Skip to main content
Journal of Virology logoLink to Journal of Virology
. 1983 Nov;48(2):396–404. doi: 10.1128/jvi.48.2.396-404.1983

Insertion mutants of herpes simplex virus have a duplication of the glycoprotein D gene and express two different forms of glycoprotein D.

M G Gibson, P G Spear
PMCID: PMC255364  PMID: 6312097

Abstract

We produced insertion mutants of herpes simplex virus (HSV) that contain two functional copies of genes encoding different forms of glycoprotein D (gD). These viruses have the gene for HSV type 2 (HSV-2) gD at the normal locus and the gene for HSV-1 gD inserted into the thymidine kinase locus. Results of immunoprecipitation experiments done with monoclonal antibodies revealed that both gD genes were expressed by these viruses, regardless of orientation of the inserted HSV-1 gD gene, and that maximal synthesis of both glycoproteins depended on viral DNA replication. This apparently normal expression of the inserted HSV-1 gD gene was from a DNA fragment (SacI fragment, 0.906 to 0.924 map units) containing nucleotide sequences extending from approximately 400 base pairs upstream of the 5' end of the gD mRNA to about 200 base pairs upstream of the 3' end. The glycoproteins expressed from both genes were incorporated into the surfaces of infected cells. Electrophoretic analyses of purified virions and neutralization studies suggest that both glycoproteins were also incorporated into virions. This nonpreferential utilization of both gene products makes these viruses ideal strains for the generation and characterization of a variety of mutations.

