method of choice for analysing splice and donor sites in tissue specimens where a limited amount of RNA is available. Results obtained with transformed cells revealed no difference in splicing whether HPV-16 was controlled by its homologous promoter or by a heterologous promoter, the Rous sarcoma virus long terminal repeat.