Skip to main content
PMC home page
Journal of Virology logoLink to Journal of Virology
. 1989 Mar;63(3):1076–1086. doi: 10.1128/jvi.63.3.1076-1086.1989

The second-largest subunit of the poxvirus RNA polymerase is similar to the corresponding subunits of procaryotic and eucaryotic RNA polymerases.

D D Patel 1, D J Pickup 1
PMCID: PMC247801  PMID: 2915377

Abstract

We have characterized the poxvirus gene encoding the second-largest subunit of the viral DNA-dependent RNA polymerase. This gene, designated rpo132, is located in the HindIII A fragment of the DNA of the Brighton Red strain of cowpox virus. A similar gene is located in the corresponding position in the HindIII A fragment of the DNA of the Western Reserve strain of vaccinia virus. The rpo132 gene is transcribed throughout the viral multiplication cycle. It has two transcriptional start sites; one is operative at late times only, and the other (80 base pairs downstream) is operative both at early times and at late times. Neither early nor late transcripts originating from the latter RNA start site contain long 5'-terminal poly(A) sequences. The rpo132 gene has the capacity to encode primary gene products of two types. The RNA transcripts whose 5' ends correspond to the early RNA start site can encode a 133-kilodalton (kDa) protein. The RNA transcripts whose 5' ends correspond to the early RNA start site can encode a 132-kDa protein. Transcripts of the latter type are more abundant, suggesting that the 132-kDa protein is the major primary product of this gene. The predicted amino acid sequences of both gene products share extensive similarities with the amino acid sequences of the second-largest subunits of the following enzymes: the RNA polymerase of Escherichia coli, the RNA polymerase II of Saccharomyces cerevisiae, and the RNA polymerase II of Drosophila melanogaster. This result provides further evidence of relatedness between multisubunit DNA-dependent RNA polymerases.