Full text

PDF
396

Images in this article

Selected References

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

  1. Balachandran N., Harnish D., Rawls W. E., Bacchetti S. Glycoproteins of herpes simplex virus type 2 as defined by monoclonal antibodies. J Virol. 1982 Oct;44(1):344–355. doi: 10.1128/jvi.44.1.344-355.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Campione-Piccardo J., Rawls W. E., Bacchetti S. Selective assay for herpes simplex viruses expressing thymidine kinase. J Virol. 1979 Aug;31(2):281–287. doi: 10.1128/jvi.31.2.281-287.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cassai E. N., Sarmiento M., Spear P. G. Comparison of the virion proteins specified by herpes simplex virus types 1 and 2. J Virol. 1975 Nov;16(5):1327–1331. doi: 10.1128/jvi.16.5.1327-1331.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cohen G. H., Katze M., Hydrean-Stern C., Eisenberg R. J. Type-common CP-1 antigen of herpes simplex virus is associated with a 59,000-molecular-weight envelope glycoprotein. J Virol. 1978 Jul;27(1):172–181. doi: 10.1128/jvi.27.1.172-181.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cohen G. H., Long D., Eisenberg R. J. Synthesis and processing of glycoproteins gD and gC of herpes simplex virus type 1. J Virol. 1980 Nov;36(2):429–439. doi: 10.1128/jvi.36.2.429-439.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Eisenberg R. J., Ponce de Leon M., Cohen G. H. Comparative structural analysis of glycoprotein gD of herpes simplex virus types 1 and 2. J Virol. 1980 Aug;35(2):428–435. doi: 10.1128/jvi.35.2.428-435.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Eisenberg R. J., Ponce de Leon M., Pereira L., Long D., Cohen G. H. Purification of glycoprotein gD of herpes simplex virus types 1 and 2 by use of monoclonal antibody. J Virol. 1982 Mar;41(3):1099–1104. doi: 10.1128/jvi.41.3.1099-1104.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ejercito P. M., Kieff E. D., Roizman B. Characterization of herpes simplex virus strains differing in their effects on social behaviour of infected cells. J Gen Virol. 1968 May;2(3):357–364. doi: 10.1099/0022-1317-2-3-357. [DOI] [PubMed] [Google Scholar]
  9. Garfinkle B., McAuslan B. R. Regulation of herpes simplex virus-induced thymidine kinase. Biochem Biophys Res Commun. 1974 Jun 4;58(3):822–829. doi: 10.1016/s0006-291x(74)80491-2. [DOI] [PubMed] [Google Scholar]
  10. Heine J. W., Honess R. W., Cassai E., Roizman B. Proteins specified by herpes simplex virus. XII. The virion polypeptides of type 1 strains. J Virol. 1974 Sep;14(3):640–651. doi: 10.1128/jvi.14.3.640-651.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Holland T. C., Sandri-Goldin R. M., Holland L. E., Marlin S. D., Levine M., Glorioso J. C. Physical mapping of the mutation in an antigenic variant of herpes simplex virus type 1 by use of an immunoreactive plaque assay. J Virol. 1983 May;46(2):649–652. doi: 10.1128/jvi.46.2.649-652.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Honess R. W., Roizman B. Regulation of herpesvirus macromolecular synthesis. I. Cascade regulation of the synthesis of three groups of viral proteins. J Virol. 1974 Jul;14(1):8–19. doi: 10.1128/jvi.14.1.8-19.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Honess R. W., Watson D. H. Herpes simplex virus-specific polypeptides studied by polyacrylamide gel electrophoresis of immune precipitates. J Gen Virol. 1974 Feb;22(2):171–185. doi: 10.1099/0022-1317-22-2-171. [DOI] [PubMed] [Google Scholar]
  14. Kessler S. W. Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: parameters of the interaction of antibody-antigen complexes with protein A. J Immunol. 1975 Dec;115(6):1617–1624. [PubMed] [Google Scholar]
  15. Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
  16. Lee G. T., Para M. F., Spear P. G. Location of the structural genes for glycoproteins gD and gE and for other polypeptides in the S component of herpes simplex virus type 1 DNA. J Virol. 1982 Jul;43(1):41–49. doi: 10.1128/jvi.43.1.41-49.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Lee G. T., Pogue-Geile K. L., Pereira L., Spear P. G. Expression of herpes simplex virus glycoprotein C from a DNA fragment inserted into the thymidine kinase gene of this virus. Proc Natl Acad Sci U S A. 1982 Nov;79(21):6612–6616. doi: 10.1073/pnas.79.21.6612. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Leiden J. M., Buttyan R., Spear P. G. Herpes simplex virus gene expression in transformed cells. I. Regulation of the viral thymidine kinase gene in transformed L cells by products of superinfecting virus. J Virol. 1976 Nov;20(2):413–424. doi: 10.1128/jvi.20.2.413-424.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Mackem S., Roizman B. Differentiation between alpha promoter and regulator regions of herpes simplex virus 1: the functional domains and sequence of a movable alpha regulator. Proc Natl Acad Sci U S A. 1982 Aug;79(16):4917–4921. doi: 10.1073/pnas.79.16.4917. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Mackem S., Roizman B. Regulation of alpha genes of herpes simplex virus: the alpha 27 gene promoter-thymidine kinase chimera is positively regulated in converted L cells. J Virol. 1982 Sep;43(3):1015–1023. doi: 10.1128/jvi.43.3.1015-1023.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Mackem S., Roizman B. Structural features of the herpes simplex virus alpha gene 4, 0, and 27 promoter-regulatory sequences which confer alpha regulation on chimeric thymidine kinase genes. J Virol. 1982 Dec;44(3):939–949. doi: 10.1128/jvi.44.3.939-949.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mao J. C., Robishaw E. E., Overby L. R. Inhibition of DNA polymerase from herpes simplex virus-infected wi-38 cells by phosphonoacetic Acid. J Virol. 1975 May;15(5):1281–1283. doi: 10.1128/jvi.15.5.1281-1283.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Marsden H. S., Stow N. D., Preston V. G., Timbury M. C., Wilkie N. M. Physical mapping of herpes simplex virus-induced polypeptides. J Virol. 1978 Nov;28(2):624–642. doi: 10.1128/jvi.28.2.624-642.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. McKnight S. L. The nucleotide sequence and transcript map of the herpes simplex virus thymidine kinase gene. Nucleic Acids Res. 1980 Dec 20;8(24):5949–5964. doi: 10.1093/nar/8.24.5949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Mocarski E. S., Post L. E., Roizman B. Molecular engineering of the herpes simplex virus genome: insertion of a second L-S junction into the genome causes additional genome inversions. Cell. 1980 Nov;22(1 Pt 1):243–255. doi: 10.1016/0092-8674(80)90172-5. [DOI] [PubMed] [Google Scholar]
