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. 1988 Mar;7(3):825–833. doi: 10.1002/j.1460-2075.1988.tb02881.x

Analysis of HPV-1 E4 gene expression using epitope-defined antibodies.

J Doorbar 1, H S Evans 1, I Coneron 1, L V Crawford 1, P H Gallimore 1
PMCID: PMC454398  PMID: 2456213

Abstract

Six monoclonal antibodies (mAbs) have been raised against the E4 proteins of HPV-1. Five of these were found to recognize denaturation-resistant epitopes as determined by Western blotting--and their binding sites were identified by determining their reactivity against a panel of bacterial E4--beta-galactosidase fusion proteins which contained progressive deletions at the C-terminal end of the E4 region. The five mAbs were found to bind to four distinct sites. By using these epitope-defined mAbs, along with anti-peptide antibodies raised against putative N- and C-terminal E4 sequences, we have determined the relationships between the eight distinct polypeptides (mol. wt 10/11 kd, 16/17 kd, 21/23 kd and 32/34 kd) previously shown to be expressed from the E4 gene of HPV-1 in productively infected papillomas. The 17 kd E4 polypeptide appears to be the product of a spliced mRNA encoding five amino acids from open reading frame (ORF) E1 joined onto 120 from the E4 ORF. The 16 kd and 10/11 kd proteins, which may be derived from this, lack sequences (approximately 15 and 70 amino acids respectively) encoded by the 5' end of the E4 gene. The 32/34 kd proteins were detected by all antibodies which reacted with the 16/17 kd polypeptides, suggesting that they represent dimers of the latter species. The 21/23 kd polypeptides, however, do not appear to be simple dimers of the 10/11 kd protein as previously predicted, and reacted with antibodies whose epitopes mapped in the N-terminal half of the E4 protein.(ABSTRACT TRUNCATED AT 250 WORDS)

