title: Biologic activity of human papillomavirus type 16 E6/E7 cDNA clones isolated from SiHa cervical carcinoma cell line.
abstract: Three species of E6/E7 cDNAs of human papillomavirus type 16 (HPV16) for the full-length E6/E7 and spliced E6*I/E7 and E6*II/E7 mRNAs were synthesized by reverse transcriptase-(RT-)PCR from RNA of the cervical carcinoma cell line SiHa. Two cDNA mutants carrying point mutations in either a splice donor site or acceptor site within the E6 open reading frame were also constructed. These HPV16 E6/E7 cDNAs were cloned under the SV40 enhancer/promoter and the MMTV LTR to examine the activities of ras-collaborative transformation and induction of cellular DNA synthesis, both of which depend on the E7 gene product. The E6*II/E7 cDNA and two mutated cDNAs deficient in the spliced mRNA transcription showed lower levels of both activities than the full-length E6/E7 and the E6*I/E7 cDNA. The rat cell lines carrying each of the E6/E7 cDNAs contained the E6/E7 mRNA species expected. A small amount of E6*I/E7-sized mRNA was transcribed from a splice-donor site mutant of the E6/E7 cDNA, which turned out to be a transcript derived from a cryptic splice donor site six bases upstream from the conventional site. Among NIH3T3 cells carrying one of the above-mentioned E6/E7 cDNAs, the cells expressing E6*I/E7 mRNA [cells carrying cF(wt) and c*I] produced an amount of E7 protein comparable with those carrying the E7 or E6E7 region. These results suggest that the E6*I/E7 is the mRNA that is important for the efficient expression of E7 product from the HPV16 E6/E7 region.
mesh_terms: D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002472:Cell Transformation, Viral; D003001:Cloning, Molecular; D004247:DNA / Q000096:biosynthesis; D004279:DNA, Viral; D005260:Female; D015967:Gene Expression Regulation, Viral; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000502:physiology*; D027383:Papillomaviridae / Q000235:genetics / Q000502:physiology*; D050725:Papillomavirus E7 Proteins; D012326:RNA Splicing / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D012367:RNA, Viral / Q000032:analysis / Q000235:genetics; D051381:Rats; D012097:Repressor Proteins; D014407:Tumor Cells, Cultured; D002583:Uterine Cervical Neoplasms / Q000821:virology*
chemical_list: D004279:DNA, Viral; C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D012333:RNA, Messenger; D012367:RNA, Viral; D012097:Repressor Proteins; C059731:oncogene protein E7, Human papillomavirus type 16; D004247:DNA