Full text

PDF
1086

Images in this article

1080Image
on p.1080
1080Image
on p.1080
1081Image
on p.1081
1082Image
on p.1082

Selected References

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

  1. Archard L. C., Mackett M., Barnes D. E., Dumbell K. R. The genome structure of cowpox virus white pock variants. J Gen Virol. 1984 May;65(Pt 5):875–886. doi: 10.1099/0022-1317-65-5-875. [DOI] [PubMed] [Google Scholar]
  2. Baroudy B. M., Moss B. Purification and characterization of a DNA-dependent RNA polymerase from vaccinia virions. J Biol Chem. 1980 May 10;255(9):4372–4380. [PubMed] [Google Scholar]
  3. Berg J. M. Potential metal-binding domains in nucleic acid binding proteins. Science. 1986 Apr 25;232(4749):485–487. doi: 10.1126/science.2421409. [DOI] [PubMed] [Google Scholar]
  4. Berk A. J., Sharp P. A. Sizing and mapping of early adenovirus mRNAs by gel electrophoresis of S1 endonuclease-digested hybrids. Cell. 1977 Nov;12(3):721–732. doi: 10.1016/0092-8674(77)90272-0. [DOI] [PubMed] [Google Scholar]
  5. Bertholet C., Drillien R., Wittek R. One hundred base pairs of 5' flanking sequence of a vaccinia virus late gene are sufficient to temporally regulate late transcription. Proc Natl Acad Sci U S A. 1985 Apr;82(7):2096–2100. doi: 10.1073/pnas.82.7.2096. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bertholet C., Stocco P., Van Meir E., Wittek R. Functional analysis of the 5' flanking sequence of a vaccinia virus late gene. EMBO J. 1986 Aug;5(8):1951–1957. doi: 10.1002/j.1460-2075.1986.tb04449.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. 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]
  8. Biggin M. D., Gibson T. J., Hong G. F. Buffer gradient gels and 35S label as an aid to rapid DNA sequence determination. Proc Natl Acad Sci U S A. 1983 Jul;80(13):3963–3965. doi: 10.1073/pnas.80.13.3963. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. 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]
  10. Caruthers M. H., Barone A. D., Beaucage S. L., Dodds D. R., Fisher E. F., McBride L. J., Matteucci M., Stabinsky Z., Tang J. Y. Chemical synthesis of deoxyoligonucleotides by the phosphoramidite method. Methods Enzymol. 1987;154:287–313. doi: 10.1016/0076-6879(87)54081-2. [DOI] [PubMed] [Google Scholar]
  11. Chambon P. Eukaryotic nuclear RNA polymerases. Annu Rev Biochem. 1975;44:613–638. doi: 10.1146/annurev.bi.44.070175.003145. [DOI] [PubMed] [Google Scholar]
  12. Cochran M. A., Puckett C., Moss B. In vitro mutagenesis of the promoter region for a vaccinia virus gene: evidence for tandem early and late regulatory signals. J Virol. 1985 Apr;54(1):30–37. doi: 10.1128/jvi.54.1.30-37.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Culp J. S., Webster L. C., Friedman D. J., Smith C. L., Huang W. J., Wu F. Y., Rosenberg M., Ricciardi R. P. The 289-amino acid E1A protein of adenovirus binds zinc in a region that is important for trans-activation. Proc Natl Acad Sci U S A. 1988 Sep;85(17):6450–6454. doi: 10.1073/pnas.85.17.6450. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dynan W. S., Tjian R. Control of eukaryotic messenger RNA synthesis by sequence-specific DNA-binding proteins. 1985 Aug 29-Sep 4Nature. 316(6031):774–778. doi: 10.1038/316774a0. [DOI] [PubMed] [Google Scholar]
  15. Earl P. L., Jones E. V., Moss B. Homology between DNA polymerases of poxviruses, herpesviruses, and adenoviruses: nucleotide sequence of the vaccinia virus DNA polymerase gene. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3659–3663. doi: 10.1073/pnas.83.11.3659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ensinger M. J. Phenotypic characterization of temperature-sensitive mutants of vaccinia virus with mutations in a 135,000-Mr subunit of the virion-associated DNA-dependent RNA polymerase. J Virol. 1987 Jun;61(6):1842–1850. doi: 10.1128/jvi.61.6.1842-1850.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Esposito J. J., Knight J. C. Orthopoxvirus DNA: a comparison of restriction profiles and maps. Virology. 1985 May;143(1):230–251. doi: 10.1016/0042-6822(85)90111-4. [DOI] [PubMed] [Google Scholar]
