Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1988 Feb;62(2):519–527. doi: 10.1128/jvi.62.2.519-527.1988

Vaccinia virus-encoded ribonucleotide reductase: sequence conservation of the gene for the small subunit and its amplification in hydroxyurea-resistant mutants.

M Slabaugh 1, N Roseman 1, R Davis 1, C Mathews 1
PMCID: PMC250563  PMID: 2826813

Abstract

The vaccinia virus gene that encodes the small subunit of ribonucleotide reductase was localized to the HindIII F fragment by using degenerate oligonucleotide probes. DNA sequencing revealed a leftward-reading open reading frame that predicted a protein of 37 kilodaltons whose amino acid sequence was much more homologous to the mouse and clam M2 sequences (approximately 80%) than to the corresponding herpesvirus (approximately 27%) or procaryotic (approximately 19%) gene products. Vaccinia virus mutants selected for the ability to grow in high concentrations of a specific inhibitor of ribonucleotide reductase, hydroxyurea, amplified the M2 gene and harbored tandem arrays (2 to 15 copies) of the gene within the HindIII F region. RNA isolated at early times after infection with wild-type virus and probed with an internal fragment of the M2 gene indicated one major (1.2 kilobases) and two minor (4.0 and 2.1 kilobases) transcripts. S1 nuclease analysis and primer extension experiments identified an RNA start site 12 nucleotides upstream of the putative initiation ATG codon.

Full text

PDF
521

Images in this article

521Image
on p.521
522Image
on p.522
523Image
on p.523
524Image
on p.524
525Image
on p.525

