The E7 open reading frame of human papillomavirus type 16 (HPV-16) encodes a protein that can immortalize primary rat cells, cooperate with the ras oncoprotein to transform low passage rat cells and transform established rodent cells to anchorage independence. The immortalizing and cooperation functions have been investigated using a series of point mutations that introduce single amino acid changes into the E7 protein in two distinct regions. Certain mutations altering amino acids conserved between the E7 protein of genital HPV types, the adenovirus E1 a protein and simian virus 40 large T antigen abolished the ability of the E7 protein to immortalize or cooperate with ras in a focus forming assay. Mutations in a consensus sequence for a casein kinase II recognition site, which is also shared by E1a and large T, reduced immortalizing activity, but did not affect the ability to cooperate with ras. Single mutations disrupting cysteine motifs, which form putative zinc-binding sites in the second region, reduced the activity of the E7 protein, whereas double mutants, in which neither of the cysteine motifs remained intact, showed no or very low activity. The activity of the mutants in immortalization and cooperation assays was essentially the same as their transforming activities in NIH 3T3 cells. This indicates that these three functions of E7 map to overlapping domains which cannot be separated by these mutations in the region of E1a/large T homology or the cysteine motifs.
ChestersP. M., MccanceD. J.1989; Human papillomavirus types 6 and 16 in cooperation with Ha-rastransform secondary rat embryo fibroblasts. Journal of General Virology 70:353–365
ColeS. T., DanosO.1987; Nucleotide sequence and comparative analysis of the human papillomavirus type 18 genome: phylogeny of papillomaviruses and repeated structure of the E6 and E7 gene products. Journal of Molecular Biology 193:599–608
CrookT., StoreyA., AlmondN., OsbornK., CrawfordL.1988; Human papillomavirus type 16 cooperates with activated ras and fos oncogenes in the hormone-dependent transformation of primary mouse cells. Proceedings of the National Academy of Sciences, U.S.A 85:8820–8824
De CaprioJ. A., LudlowJ. W., FiggeJ., ShewJ.-Y., HuangC.-M., LeeW.-H., MasilioE., PauchaE., LivingstonD. M.1988; SV40 large tumor antigen forms a specific complex with the product of the retinoblastoma susceptibility gene. Cell 54:275–283
DürstM., GissmannL., IkenbergH., Zur HausenH.1983; A papillomavirus DNA from a cervical carcinoma and its prevalence in cancer biopsy samples from different geographic regions. Proceedings of the National Academy of Sciences, U.S.A 80:3812–3815
DysonN., HowleyP. M., MungerK., HarlowE.1989; The human papillomavirus-16 E7 oncoprotein is able to bind to the retinoblastoma gene product. Science 243:934–937
GissmannL., BoshartM., DurstM., IkenbergH., WagnerD.1984; Presence of human papillomavirus (HPV) DNA in genital tumours. Journal of Investigative Dermatology 83:265–288
JatP. S., SharpP. A.1989; Cell lines established by a temperature sensitive SV40 large T antigen are growth restricted at the nonpermissive temperature. Molecular and Cellular Biology 9:1672–1681
KandaT., WatanabeS., YoshiikeK.1988; Immortalisation of primary rat cells by human papillomavirus type 16 subgenomic DNA fragments controlled by the SV40 promoter. Virology 165:321–325
LandH., ParadaL. F., WeinbergR.1983; Tumorigenic conversion of primary embryo fibroblasts requires at least two cooperating oncogenes. Nature; London: 304596–602
MoranE., ZerlerB., HarrisonT. M., MathewsM. B.1986; Identification of separate domains in the adenovirus E1A gene for immortalisation activity and the activation of virus early genes. Molecular and Cellular Biology 6:3470–3480
PhelpsW. C., YeeC. L., MungerK., HowleyP. M.1988; The human papillomavirus type 16 E7 gene encodes transactivation and transformation functions similar to those of adenovirus E1a. Cell 53:539–547
SmithD. H., ZiffE. B.1988; The amino-terminal region of the adenovirus serotype 5 Elaprotein performs two separate functions when expressed in primary rat kidney cells. Molecular and Cellular Biology 8:3882–3890
SmotkinD., WettsteinF. O.1987; The major human papillomavirus protein in cervical cancers is a cytoplasmic phosphoprotein. Journal of Virology 61:1686–1689
StoreyA., PimD., MurrayA., OsbornK., BanksL., CrawfordL.1988; Comparison of the in vitro transforming activities of human papillomavirus types. EMBO Journal 1:1815–1820
SugdenB., MarshK., YatesJ.1985; A vector that replicates as a plasmid and can be efficiently selected in B-lymphoblasts transformed by Epstein-Barr virus. Molecular and Cellular Biology 5:410–413
VousdenK. H., DonigerJ., DipaoloJ. A., LowyD. R.1988; The E7 open reading frame of human papillomavirus type 16 encodes a transforming gene. Oncogene Research 3:167–175
WhyteP., RuleyH. E., HarlowE.1988; Two regions of the adenovirus early region 1A protein are required for transformation. Journal of Virology 62:257–265