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Characterization of a herpes simplex virus regulatory protein: Aggregation and phosphorylation of a temperature-sensitive variant of ICP 4

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Summary

The viral polypeptide ICP 4 (or Vmw 175) is synthesized during the immediate early phase of infection by herpes simplex virus (HSV) and is required during the viral reproductive cycle for efficient transcription of delayed early viral genes. Replication of mutant strains of HSV-1 such as tsLB 2 that encode a temperature-sensitive variant of ICP 4 does not proceed beyond the immediate early phase in cells that are infected and maintained at the nonpermissive temperature (NPT). Under these conditions, the immediate early viral polypeptides accumulate to levels that are 10 to 100 fold greater than normal. We have investigated the use of tsLB 2-infected cells maintained at the NPT as a source for substantial amounts of ICP 4 for further characterization. Extraction of ICP 4 from tsLB 2-infected cells requires 0.5m NaCl and yields aggregates that contain ICP 4, ICP 6, ICP 27, and lesser amounts of other proteins. These large aggregates cannot be disrupted under nondenaturing conditions and thus are not a suitable source for native ICP 4. We have used this overproduced ICP 4 as an antigen to generate ICP 4-specific antibody and for characterization of the primary structure of ICP 4. Analysis of acid-hydrolysed32P-labeled ICP 4 revealed that the major phosphorylated residues in ICP 4 are phosphoserine and phosphothreonine.

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References

  1. Boyer PD, Deluca M, Ebner K, Hultquist D, Peter J (1962) Identification of phosphohistidine in digests from a probable intermediate of oxidative phosphorylation. J Biol Chem 237: PC 3306–3308

    Google Scholar 

  2. Brown SM, Ritchie DA, Subak-Sharpe HH (1973) Genetic studies with herpes simplex virus type 1. The isolation of temperature-sensitive mutants, their arrangement into complementation groups, and recombination analysis leading to a linkage map. J Gen Virol 18: 329–346

    Google Scholar 

  3. Burnette W (1981) “Western Blotting”: Electrophoretic transfer of proteins from sodium dodecyl sulfate-polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem 112: 195–203

    Google Scholar 

  4. Cabral GA, Courtney RJ, Schaffer PA, Marciano-Cabral F (1980) Ultra-structural characterization of an early, nonstructural polypeptide of herpes simplex virus type 1. J Virol 33: 1192–1198

    Google Scholar 

  5. Courtney RJ, Benyesh-Melnick M (1974) Isolation and characterization of a large molecular weight polypeptide of herpes simplex virus type 1. Virology 62: 539–551

    Google Scholar 

  6. Courtney RJ, Schaffer P, Powell K (1976) Synthesis of virus-specific polypeptides by temperature-sensitive mutants of herpes simplex virus type 1. Virology 75: 306–318

    Google Scholar 

  7. Davison MJ, Preston V, McGeoch DJ (1984) Determination of the sequence alteration in the DNA of the herpes simplex virus type 1 temperature-sensitive mutant tsK. J Gen Virol 65: 859–863

    Google Scholar 

  8. Deluca NA, Courtney MA, Schaffer PA (1984) Temperature-sensitive mutants of herpes simplex virus type 1 ICP 4 permissive for early gene expression. J Virol 52: 767–776

    Google Scholar 

  9. Dixon R, Schaffer P (1980) Fine-structure mapping and functional analysis of temperature-sensitive mutants in the gene encoding the herpes simplex virus type 1 immediate early protein VP 175. J Virol 36: 189–203

    Google Scholar 

  10. Dulbecco R, Vogt M (1954) Plaque formation and isolation of pure lines with poliomyelitis virus. J Exp Med 99: 167–182

    Google Scholar 

  11. Ejercito P, Kieff E, Roizman B (1968) Characterization of herpes simplex virus strains differing in their effect on social behavior of infected cells. J Gen Virol 3: 357–364

    Google Scholar 

  12. Fenwick ML, Walker MJ (1979) Phosphorylation of a ribosomal protein and of virus-specific proteins in cells infected with herpes simplex virus. J Gen Virol 45: 397–405

    Google Scholar 

  13. Fenwick ML, Walker MJ, Petkevich JM (1978) On the association of virus proteins with the nuclei of cells infected with herpes simplex virus. J Gen Virol 39: 519–529

    Google Scholar 

  14. Gabriel O (1971) Locating enzymes on gels. In:Jackoby W (ed) Methods in enzymology, vol XXII. Academic Press, New York, pp 583–584

    Google Scholar 

  15. Halliburton I, Randall R, Killington R, Watson D (1977) Some properties of recombinants between type 1 and type 2 herpes simplex viruses. J Gen Virol 36: 471–484

    Google Scholar 

  16. Heine J, Honess R, Cassai E, Roizman B (1974) Proteins specified by herpes simplex virus. XII. The virion polypeptides of type 1 strains. J Virol 14: 640–651

    Google Scholar 

  17. Honess R, Roizman B (1974) Regulation of herpesvirus macromolecular synthesis. I. Cascade regulation of the synthesis of three groups of viral proteins. J Virol 12: 1347–1365