  18. Evans R. M., Hollenberg S. M. Zinc fingers: gilt by association. Cell. 1988 Jan 15;52(1):1–3. doi: 10.1016/0092-8674(88)90522-3. [DOI] [PubMed] [Google Scholar]
  19. Falkenburg D., Dworniczak B., Faust D. M., Bautz E. K. RNA polymerase II of Drosophila. Relation of its 140,000 Mr subunit to the beta subunit of Escherichia coli RNA polymerase. J Mol Biol. 1987 Jun 20;195(4):929–937. doi: 10.1016/0022-2836(87)90496-7. [DOI] [PubMed] [Google Scholar]
  20. Fritz H. J., Belagaje R., Brown E. L., Fritz R. H., Jones R. A., Lees R. G., Khorana H. G. High-pressure liquid chromatography in polynucleotide synthesis. Biochemistry. 1978 Apr 4;17(7):1257–1267. doi: 10.1021/bi00600a020. [DOI] [PubMed] [Google Scholar]
  21. Funahashi S., Sato T., Shida H. Cloning and characterization of the gene encoding the major protein of the A-type inclusion body of cowpox virus. J Gen Virol. 1988 Jan;69(Pt 1):35–47. doi: 10.1099/0022-1317-69-1-35. [DOI] [PubMed] [Google Scholar]
  22. Ghosh P. K., Reddy V. B., Swinscoe J., Lebowitz P., Weissman S. M. Heterogeneity and 5'-terminal structures of the late RNAs of simian virus 40. J Mol Biol. 1978 Dec 25;126(4):813–846. doi: 10.1016/0022-2836(78)90022-0. [DOI] [PubMed] [Google Scholar]
  23. Glass R. E., Jones S. T., Nene V., Nomura T., Fujita N., Ishihama A. Genetic studies on the beta subunit of Escherichia coli RNA polymerase. VIII. Localisation of a region involved in promoter selectivity. Mol Gen Genet. 1986 Jun;203(3):487–491. doi: 10.1007/BF00422074. [DOI] [PubMed] [Google Scholar]
  24. Godovikova T. S., Grachev M. A., Kutyavin I. V., Tsarev I. G., Zarytova V. F., Zaychikov E. F. Studies of the functional topography of Escherichia coli RNA polymerase. Affinity labelling of RNA polymerase in a promoter complex by phosphorylating derivatives of primer oligonucleotides. Eur J Biochem. 1987 Aug 3;166(3):611–616. doi: 10.1111/j.1432-1033.1987.tb13557.x. [DOI] [PubMed] [Google Scholar]
  25. Hooda-Dhingra U., Thompson C. L., Condit R. C. Detailed phenotypic characterization of five temperature-sensitive mutants in the 22- and 147-kilodalton subunits of vaccinia virus DNA-dependent RNA polymerase. J Virol. 1989 Feb;63(2):714–729. doi: 10.1128/jvi.63.2.714-729.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Hänggi M., Bannwarth W., Stunnenberg H. G. Conserved TAAAT motif in vaccinia virus late promoters: overlapping TATA box and site of transcription initiation. EMBO J. 1986 May;5(5):1071–1076. doi: 10.1002/j.1460-2075.1986.tb04324.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. JOKLIK W. K. The purification fo four strains of poxvirus. Virology. 1962 Sep;18:9–18. doi: 10.1016/0042-6822(62)90172-1. [DOI] [PubMed] [Google Scholar]
  28. Jones E. V., Puckett C., Moss B. DNA-dependent RNA polymerase subunits encoded within the vaccinia virus genome. J Virol. 1987 Jun;61(6):1765–1771. doi: 10.1128/jvi.61.6.1765-1771.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Kates J. R., McAuslan B. R. Poxvirus DNA-dependent RNA polymerase. Proc Natl Acad Sci U S A. 1967 Jul;58(1):134–141. doi: 10.1073/pnas.58.1.134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kates J., Beeson J. Ribonucleic acid synthesis in vaccinia virus. II. Synthesis of polyriboadenylic acid. J Mol Biol. 1970 May 28;50(1):19–33. doi: 10.1016/0022-2836(70)90101-4. [DOI] [PubMed] [Google Scholar]
  31. Kozak M. Point mutations define a sequence flanking the AUG initiator codon that modulates translation by eukaryotic ribosomes. Cell. 1986 Jan 31;44(2):283–292. doi: 10.1016/0092-8674(86)90762-2. [DOI] [PubMed] [Google Scholar]
  32. Lee-Chen G. J., Bourgeois N., Davidson K., Condit R. C., Niles E. G. Structure of the transcription initiation and termination sequences of seven early genes in the vaccinia virus HindIII D fragment. Virology. 1988 Mar;163(1):64–79. doi: 10.1016/0042-6822(88)90234-6. [DOI] [PubMed] [Google Scholar]