Selected References

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

  1. Averett D. R., Lubbers C., Elion G. B., Spector T. Ribonucleotide reductase induced by herpes simplex type 1 virus. Characterization of a distinct enzyme. J Biol Chem. 1983 Aug 25;258(16):9831–9838. [PubMed] [Google Scholar]
  2. Ball L. A. High-frequency homologous recombination in vaccinia virus DNA. J Virol. 1987 Jun;61(6):1788–1795. doi: 10.1128/jvi.61.6.1788-1795.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Baltimore D. Retroviruses and retrotransposons: the role of reverse transcription in shaping the eukaryotic genome. Cell. 1985 Mar;40(3):481–482. doi: 10.1016/0092-8674(85)90190-4. [DOI] [PubMed] [Google Scholar]
  4. Berglund O. Ribonucleoside diphosphate reductase induced by bacteriophage T4. II. Allosteric regulation of substrate sepecificity and catalytic activity. J Biol Chem. 1972 Nov 25;247(22):7276–7281. [PubMed] [Google Scholar]
  5. Bertholet C., Van Meir E., ten Heggeler-Bordier B., Wittek R. Vaccinia virus produces late mRNAs by discontinuous synthesis. Cell. 1987 Jul 17;50(2):153–162. doi: 10.1016/0092-8674(87)90211-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Broyles S. S., Moss B. Homology between RNA polymerases of poxviruses, prokaryotes, and eukaryotes: nucleotide sequence and transcriptional analysis of vaccinia virus genes encoding 147-kDa and 22-kDa subunits. Proc Natl Acad Sci U S A. 1986 May;83(10):3141–3145. doi: 10.1073/pnas.83.10.3141. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Carlson J., Fuchs J. A., Messing J. Primary structure of the Escherichia coli ribonucleoside diphosphate reductase operon. Proc Natl Acad Sci U S A. 1984 Jul;81(14):4294–4297. doi: 10.1073/pnas.81.14.4294. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Condit R. C., Motyczka A., Spizz G. Isolation, characterization, and physical mapping of temperature-sensitive mutants of vaccinia virus. Virology. 1983 Jul 30;128(2):429–443. doi: 10.1016/0042-6822(83)90268-4. [DOI] [PubMed] [Google Scholar]
  9. DeFilippes F. M. Restriction enzyme mapping of vaccinia virus DNA. J Virol. 1982 Jul;43(1):136–149. doi: 10.1128/jvi.43.1.136-149.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Elledge S. J., Davis R. W. Identification and isolation of the gene encoding the small subunit of ribonucleotide reductase from Saccharomyces cerevisiae: DNA damage-inducible gene required for mitotic viability. Mol Cell Biol. 1987 Aug;7(8):2783–2793. doi: 10.1128/mcb.7.8.2783. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Eriksson S., Gräslund A., Skog S., Thelander L., Tribukait B. Cell cycle-dependent regulation of mammalian ribonucleotide reductase. The S phase-correlated increase in subunit M2 is regulated by de novo protein synthesis. J Biol Chem. 1984 Oct 10;259(19):11695–11700. [PubMed] [Google Scholar]
  12. Esposito J., Condit R., Obijeski J. The preparation of orthopoxvirus DNA. J Virol Methods. 1981 Feb;2(3):175–179. doi: 10.1016/0166-0934(81)90036-7. [DOI] [PubMed] [Google Scholar]
  13. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  14. Fitch W. M., Margoliash E. Construction of phylogenetic trees. Science. 1967 Jan 20;155(3760):279–284. doi: 10.1126/science.155.3760.279. [DOI] [PubMed] [Google Scholar]
  15. Gibson T., Stockwell P., Ginsburg M., Barrell B. Homology between two EBV early genes and HSV ribonucleotide reductase and 38K genes. Nucleic Acids Res. 1984 Jun 25;12(12):5087–5099. doi: 10.1093/nar/12.12.5087. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gotoh O. Pattern matching of biological sequences with limited storage. Comput Appl Biosci. 1987 Mar;3(1):17–20. doi: 10.1093/bioinformatics/3.1.17. [DOI] [PubMed] [Google Scholar]
  17. Groyon R. M., Kniazeff A. J. Vaccinia virus infection of synchronized pig kidney cells. J Virol. 1967 Dec;1(6):1255–1264. doi: 10.1128/jvi.1.6.1255-1264.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Larsson A., Sjöberg B. M. Identification of the stable free radical tyrosine residue in ribonucleotide reductase. EMBO J. 1986 Aug;5(8):2037–2040. doi: 10.1002/j.1460-2075.1986.tb04461.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lehrach H., Diamond D., Wozney J. M., Boedtker H. RNA molecular weight determinations by gel electrophoresis under denaturing conditions, a critical reexamination. Biochemistry. 1977 Oct 18;16(21):4743–4751. doi: 10.1021/bi00640a033. [DOI] [PubMed] [Google Scholar]
  20. Linial M. Creation of a processed pseudogene by retroviral infection. Cell. 1987 Apr 10;49(1):93–102. doi: 10.1016/0092-8674(87)90759-8. [DOI] [PubMed] [Google Scholar]
  21. McLauchlan J., Clements J. B. A 3' co-terminus of two early herpes simplex virus type 1 mRNAs. Nucleic Acids Res. 1982 Jan 22;10(2):501–512. doi: 10.1093/nar/10.2.501. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Moyer R. W., Graves R. L. The mechanism of cytoplasmic orthopoxvirus DNA replication. Cell. 1981 Dec;27(2 Pt 1):391–401. doi: 10.1016/0092-8674(81)90422-0. [DOI] [PubMed] [Google Scholar]
  23. Needleman S. B., Wunsch C. D. A general method applicable to the search for similarities in the amino acid sequence of two proteins. J Mol Biol. 1970 Mar;48(3):443–453. doi: 10.1016/0022-2836(70)90057-4. [DOI] [PubMed] [Google Scholar]