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Androphy E. J., Schiller J. T., Lowy D. R. Identification of the protein encoded by the E6 transforming gene of bovine papillomavirus. Science. 1985 Oct 25;230(4724):442–445. doi: 10.1126/science.2996134. [DOI] [PubMed] [Google Scholar]
  2. Baker C. C., Howley P. M. Differential promoter utilization by the bovine papillomavirus in transformed cells and productively infected wart tissues. EMBO J. 1987 Apr;6(4):1027–1035. doi: 10.1002/j.1460-2075.1987.tb04855.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berzofsky J. A., Hicks G., Fedorko J., Minna J. Properties of monoclonal antibodies specific for determinants of a protein antigen, myoglobin. J Biol Chem. 1980 Dec 10;255(23):11188–11191. [PubMed] [Google Scholar]
  4. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chow L. T., Nasseri M., Wolinsky S. M., Broker T. R. Human papillomavirus types 6 and 11 mRNAs from genital condylomata acuminata. J Virol. 1987 Aug;61(8):2581–2588. doi: 10.1128/jvi.61.8.2581-2588.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chow L. T., Reilly S. S., Broker T. R., Taichman L. B. Identification and mapping of human papillomavirus type 1 RNA transcripts recovered from plantar warts and infected epithelial cell cultures. J Virol. 1987 Jun;61(6):1913–1918. doi: 10.1128/jvi.61.6.1913-1918.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cole S. T., Streeck R. E. Genome organization and nucleotide sequence of human papillomavirus type 33, which is associated with cervical cancer. J Virol. 1986 Jun;58(3):991–995. doi: 10.1128/jvi.58.3.991-995.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Crawford L. V., O'Farrell P. Z. Effect of alkylation on the physical properties of simian virus 40 T-antigen species. J Virol. 1979 Feb;29(2):587–596. doi: 10.1128/jvi.29.2.587-596.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Danos O., Katinka M., Yaniv M. Human papillomavirus 1a complete DNA sequence: a novel type of genome organization among papovaviridae. EMBO J. 1982;1(2):231–236. doi: 10.1002/j.1460-2075.1982.tb01152.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dartmann K., Schwarz E., Gissmann L., zur Hausen H. The nucleotide sequence and genome organization of human papilloma virus type 11. Virology. 1986 May;151(1):124–130. doi: 10.1016/0042-6822(86)90110-8. [DOI] [PubMed] [Google Scholar]
  11. DiMaio D., Guralski D., Schiller J. T. Translation of open reading frame E5 of bovine papillomavirus is required for its transforming activity. Proc Natl Acad Sci U S A. 1986 Mar;83(6):1797–1801. doi: 10.1073/pnas.83.6.1797. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Doorbar J., Campbell D., Grand R. J., Gallimore P. H. Identification of the human papilloma virus-1a E4 gene products. EMBO J. 1986 Feb;5(2):355–362. doi: 10.1002/j.1460-2075.1986.tb04219.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Doorbar J., Gallimore P. H. Identification of proteins encoded by the L1 and L2 open reading frames of human papillomavirus 1a. J Virol. 1987 Sep;61(9):2793–2799. doi: 10.1128/jvi.61.9.2793-2799.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fuchs P. G., Iftner T., Weninger J., Pfister H. Epidermodysplasia verruciformis-associated human papillomavirus 8: genomic sequence and comparative analysis. J Virol. 1986 May;58(2):626–634. doi: 10.1128/jvi.58.2.626-634.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Giri I., Danos O., Yaniv M. Genomic structure of the cottontail rabbit (Shope) papillomavirus. Proc Natl Acad Sci U S A. 1985 Mar;82(6):1580–1584. doi: 10.1073/pnas.82.6.1580. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Guiso N., Dreyfus M., Siffert O., Danchin A., Spyridakis A., Gargouri A., Claisse M., Slonimski P. P. Antibodies against synthetic oligopeptides allow identification of the mRNA-maturase encoded by the second intron of the yeast cob-box gene. EMBO J. 1984 Aug;3(8):1769–1772. doi: 10.1002/j.1460-2075.1984.tb02044.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  18. Hopp T. P., Woods K. R. Prediction of protein antigenic determinants from amino acid sequences. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3824–3828. doi: 10.1073/pnas.78.6.3824. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Komly C. A., Breitburd F., Croissant O., Streeck R. E. The L2 open reading frame of human papillomavirus type 1a encodes a minor structural protein carrying type-specific antigens. J Virol. 1986 Nov;60(2):813–816. doi: 10.1128/jvi.60.2.813-816.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lowy D. R., Dvoretzky I., Shober R., Law M. F., Engel L., Howley P. M. In vitro tumorigenic transformation by a defined sub-genomic fragment of bovine papilloma virus DNA. Nature. 1980 Sep 4;287(5777):72–74. doi: 10.1038/287072a0. [DOI] [PubMed] [Google Scholar]