    Google Scholar 

  18. Hunter T, Sefton BM (1980) Transforming gene product of Rous sarcoma virus phosphorylates tyrosine. Proc Natl Acad Sci USA 77: 1311–1315

    Google Scholar 

  19. Kabat D (1970) Phosphorylation of ribosomal proteins in rabbit reticulocytes. Characterization and regulatory aspects. Biochem 9: 4160–4175

    Google Scholar 

  20. Kessler SW (1975) 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 Immun 115: 1617–1624

    Google Scholar 

  21. Klotz IM, Haney DN, King LC (1981) Rational approaches to chemotherapy: antisickling agents. Science 213: 724–731

    Google Scholar 

  22. Lee FY, Nomoto A, Detjen BM, Wimmer E (1977) A protein covalently linked to poliovirus genome RNA. Proc Natl Acad Sci USA 74: 59–63

    Google Scholar 

  23. Marsden HS, Chrombie IK, Subak-Sharpe JH (1976) Control of protein synthesis in herpes virus-infected cells: analysis of the polypeptides induced by wild type and sixteen temperature-sensitive mutants of HSV strain 17. J Gen Virol 31: 347–372

    Google Scholar 

  24. Marsden HS, Stow N, Preston V, Timbury M, Wilkie N (1978) Physical mapping of herpes simplex virus-induced polypeptides. J Virol 28: 624–642

    Google Scholar 

  25. Metzler DW, Wilcox KW (1985) Isolation of herpes simplex virus regulatory protein ICP 4 as a homodimeric complex. J Virol 55: 329–337

    Google Scholar 

  26. Palfreyman JW, MacLean JB, Messeder E, Sheppard RC (1984) Successful use of oligopeptides as immunogens in the preparation of antisera to immediate-early gene products of herpes simplex virus type 1. J Gen Virol 65: 865–874

    Google Scholar 

  27. Pereira L, Wolff M, Fenwick M, Roizman B (1977) Regulation of herpesvirus macromolecular synthesis. V. Properties of α polypeptides made in HSV-1 and HSV-2 infected cells. Virology 77: 733–749

    Google Scholar 

  28. Plimmer RH (1941) Esters of phosphoric acid. 4. Phosphoryl hydroxyamino acids. Biochem J 35: 461–469

    Google Scholar 

  29. Powell KL, Purifoy DJM (1976) DNA-binding proteins of cells infected by herpes simplex virus type 1 and 2. Intervirology 7: 225–239

    Google Scholar 

  30. Preston CM (1979) Control of herpes simplex virus type 1 mRNA synthesis in cells infected with wild-type virus or the temperature-sensitive mutant tsK. J Virol 29: 275–284

    Google Scholar 

  31. Preston CM (1979) Abnormal properties of an immediate early polypeptide in cells infected with the herpes simplex virus type 1 mutant tsK. J Virol 32: 357–369

    Google Scholar 

  32. Preston CM (1981) Fine-structure mapping of herpes simplex virus type 1 temperature-sensitive mutations within the short repeat region of the genome. J Virol 39: 150–161

    Google Scholar 

  33. Preston CM, Notarianni EL (1983) Poly(ADP-ribosyl)ation of a herpes simplex virus immediate early polypeptide. Virology 131: 492–501

    Google Scholar 

  34. Prouty WF, Karnovsky NJ, Goldberg AL (1975) Degradation of abnormal proteins inEscherichia coli. J Biol Chem 250: 1112–1122

    Google Scholar 

  35. Purnell M, Stone P, Whish W (1980) ADP-ribosylation of nuclear proteins. Biochem Soc Trans 8: 215–227

    Google Scholar 

  36. Schaffer PA (1975) Temperature-sensitive mutants of herpesviruses. Curr Top Microb Immunol 70: 51–100

    Google Scholar 

  37. Schaffer PA, Carter VC, Timbury MC (1978) Collaborative complementation study of temperature-sensitive mutants of herpes simplex virus types 1 and 2. J Virol 27: 490–504

    Google Scholar 

  38. Threfall CJ (1957) An analytical procedure for the acid-soluble phosphorus compounds in rat-skeletal muscle. Biochem J 65: 694–699

    Google Scholar 

  39. Watson RJ, Clements JB (1978) Characterization of transcription deficient temperature-sensitive mutants of herpes simplex virus type 1. Virology 91: 364–379

    Google Scholar 

  40. Wilcox KW, Kohn A, Sklyanskaya E, Roizman B (1980) Herpes simplex virus phosphoproteins. I. Phosphate cycles on and off some viral polypeptides and can alter their affinity for DNA. J Virol 33: 167–182

    Google Scholar 

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  1. Department of Microbiology, Medical College of Wisconsin, Milwaukee, Wisconsin, USA

    S. W. Faber & K. W. Wilcox

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  1. S. W. Faber
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  2. K. W. Wilcox
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Faber, S.W., Wilcox, K.W. Characterization of a herpes simplex virus regulatory protein: Aggregation and phosphorylation of a temperature-sensitive variant of ICP 4. Archives of Virology 91, 297–312 (1986). https://doi.org/10.1007/BF01314289

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  • DOI: https://doi.org/10.1007/BF01314289

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