  26. Overby L. R., Robishaw E. E., Schleicher J. B., Rueter A., Shipkowitz N. L., Mao J. C. Inhibition of herpes simplex virus replication by phosphonoacetic acid. Antimicrob Agents Chemother. 1974 Sep;6(3):360–365. doi: 10.1128/aac.6.3.360. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Para M. F., Baucke R. B., Spear P. G. Glycoprotein gE of herpes simplex virus type 1: effects of anti-gE on virion infectivity and on virus-induced fc-binding receptors. J Virol. 1982 Jan;41(1):129–136. doi: 10.1128/jvi.41.1.129-136.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Peake M. L., Nystrom P., Pizer L. I. Herpesvirus glycoprotein synthesis and insertion into plasma membranes. J Virol. 1982 May;42(2):678–690. doi: 10.1128/jvi.42.2.678-690.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Pereira L., Dondero D. V., Gallo D., Devlin V., Woodie J. D. Serological analysis of herpes simplex virus types 1 and 2 with monoclonal antibodies. Infect Immun. 1982 Jan;35(1):363–367. doi: 10.1128/iai.35.1.363-367.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Pereira L., Klassen T., Baringer J. R. Type-common and type-specific monoclonal antibody to herpes simplex virus type 1. Infect Immun. 1980 Aug;29(2):724–732. doi: 10.1128/iai.29.2.724-732.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Post L. E., Conley A. J., Mocarski E. S., Roizman B. Cloning of reiterated and nonreiterated herpes simplex virus 1 sequences as BamHI fragments. Proc Natl Acad Sci U S A. 1980 Jul;77(7):4201–4205. doi: 10.1073/pnas.77.7.4201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Post L. E., Mackem S., Roizman B. Regulation of alpha genes of herpes simplex virus: expression of chimeric genes produced by fusion of thymidine kinase with alpha gene promoters. Cell. 1981 May;24(2):555–565. doi: 10.1016/0092-8674(81)90346-9. [DOI] [PubMed] [Google Scholar]
  33. Post L. E., Roizman B. A generalized technique for deletion of specific genes in large genomes: alpha gene 22 of herpes simplex virus 1 is not essential for growth. Cell. 1981 Jul;25(1):227–232. doi: 10.1016/0092-8674(81)90247-6. [DOI] [PubMed] [Google Scholar]
  34. Ruyechan W. T., Morse L. S., Knipe D. M., Roizman B. Molecular genetics of herpes simplex virus. II. Mapping of the major viral glycoproteins and of the genetic loci specifying the social behavior of infected cells. J Virol. 1979 Feb;29(2):677–697. doi: 10.1128/jvi.29.2.677-697.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Showalter S. D., Zweig M., Hampar B. Monoclonal antibodies to herpes simplex virus type 1 proteins, including the immediate-early protein ICP 4. Infect Immun. 1981 Dec;34(3):684–692. doi: 10.1128/iai.34.3.684-692.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Sim C., Watson D. H. The role of type specific and cross reacting structural antigens in the neutralization of herpes simplex virus types 1 and 2. J Gen Virol. 1973 May;19(2):217–233. doi: 10.1099/0022-1317-19-2-217. [DOI] [PubMed] [Google Scholar]
  37. Smiley J. R., Fong B. S., Leung W. C. Construction of a double-jointed herpes simplex viral DNA molecule: inverted repeats are required for segment inversion, and direct repeats promote deletions. Virology. 1981 Aug;113(1):345–362. doi: 10.1016/0042-6822(81)90161-6. [DOI] [PubMed] [Google Scholar]
  38. Smith J. F., Brown D. T. Envelopments of Sindbis virus: synthesis and organization of proteins in cells infected with wild type and maturation-defective mutants. J Virol. 1977 Jun;22(3):662–678. doi: 10.1128/jvi.22.3.662-678.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Tognon M., Furlong D., Conley A. J., Roizman B. Molecular genetics of herpes simplex virus. V. Characterization of a mutant defective in ability to form plaques at low temperatures and in a viral fraction which prevents accumulation of coreless capsids at nuclear pores late in infection. J Virol. 1981 Dec;40(3):870–880. doi: 10.1128/jvi.40.3.870-880.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Wagner M. J., Sharp J. A., Summers W. C. Nucleotide sequence of the thymidine kinase gene of herpes simplex virus type 1. Proc Natl Acad Sci U S A. 1981 Mar;78(3):1441–1445. doi: 10.1073/pnas.78.3.1441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Watson D. H., Wildy P. The preparation of 'monoprecipitin' antisera to herpes virus specific antigens. J Gen Virol. 1969 Mar;4(2):163–168. doi: 10.1099/0022-1317-4-2-163. [DOI] [PubMed] [Google Scholar]
  42. Watson R. J., Colberg-Poley A. M., Marcus-Sekura C. J., Carter B. J., Enquist L. W. Characterization of the herpes simplex virus type 1 glycoprotein D mRNA and expression of this protein in Xenopus oocytes. Nucleic Acids Res. 1983 Mar 11;11(5):1507–1522. doi: 10.1093/nar/11.5.1507. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Watson R. J., Weis J. H., Salstrom J. S., Enquist L. W. Herpes simplex virus type-1 glycoprotein D gene: nucleotide sequence and expression in Escherichia coli. Science. 1982 Oct 22;218(4570):381–384. doi: 10.1126/science.6289440. [DOI] [PubMed] [Google Scholar]
  44. Weis J. H., Enquist L. W., Salstrom J. S., Watson R. J. An immunologically active chimaeric protein containing herpes simplex virus type 1 glycoprotein D. Nature. 1983 Mar 3;302(5903):72–74. doi: 10.1038/302072a0. [DOI] [PubMed] [Google Scholar]
  45. Wigler M., Pellicer A., Silverstein S., Axel R., Urlaub G., Chasin L. DNA-mediated transfer of the adenine phosphoribosyltransferase locus into mammalian cells. Proc Natl Acad Sci U S A. 1979 Mar;76(3):1373–1376. doi: 10.1073/pnas.76.3.1373. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES