title: A point mutational analysis of human papillomavirus type 16 E7 protein.
abstract: The E7 open reading frame of human papillomavirus type 16 (HPV16) has been shown to be selectively retained in cervical tumors and to encode both transforming and trans-activating functions in murine cells, supporting the notion that expression of E7 contributes towards the progression of premalignant cervical lesions. A comparison among E7 sequences of different HPV types reveals some homology at the amino acid level. Of particular interest are two regions, one which contains significant homology to a region of adenovirus E1a and simian virus 40 large T (LT), and a second region which contains two conserved Cys-X-X-Cys motifs. To determine the importance of these domains to the function of the E7 protein, a series of mutants carrying substitutions at amino acids in the region of E1a-LT homology and at the Cys-X-X-Cys motifs were constructed. The mutated E7 sequences were placed under the control of a strong heterologous promoter (Moloney long terminal repeat), and the activity of the mutants was assayed in NIH 3T3 cells, a cell line in which both the transforming function and the trans-activating function of E7 could be determined. A single amino acid substitution analogous to a mutation in E1a which destroys the transforming ability of this protein abolished both transformation and trans-activation by E7. Mutations at the Cys-X-X-Cys motifs demonstrated that this region contributes to the transforming potential of E7, although proteins in which both motifs were interrupted retained a low level of transforming activity. Mutations in the region of E1a-LT homology which occur within a recognition sequence for casein kinase II did not markedly affect transforming activity of E7 but severely reduced trans-activating ability. This indicates that efficient trans-activation is not required for transformation by HPV16 E7 in these cells.
authors.affiliation: Ludwig Institute for Cancer Research, St. Mary's Hospital Medical School, London, England.
authors.forename: C
authors.initials: C
authors.lastname: Edmonds
authors.forename: K H
authors.initials: KH
authors.lastname: Vousden
mesh_terms: D000595:Amino Acid Sequence; D015153:Blotting, Western; D002472:Cell Transformation, Viral; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D027383:Papillomaviridae / Q000235:genetics*; D050725:Papillomavirus E7 Proteins; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic
chemical_list: D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; C059731:oncogene protein E7, Human papillomavirus type 16