  33. Lee-Chen G. J., Niles E. G. Map positions of the 5' ends of eight mRNAs synthesized from the late genes in the vaccinia virus HindIII D fragment. Virology. 1988 Mar;163(1):80–92. doi: 10.1016/0042-6822(88)90235-8. [DOI] [PubMed] [Google Scholar]
  34. Mackett M., Archard L. C. Conservation and variation in Orthopoxvirus genome structure. J Gen Virol. 1979 Dec;45(3):683–701. doi: 10.1099/0022-1317-45-3-683. [DOI] [PubMed] [Google Scholar]
  35. Maizel J. V., Jr, Lenk R. P. Enhanced graphic matrix analysis of nucleic acid and protein sequences. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7665–7669. doi: 10.1073/pnas.78.12.7665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Morrison D. K., Carter J. K., Moyer R. W. Isolation and characterization of monoclonal antibodies directed against two subunits of rabbit poxvirus-associated, DNA-directed RNA polymerase. J Virol. 1985 Sep;55(3):670–680. doi: 10.1128/jvi.55.3.670-680.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Moss B., Salzman N. P. Sequential protein synthesis following vaccinia virus infection. J Virol. 1968 Oct;2(10):1016–1027. doi: 10.1128/jvi.2.10.1016-1027.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Munyon W., Paoletti E., Grace J. T., Jr RNA polymerase activity in purified infectious vaccinia virus. Proc Natl Acad Sci U S A. 1967 Dec;58(6):2280–2287. doi: 10.1073/pnas.58.6.2280. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. 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]
  40. Nevins J. R., Joklik W. K. Isolation and properties of the vaccinia virus DNA-dependent RNA polymerase. J Biol Chem. 1977 Oct 10;252(19):6930–6938. [PubMed] [Google Scholar]
  41. Norrander J., Kempe T., Messing J. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene. 1983 Dec;26(1):101–106. doi: 10.1016/0378-1119(83)90040-9. [DOI] [PubMed] [Google Scholar]
  42. Ovchinnikov Y. A., Monastyrskaya G. S., Gubanov V. V., Guryev S. O., Chertov OYu, Modyanov N. N., Grinkevich V. A., Makarova I. A., Marchenko T. V., Polovnikova I. N. The primary structure of Escherichia coli RNA polymerase. Nucleotide sequence of the rpoB gene and amino-acid sequence of the beta-subunit. Eur J Biochem. 1981 Jun 1;116(3):621–629. doi: 10.1111/j.1432-1033.1981.tb05381.x. [DOI] [PubMed] [Google Scholar]
  43. Panka D., Dennis D. RNA polymerase. Direct evidence for two active sites involved in transcription. J Biol Chem. 1985 Feb 10;260(3):1427–1431. [PubMed] [Google Scholar]
  44. Patel D. D., Pickup D. J., Joklik W. K. Isolation of cowpox virus A-type inclusions and characterization of their major protein component. Virology. 1986 Mar;149(2):174–189. doi: 10.1016/0042-6822(86)90119-4. [DOI] [PubMed] [Google Scholar]
  45. Patel D. D., Pickup D. J. Messenger RNAs of a strongly-expressed late gene of cowpox virus contain 5'-terminal poly(A) sequences. EMBO J. 1987 Dec 1;6(12):3787–3794. doi: 10.1002/j.1460-2075.1987.tb02714.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Pennington T. H. Vaccinia virus polypeptide synthesis: sequential appearance and stability of pre- and post-replicative polypeptides. J Gen Virol. 1974 Dec;25(3):433–444. doi: 10.1099/0022-1317-25-3-433. [DOI] [PubMed] [Google Scholar]
  47. Pickup D. J., Ink B. S., Parsons B. L., Hu W., Joklik W. K. Spontaneous deletions and duplications of sequences in the genome of cowpox virus. Proc Natl Acad Sci U S A. 1984 Nov;81(21):6817–6821. doi: 10.1073/pnas.81.21.6817. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Plucienniczak A., Schroeder E., Zettlmeissl G., Streeck R. E. Nucleotide sequence of a cluster of early and late genes in a conserved segment of the vaccinia virus genome. Nucleic Acids Res. 1985 Feb 11;13(3):985–998. doi: 10.1093/nar/13.3.985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. 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]
  50. Sanger F., Coulson A. R., Barrell B. G., Smith A. J., Roe B. A. Cloning in single-stranded bacteriophage as an aid to rapid DNA sequencing. J Mol Biol. 1980 Oct 25;143(2):161–178. doi: 10.1016/0022-2836(80)90196-5. [DOI] [PubMed] [Google Scholar]