  24. Nikas I., McLauchlan J., Davison A. J., Taylor W. R., Clements J. B. Structural features of ribonucleotide reductase. Proteins. 1986 Dec;1(4):376–384. doi: 10.1002/prot.340010411. [DOI] [PubMed] [Google Scholar]
  25. Niles E. G., Condit R. C., Caro P., Davidson K., Matusick L., Seto J. Nucleotide sequence and genetic map of the 16-kb vaccinia virus HindIII D fragment. Virology. 1986 Aug;153(1):96–112. doi: 10.1016/0042-6822(86)90011-5. [DOI] [PubMed] [Google Scholar]
  26. Perkus M. E., Panicali D., Mercer S., Paoletti E. Insertion and deletion mutants of vaccinia virus. Virology. 1986 Jul 30;152(2):285–297. doi: 10.1016/0042-6822(86)90132-7. [DOI] [PubMed] [Google Scholar]
  27. Rosel J. L., Earl P. L., Weir J. P., Moss B. Conserved TAAATG sequence at the transcriptional and translational initiation sites of vaccinia virus late genes deduced by structural and functional analysis of the HindIII H genome fragment. J Virol. 1986 Nov;60(2):436–449. doi: 10.1128/jvi.60.2.436-449.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Roseman N. A., Hruby D. E. Nucleotide sequence and transcript organization of a region of the vaccinia virus genome which encodes a constitutively expressed gene required for DNA replication. J Virol. 1987 May;61(5):1398–1406. doi: 10.1128/jvi.61.5.1398-1406.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. 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]
  30. Schimke R. T. Gene amplification in cultured animal cells. Cell. 1984 Jul;37(3):705–713. doi: 10.1016/0092-8674(84)90406-9. [DOI] [PubMed] [Google Scholar]
  31. Sjöberg B. M., Eklund H., Fuchs J. A., Carlson J., Standart N. M., Ruderman J. V., Bray S. J., Hunt T. Identification of the stable free radical tyrosine residue in ribonucleotide reductase. A sequence comparison. FEBS Lett. 1985 Apr 8;183(1):99–102. doi: 10.1016/0014-5793(85)80962-5. [DOI] [PubMed] [Google Scholar]
  32. Sjöberg B. M., Hahne S., Mathews C. Z., Mathews C. K., Rand K. N., Gait M. J. The bacteriophage T4 gene for the small subunit of ribonucleotide reductase contains an intron. EMBO J. 1986 Aug;5(8):2031–2036. doi: 10.1002/j.1460-2075.1986.tb04460.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Slabaugh M. B., Johnson T. L., Mathews C. K. Vaccinia virus induces ribonucleotide reductase in primate cells. J Virol. 1984 Nov;52(2):507–514. doi: 10.1128/jvi.52.2.507-514.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Slabaugh M. B., Mathews C. K. Hydroxyurea-resistant vaccinia virus: overproduction of ribonucleotide reductase. J Virol. 1986 Nov;60(2):506–514. doi: 10.1128/jvi.60.2.506-514.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Slabaugh M. B., Mathews C. K. Vaccinia virus-induced ribonucleotide reductase can be distinguished from host cell activity. J Virol. 1984 Nov;52(2):501–506. doi: 10.1128/jvi.52.2.501-506.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Standart N. M., Bray S. J., George E. L., Hunt T., Ruderman J. V. The small subunit of ribonucleotide reductase is encoded by one of the most abundant translationally regulated maternal RNAs in clam and sea urchin eggs. J Cell Biol. 1985 Jun;100(6):1968–1976. doi: 10.1083/jcb.100.6.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Standart N., Hunt T., Ruderman J. V. Differential accumulation of ribonucleotide reductase subunits in clam oocytes: the large subunit is stored as a polypeptide, the small subunit as untranslated mRNA. J Cell Biol. 1986 Dec;103(6 Pt 1):2129–2136. doi: 10.1083/jcb.103.6.2129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Stark G. R., Wahl G. M. Gene amplification. Annu Rev Biochem. 1984;53:447–491. doi: 10.1146/annurev.bi.53.070184.002311. [DOI] [PubMed] [Google Scholar]
  39. Tabor S., Richardson C. C. DNA sequence analysis with a modified bacteriophage T7 DNA polymerase. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4767–4771. doi: 10.1073/pnas.84.14.4767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Thelander L., Berg P. Isolation and characterization of expressible cDNA clones encoding the M1 and M2 subunits of mouse ribonucleotide reductase. Mol Cell Biol. 1986 Oct;6(10):3433–3442. doi: 10.1128/mcb.6.10.3433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Upton C., McFadden G. Identification and nucleotide sequence of the thymidine kinase gene of Shope fibroma virus. J Virol. 1986 Dec;60(3):920–927. doi: 10.1128/jvi.60.3.920-927.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Weinrich S. L., Niles E. G., Hruby D. E. Transcriptional and translational analysis of the vaccinia virus late gene L65. J Virol. 1985 Aug;55(2):450–457. doi: 10.1128/jvi.55.2.450-457.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Wright J. A., Alam T. G., McClarty G. A., Tagger A. Y., Thelander L. Altered expression of ribonucleotide reductase and role of M2 gene amplification in hydroxyurea-resistant hamster, mouse, rat, and human cell lines. Somat Cell Mol Genet. 1987 Mar;13(2):155–165. doi: 10.1007/BF01534695. [DOI] [PubMed] [Google Scholar]
  44. Wu D. G., Black L. W. Gene amplification mechanism for the hyperproduction of T4 bacteriophage gene 17 and 18 proteins. J Mol Biol. 1987 Jun 20;195(4):769–783. doi: 10.1016/0022-2836(87)90483-9. [DOI] [PubMed] [Google Scholar]

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

RESOURCES