  21. Lusky M., Botchan M. R. Genetic analysis of bovine papillomavirus type 1 trans-acting replication factors. J Virol. 1985 Mar;53(3):955–965. doi: 10.1128/jvi.53.3.955-965.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Messing J., Crea R., Seeburg P. H. A system for shotgun DNA sequencing. Nucleic Acids Res. 1981 Jan 24;9(2):309–321. doi: 10.1093/nar/9.2.309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mount S. M. A catalogue of splice junction sequences. Nucleic Acids Res. 1982 Jan 22;10(2):459–472. doi: 10.1093/nar/10.2.459. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Nasseri M., Wettstein F. O. Differences exist between viral transcripts in cottontail rabbit papillomavirus-induced benign and malignant tumors as well as non-virus-producing and virus-producing tumors. J Virol. 1984 Sep;51(3):706–712. doi: 10.1128/jvi.51.3.706-712.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Neary K., Horwitz B. H., DiMaio D. Mutational analysis of open reading frame E4 of bovine papillomavirus type 1. J Virol. 1987 Apr;61(4):1248–1252. doi: 10.1128/jvi.61.4.1248-1252.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Pfister H. Biology and biochemistry of papillomaviruses. Rev Physiol Biochem Pharmacol. 1984;99:111–181. doi: 10.1007/BFb0027716. [DOI] [PubMed] [Google Scholar]
  27. Pfister H., Krubke J., Dietrich W., Iftner T., Fuchs P. G. Classification of the papillomaviruses--mapping the genome. Ciba Found Symp. 1986;120:3–22. doi: 10.1002/9780470513309.ch2. [DOI] [PubMed] [Google Scholar]
  28. Phelps W. C., Leary S. L., Faras A. J. Shope papillomavirus transcription in benign and malignant rabbit tumors. Virology. 1985 Oct 15;146(1):120–129. doi: 10.1016/0042-6822(85)90058-3. [DOI] [PubMed] [Google Scholar]
  29. Raibaud O., Mock M., Schwartz M. A technique for integrating any DNA fragment into the chromosome of Escherichia coli. Gene. 1984 Jul-Aug;29(1-2):231–241. doi: 10.1016/0378-1119(84)90183-5. [DOI] [PubMed] [Google Scholar]
  30. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  31. Robson B., Fishleigh R. V., Morrison C. A. Prediction of HIV vaccine. 1987 Jan 29-Feb 4Nature. 325(6103):395–395. doi: 10.1038/325395a0. [DOI] [PubMed] [Google Scholar]
  32. Rothnagel J. A., Rogers G. E. Transglutaminase-mediated cross-linking in mammalian epidermis. Mol Cell Biochem. 1984;58(1-2):113–119. doi: 10.1007/BF00240610. [DOI] [PubMed] [Google Scholar]
  33. Rowe M., Evans H. S., Young L. S., Hennessy K., Kieff E., Rickinson A. B. Monoclonal antibodies to the latent membrane protein of Epstein-Barr virus reveal heterogeneity of the protein and inducible expression in virus-transformed cells. J Gen Virol. 1987 Jun;68(Pt 6):1575–1586. doi: 10.1099/0022-1317-68-6-1575. [DOI] [PubMed] [Google Scholar]
  34. Rowe M., Hildreth J. E., Rickinson A. B., Epstein M. A. Monoclonal antibodies to Epstein-Barr virus-induced, transformation-associated cell surface antigens: binding patterns and effect upon virus-specific T-cell cytotoxicity. Int J Cancer. 1982 Apr 15;29(4):373–381. doi: 10.1002/ijc.2910290403. [DOI] [PubMed] [Google Scholar]
  35. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Schwarz E., Dürst M., Demankowski C., Lattermann O., Zech R., Wolfsperger E., Suhai S., zur Hausen H. DNA sequence and genome organization of genital human papillomavirus type 6b. EMBO J. 1983;2(12):2341–2348. doi: 10.1002/j.1460-2075.1983.tb01744.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Seedorf K., Krämmer G., Dürst M., Suhai S., Röwekamp W. G. Human papillomavirus type 16 DNA sequence. Virology. 1985 Aug;145(1):181–185. doi: 10.1016/0042-6822(85)90214-4. [DOI] [PubMed] [Google Scholar]
  38. Smith-Gill S. J., Wilson A. C., Potter M., Prager E. M., Feldmann R. J., Mainhart C. R. Mapping the antigenic epitope for a monoclonal antibody against lysozyme. J Immunol. 1982 Jan;128(1):314–322. [PubMed] [Google Scholar]
  39. Stanley K. K., Luzio J. P. Construction of a new family of high efficiency bacterial expression vectors: identification of cDNA clones coding for human liver proteins. EMBO J. 1984 Jun;3(6):1429–1434. doi: 10.1002/j.1460-2075.1984.tb01988.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Sun T. T., Tseng S. C., Huang A. J., Cooper D., Schermer A., Lynch M. H., Weiss R., Eichner R. Monoclonal antibody studies of mammalian epithelial keratins: a review. Ann N Y Acad Sci. 1985;455:307–329. doi: 10.1111/j.1749-6632.1985.tb50419.x. [DOI] [PubMed] [Google Scholar]
  41. Tamura T., Bauer H., Birr C., Pipkorn R. Antibodies against synthetic peptides as a tool for functional analysis of the transforming protein pp60src. Cell. 1983 Sep;34(2):587–596. doi: 10.1016/0092-8674(83)90391-4. [DOI] [PubMed] [Google Scholar]
  42. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Warren G. Protein transport. Signals and salvage sequences. Nature. 1987 May 7;327(6117):17–18. doi: 10.1038/327017a0. [DOI] [PubMed] [Google Scholar]
  44. Watt F. M., Green H. Involucrin synthesis is correlated with cell size in human epidermal cultures. J Cell Biol. 1981 Sep;90(3):738–742. doi: 10.1083/jcb.90.3.738. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Winter G., Fields S. Cloning of influenza cDNA ino M13: the sequence of the RNA segment encoding the A/PR/8/34 matrix protein. Nucleic Acids Res. 1980 May 10;8(9):1965–1974. doi: 10.1093/nar/8.9.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Yoshitake S., Yamada Y., Ishikawa E., Masseyeff R. Conjugation of glucose oxidase from Aspergillus niger and rabbit antibodies using N-hydroxysuccinimide ester of N-(4-carboxycyclohexylmethyl)-maleimide. Eur J Biochem. 1979 Nov;101(2):395–399. doi: 10.1111/j.1432-1033.1979.tb19731.x. [DOI] [PubMed] [Google Scholar]
  47. von Heijne G. Signal sequences. The limits of variation. J Mol Biol. 1985 Jul 5;184(1):99–105. doi: 10.1016/0022-2836(85)90046-4. [DOI] [PubMed] [Google Scholar]

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