  51. Schwer B., Stunnenberg H. G. Vaccinia virus late transcripts generated in vitro have a poly(A) head. EMBO J. 1988 Apr;7(4):1183–1190. doi: 10.1002/j.1460-2075.1988.tb02929.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Schwer B., Visca P., Vos J. C., Stunnenberg H. G. Discontinuous transcription or RNA processing of vaccinia virus late messengers results in a 5' poly(A) leader. Cell. 1987 Jul 17;50(2):163–169. doi: 10.1016/0092-8674(87)90212-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Schümperli D., Menna A., Schwendimann F., Wittek R., Wyler R. Symmetrical arrangement of the heterologous regions of rabbit poxvirus and vaccinia virus DNA. J Gen Virol. 1980 Apr;47(2):385–398. doi: 10.1099/0022-1317-47-2-385. [DOI] [PubMed] [Google Scholar]
  54. Shenk T. E., Rhodes C., Rigby P. W., Berg P. Biochemical method for mapping mutational alterations in DNA with S1 nuclease: the location of deletions and temperature-sensitive mutations in simian virus 40. Proc Natl Acad Sci U S A. 1975 Mar;72(3):989–993. doi: 10.1073/pnas.72.3.989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Shuman S., Moss B. Factor-dependent transcription termination by vaccinia virus RNA polymerase. Evidence that the cis-acting termination signal is in nascent RNA. J Biol Chem. 1988 May 5;263(13):6220–6225. [PubMed] [Google Scholar]
  56. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  57. Spencer E., Shuman S., Hurwitz J. Purification and properties of vaccinia virus DNA-dependent RNA polymerase. J Biol Chem. 1980 Jun 10;255(11):5388–5395. [PubMed] [Google Scholar]
  58. Struhl K. Promoters, activator proteins, and the mechanism of transcriptional initiation in yeast. Cell. 1987 May 8;49(3):295–297. doi: 10.1016/0092-8674(87)90277-7. [DOI] [PubMed] [Google Scholar]
  59. Sweetser D., Nonet M., Young R. A. Prokaryotic and eukaryotic RNA polymerases have homologous core subunits. Proc Natl Acad Sci U S A. 1987 Mar;84(5):1192–1196. doi: 10.1073/pnas.84.5.1192. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. 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]
  61. Thompson C. L., Hooda-Dhingra U., Condit R. C. Fine structure mapping of five temperature-sensitive mutants in the 22- and 147-kilodalton subunits of vaccinia virus DNA-dependent RNA polymerase. J Virol. 1989 Feb;63(2):705–713. doi: 10.1128/jvi.63.2.705-713.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. Vos J. C., Stunnenberg H. G. Derepression of a novel class of vaccinia virus genes upon DNA replication. EMBO J. 1988 Nov;7(11):3487–3492. doi: 10.1002/j.1460-2075.1988.tb03224.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  63. Wei C. M., Moss B. Methylated nucleotides block 5'-terminus of vaccinia virus messenger RNA. Proc Natl Acad Sci U S A. 1975 Jan;72(1):318–322. doi: 10.1073/pnas.72.1.318. [DOI] [PMC free article] [PubMed] [Google Scholar]
  64. Weinrich S. L., Hruby D. E. A tandemly-oriented late gene cluster within the vaccinia virus genome. Nucleic Acids Res. 1986 Apr 11;14(7):3003–3016. doi: 10.1093/nar/14.7.3003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  65. Weir J. P., Moss B. Determination of the transcriptional regulatory region of a vaccinia virus late gene. J Virol. 1987 Jan;61(1):75–80. doi: 10.1128/jvi.61.1.75-80.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  66. Weir J. P., Moss B. Regulation of expression and nucleotide sequence of a late vaccinia virus gene. J Virol. 1984 Sep;51(3):662–669. doi: 10.1128/jvi.51.3.662-669.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  67. Wright C. F., Moss B. In vitro synthesis of vaccinia virus late mRNA containing a 5' poly(A) leader sequence. Proc Natl Acad Sci U S A. 1987 Dec;84(24):8883–8887. doi: 10.1073/pnas.84.24.8883. [DOI] [PMC free article] [PubMed] [Google Scholar]
  68. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]
  69. Yuen L., Moss B. Oligonucleotide sequence signaling transcriptional termination of vaccinia virus early genes. Proc Natl Acad Sci U S A. 1987 Sep;84(18):6417–6421. doi: 10.1073/pnas.84.18.6417. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES