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Journal of Virology logoLink to Journal of Virology
. 1987 Jan;61(1):119–124. doi: 10.1128/jvi.61.1.119-124.1987

Genomic localization, sequence analysis, and transcription of the putative human cytomegalovirus DNA polymerase gene.

R Heilbronn, G Jahn, A Bürkle, U K Freese, B Fleckenstein, H zur Hausen
PMCID: PMC255217  PMID: 3023689

Abstract

The human cytomegalovirus (HCMV)-induced DNA polymerase has been well characterized biochemically and functionally, but its genomic location has not yet been assigned. To identify the coding sequence, cross-hybridization with the herpes simplex virus type 1 (HSV-1) polymerase gene was used, as suggested by the close similarity of the herpes group virus-induced DNA polymerases to the HCMV DNA polymerase. A cosmid and plasmid library of the entire HCMV genome was screened with the BamHI Q fragment of HSV-1 at different stringency conditions. One PstI-HincII restriction fragment of 850 base pairs mapping within the EcoRI M fragment of HCMV cross-hybridized at Tm - 25 degrees C. Sequence analysis revealed one open reading frame spanning the entire sequence. The amino acid sequence showed a highly conserved domain of 133 amino acids shared with the HSV and putative Epstein-Barr virus polymerase sequences. This domain maps within the C-terminal part of the HSV polymerase gene, which has been suggested to contain part of the catalytic center of the enzyme. Transcription analysis revealed one 5.4-kilobase early transcript in the sense orientation with respect to the open reading frame identified. This transcript appears to code for the 140-kilodalton HCMV polymerase protein.

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Selected References

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  1. Baer R., Bankier A. T., Biggin M. D., Deininger P. L., Farrell P. J., Gibson T. J., Hatfull G., Hudson G. S., Satchwell S. C., Séguin C. DNA sequence and expression of the B95-8 Epstein-Barr virus genome. Nature. 1984 Jul 19;310(5974):207–211. doi: 10.1038/310207a0. [DOI] [PubMed] [Google Scholar]
  2. Beltz G. A., Jacobs K. A., Eickbush T. H., Cherbas P. T., Kafatos F. C. Isolation of multigene families and determination of homologies by filter hybridization methods. Methods Enzymol. 1983;100:266–285. doi: 10.1016/0076-6879(83)00061-0. [DOI] [PubMed] [Google Scholar]
  3. Chartrand P., Crumpacker C. S., Schaffer P. A., Wilkie N. M. Physical and genetic analysis of the herpes simplex virus DNA polymerase locus. Virology. 1980 Jun;103(2):311–326. doi: 10.1016/0042-6822(80)90190-7. [DOI] [PubMed] [Google Scholar]
  4. Chartrand P., Stow N. D., Timbury M. C., Wilkie N. M. Physical mapping of paar mutations of herpes simplex virus type 1 and type 2 by intertypic marker rescue. J Virol. 1979 Aug;31(2):265–276. doi: 10.1128/jvi.31.2.265-276.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Crouse G. F., Frischauf A., Lehrach H. An integrated and simplified approach to cloning into plasmids and single-stranded phages. Methods Enzymol. 1983;101:78–89. doi: 10.1016/0076-6879(83)01006-x. [DOI] [PubMed] [Google Scholar]
  6. Crumpacker C. S., Schnipper L. E., Zaia J. A., Levin M. J. Growth inhibition by acycloguanosine of herpesviruses isolated from human infections. Antimicrob Agents Chemother. 1979 May;15(5):642–645. doi: 10.1128/aac.15.5.642. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Datta A. K., Feighny R. J., Pagano J. S. Induction of Epstein-Barr virus-associated DNA polymerase by 12-O-tetradecanoylphorbol-13-acetate. Purification and characterization. J Biol Chem. 1980 Jun 10;255(11):5120–5125. [PubMed] [Google Scholar]
  8. Elion G. B., Furman P. A., Fyfe J. A., de Miranda P., Beauchamp L., Schaeffer H. J. Selectivity of action of an antiherpetic agent, 9-(2-hydroxyethoxymethyl) guanine. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5716–5720. doi: 10.1073/pnas.74.12.5716. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Estes J. E., Huang E. S. Stimulation of cellular thymidine kinases by human cytomegalovirus. J Virol. 1977 Oct;24(1):13–21. doi: 10.1128/jvi.24.1.13-21.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fleckenstein B., Müller I., Collins J. Cloning of the complete human cytomegalovirus genome in cosmids. Gene. 1982 Apr;18(1):39–46. doi: 10.1016/0378-1119(82)90054-3. [DOI] [PubMed] [Google Scholar]
  11. Gibbs J. S., Chiou H. C., Hall J. D., Mount D. W., Retondo M. J., Weller S. K., Coen D. M. Sequence and mapping analyses of the herpes simplex virus DNA polymerase gene predict a C-terminal substrate binding domain. Proc Natl Acad Sci U S A. 1985 Dec;82(23):7969–7973. doi: 10.1073/pnas.82.23.7969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Greenaway P. J., Oram J. D., Downing R. G., Patel K. Human cytomegalovirus DNA: BamHI, EcoRI and PstI restriction endonuclease cleavage maps. Gene. 1982 Jun;18(3):355–360. doi: 10.1016/0378-1119(82)90174-3. [DOI] [PubMed] [Google Scholar]
  13. Holland L. E., Sandri-Goldin R. M., Goldin A. L., Glorioso J. C., Levine M. Transcriptional and genetic analyses of the herpes simplex virus type 1 genome: coordinates 0.29 to 0.45. J Virol. 1984 Mar;49(3):947–959. doi: 10.1128/jvi.49.3.947-959.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Huang E. S. Human cytomegalovirus. III. Virus-induced DNA polymerase. J Virol. 1975 Aug;16(2):298–310. doi: 10.1128/jvi.16.2.298-310.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Huang E. S. Human cytomegalovirus. IV. Specific inhibition of virus-induced DNA polymerase activity and viral DNA replication by phosphonoacetic acid. J Virol. 1975 Dec;16(6):1560–1565. doi: 10.1128/jvi.16.6.1560-1565.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jahn G., Knust E., Schmolla H., Sarre T., Nelson J. A., McDougall J. K., Fleckenstein B. Predominant immediate-early transcripts of human cytomegalovirus AD 169. J Virol. 1984 Feb;49(2):363–370. doi: 10.1128/jvi.49.2.363-370.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Jordan M. C. Latent infection and the elusive cytomegalovirus. Rev Infect Dis. 1983 Mar-Apr;5(2):205–215. doi: 10.1093/clinids/5.2.205. [DOI] [PubMed] [Google Scholar]
  18. Kallin B., Sternås L., Saemundssen A. K., Luka J., Jörnvall H., Eriksson B., Tao P. Z., Nilsson M. T., Klein G. Purification of Epstein-Barr virus DNA polymerase from P3HR-1 cells. J Virol. 1985 May;54(2):561–568. doi: 10.1128/jvi.54.2.561-568.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Knopf K. W., Kaufman E. R., Crumpacker C. Physical mapping of drug resistance mutations defines an active center of the herpes simplex virus DNA polymerase enzyme. J Virol. 1981 Sep;39(3):746–757. doi: 10.1128/jvi.39.3.746-757.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Knopf K. W. Properties of herpes simplex virus DNA polymerase and characterization of its associated exonuclease activity. Eur J Biochem. 1979 Jul;98(1):231–244. doi: 10.1111/j.1432-1033.1979.tb13181.x. [DOI] [PubMed] [Google Scholar]
  21. Kouzarides T., Bankier A. T., Satchwell S. C., Weston K., Tomlinson P., Barrell B. G. Sequence and transcription analysis of the human cytomegalovirus DNA polymerase gene. J Virol. 1987 Jan;61(1):125–133. doi: 10.1128/jvi.61.1.125-133.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mar E. C., Chiou J. F., Cheng Y. C., Huang E. S. Human cytomegalovirus-induced DNA polymerase and its interaction with the triphosphates of 1-(2'-deoxy-2'-fluoro-beta-D-arabinofuranosyl)-5-methyluracil, -5-iodocytosine, and -5-methylcytosine. J Virol. 1985 Dec;56(3):846–851. doi: 10.1128/jvi.56.3.846-851.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mar E. C., Chiou J. F., Cheng Y. C., Huang E. S. Inhibition of cellular DNA polymerase alpha and human cytomegalovirus-induced DNA polymerase by the triphosphates of 9-(2-hydroxyethoxymethyl)guanine and 9-(1,3-dihydroxy-2-propoxymethyl)guanine. J Virol. 1985 Mar;53(3):776–780. doi: 10.1128/jvi.53.3.776-780.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Mar E. C., Patel P. C., Cheng Y. C., Fox J. J., Watanabe K. A., Huang E. S. Effects of certain nucleoside analogues on human cytomegalovirus replication in vitro. J Gen Virol. 1984 Jan;65(Pt 1):47–53. doi: 10.1099/0022-1317-65-1-47. [DOI] [PubMed] [Google Scholar]
  25. Melton D. A., Krieg P. A., Rebagliati M. R., Maniatis T., Zinn K., Green M. R. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res. 1984 Sep 25;12(18):7035–7056. doi: 10.1093/nar/12.18.7035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Messing J., Vieira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982 Oct;19(3):269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
  27. Nishiyama Y., Maeno K., Yoshida S. Characterization of human cytomegalovirus-induced DNA polymerase and the associated 3'-to-5', exonuclease. Virology. 1983 Jan 30;124(2):221–231. doi: 10.1016/0042-6822(83)90339-2. [DOI] [PubMed] [Google Scholar]
  28. Polack A., Hartl G., Zimber U., Freese U. K., Laux G., Takaki K., Hohn B., Gissmann L., Bornkamm G. W. A complete set of overlapping cosmid clones of M-ABA virus derived from nasopharyngeal carcinoma and its similarity to other Epstein-Barr virus isolates. Gene. 1984 Mar;27(3):279–288. doi: 10.1016/0378-1119(84)90072-6. [DOI] [PubMed] [Google Scholar]
  29. Quinn J. P., McGeoch D. J. DNA sequence of the region in the genome of herpes simplex virus type 1 containing the genes for DNA polymerase and the major DNA binding protein. Nucleic Acids Res. 1985 Nov 25;13(22):8143–8163. doi: 10.1093/nar/13.22.8143. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Richardson W. P., Colvin R. B., Cheeseman S. H., Tolkoff-Rubin N. E., Herrin J. T., Cosimi A. B., Collins A. B., Hirsch M. S., McCluskey R. T., Russell P. S. Glomerulopathy associated with cytomegalovirus viremia in renal allografts. N Engl J Med. 1981 Jul 9;305(2):57–63. doi: 10.1056/NEJM198107093050201. [DOI] [PubMed] [Google Scholar]
  31. 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]
  32. 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]
  33. Schrier R. D., Nelson J. A., Oldstone M. B. Detection of human cytomegalovirus in peripheral blood lymphocytes in a natural infection. Science. 1985 Nov 29;230(4729):1048–1051. doi: 10.1126/science.2997930. [DOI] [PubMed] [Google Scholar]
  34. St Clair M. H., Furman P. A., Lubbers C. M., Elion G. B. Inhibition of cellular alpha and virally induced deoxyribonucleic acid polymerases by the triphosphate of acyclovir. Antimicrob Agents Chemother. 1980 Nov;18(5):741–745. doi: 10.1128/aac.18.5.741. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Stagno S., Pass R. F., Dworsky M. E., Henderson R. E., Moore E. G., Walton P. D., Alford C. A. Congenital cytomegalovirus infection: The relative importance of primary and recurrent maternal infection. N Engl J Med. 1982 Apr 22;306(16):945–949. doi: 10.1056/NEJM198204223061601. [DOI] [PubMed] [Google Scholar]
  36. Vaughan P. J., Purifoy D. J., Powell K. L. DNA-binding protein associated with herpes simplex virus DNA polymerase. J Virol. 1985 Feb;53(2):501–508. doi: 10.1128/jvi.53.2.501-508.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Werner D., Chemla Y., Herzberg M. Isolation of poly(A)+ RNA by paper affinity chromatography. Anal Biochem. 1984 Sep;141(2):329–336. doi: 10.1016/0003-2697(84)90050-2. [DOI] [PubMed] [Google Scholar]
  38. Závada V., Erban V., Rezácová D., Vonka V. Thymidine-kinase in cytomegalovirus infected cells. Arch Virol. 1976;52(4):333–339. doi: 10.1007/BF01315622. [DOI] [PubMed] [Google Scholar]

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