Skip to main content
NCBI home page
Journal of Virology logoLink to Journal of Virology
. 1993 Feb;67(2):1024–1033. doi: 10.1128/jvi.67.2.1024-1033.1993

Analysis of the thymidine kinase genes from acyclovir-resistant mutants of varicella-zoster virus isolated from patients with AIDS.

C L Talarico 1, W C Phelps 1, K K Biron 1
PMCID: PMC237457  PMID: 8380452

Abstract

Patients with AIDS often experience recurrent infections with varicella-zoster virus (VZV) requiring repeated or prolonged treatment with acyclovir (ACV), which may lead to the development of ACV resistance. The ACV resistance of isolates recovered from such patients is associated with diminished VZV thymidine kinase (TK) function. We determined the nucleotide sequences of the TK genes of 12 ACV-resistant VZV strains purified from nine patients with AIDS. Five VZV strains contained nucleotide deletions in their TK genes, introducing a premature termination codon which is expected to result in the production of a truncated protein. No detectable full-length TK protein could be immunoprecipitated from extracts of cells infected with these virus strains. These TK-deficient strains were cross resistant to the TK-dependent antiviral agents ACV, 9-(4-hydroxy-3-hydroxymethylbutyl-yl)guanine (penciclovir), and 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl) uracil (BVaraU). The remaining seven strains each contained a nucleotide change that resulted in an amino acid substitution in the TK protein. These substitutions occurred throughout the TK protein, namely, in the ATP-binding site, the nucleoside-binding site, between the two binding sites, and at the carboxy terminus of the protein. We determined the effects of these mutations on the stability of TK protein expression in virus-infected cells and on the sensitivity of mutants to the TK-dependent antiviral agents ACV, BVaraU, and penciclovir.

Full text

PDF
1024

Images in this article

1026Image
on p.1026
1028Image
on p.1028

Selected References

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

  1. Biron K. K., Fyfe J. A., Noblin J. E., Elion G. B. Selection and preliminary characterization of acyclovir-resistant mutants of varicella zoster virus. Am J Med. 1982 Jul 20;73(1A):383–386. doi: 10.1016/0002-9343(82)90128-0. [DOI] [PubMed] [Google Scholar]
  2. Boyd M. R., Bacon T. H., Sutton D. Antiherpesvirus activity of 9-(4-hydroxy-3-hydroxymethylbut-1-yl) guanine (BRL 39123) in animals. Antimicrob Agents Chemother. 1988 Mar;32(3):358–363. doi: 10.1128/aac.32.3.358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Boyd M. R., Bacon T. H., Sutton D., Cole M. Antiherpesvirus activity of 9-(4-hydroxy-3-hydroxy-methylbut-1-yl)guanine (BRL 39123) in cell culture. Antimicrob Agents Chemother. 1987 Aug;31(8):1238–1242. doi: 10.1128/aac.31.8.1238. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chatis P. A., Crumpacker C. S. Analysis of the thymidine kinase gene from clinically isolated acyclovir-resistant herpes simplex viruses. Virology. 1991 Feb;180(2):793–797. doi: 10.1016/0042-6822(91)90093-q. [DOI] [PubMed] [Google Scholar]
  5. Chatis P. A., Miller C. H., Schrager L. E., Crumpacker C. S. Successful treatment with foscarnet of an acyclovir-resistant mucocutaneous infection with herpes simplex virus in a patient with acquired immunodeficiency syndrome. N Engl J Med. 1989 Feb 2;320(5):297–300. doi: 10.1056/NEJM198902023200507. [DOI] [PubMed] [Google Scholar]
  6. Coen D. M., Schaffer P. A. Two distinct loci confer resistance to acycloguanosine in herpes simplex virus type 1. Proc Natl Acad Sci U S A. 1980 Apr;77(4):2265–2269. doi: 10.1073/pnas.77.4.2265. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Collins P., Larder B. A., Oliver N. M., Kemp S., Smith I. W., Darby G. Characterization of a DNA polymerase mutant of herpes simplex virus from a severely immunocompromised patient receiving acyclovir. J Gen Virol. 1989 Feb;70(Pt 2):375–382. doi: 10.1099/0022-1317-70-2-375. [DOI] [PubMed] [Google Scholar]
  8. Dankner W. M., Scholl D., Stanat S. C., Martin M., Sonke R. L., Spector S. A. Rapid antiviral DNA-DNA hybridization assay for human cytomegalovirus. J Virol Methods. 1990 Jun;28(3):293–298. doi: 10.1016/0166-0934(90)90122-v. [DOI] [PubMed] [Google Scholar]
  9. Darby G., Churcher M. J., Larder B. A. Cooperative effects between two acyclovir resistance loci in herpes simplex virus. J Virol. 1984 Jun;50(3):838–846. doi: 10.1128/jvi.50.3.838-846.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Davison A. J., Scott J. E. The complete DNA sequence of varicella-zoster virus. J Gen Virol. 1986 Sep;67(Pt 9):1759–1816. doi: 10.1099/0022-1317-67-9-1759. [DOI] [PubMed] [Google Scholar]
  11. Derse D., Cheng Y. C., Furman P. A., St Clair M. H., Elion G. B. Inhibition of purified human and herpes simplex virus-induced DNA polymerases by 9-(2-hydroxyethoxymethyl)guanine triphosphate. Effects on primer-template function. J Biol Chem. 1981 Nov 25;256(22):11447–11451. [PubMed] [Google Scholar]
  12. Efstathiou S., Kemp S., Darby G., Minson A. C. The role of herpes simplex virus type 1 thymidine kinase in pathogenesis. J Gen Virol. 1989 Apr;70(Pt 4):869–879. doi: 10.1099/0022-1317-70-4-869. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Field H. J., Coen D. M. Pathogenicity of herpes simplex virus mutants containing drug resistance mutations in the viral DNA polymerase gene. J Virol. 1986 Oct;60(1):286–289. doi: 10.1128/jvi.60.1.286-289.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Field H., McMillan A., Darby G. The sensitivity of acyclovir-resistant mutants of herpes simplex virus to other antiviral drugs. J Infect Dis. 1981 Feb;143(2):281–285. doi: 10.1093/infdis/143.2.281. [DOI] [PubMed] [Google Scholar]
  16. Furman P. A., St Clair M. H., Fyfe J. A., Rideout J. L., Keller P. M., Elion G. B. Inhibition of herpes simplex virus-induced DNA polymerase activity and viral DNA replication by 9-(2-hydroxyethoxymethyl)guanine and its triphosphate. J Virol. 1979 Oct;32(1):72–77. doi: 10.1128/jvi.32.1.72-77.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Furman P. A., St Clair M. H., Spector T. Acyclovir triphosphate is a suicide inactivator of the herpes simplex virus DNA polymerase. J Biol Chem. 1984 Aug 10;259(15):9575–9579. [PubMed] [Google Scholar]
  18. Fyfe J. A. Differential phosphorylation of (E)-5-(2-bromovinyl)-2'-deoxyuridine monophosphate by thymidylate kinases from herpes simplex viruses types 1 and 2 and varicella zoster virus. Mol Pharmacol. 1982 Mar;21(2):432–437. [PubMed] [Google Scholar]
  19. Fyfe J. A., McKee S. A., Keller P. M. Altered thymidine-thymidylate kinases from strains of herpes simplex virus with modified drug sensitivities to acyclovir and (E)-5-(2-bromovinyl)-2'-deoxyuridine. Mol Pharmacol. 1983 Sep;24(2):316–323. [PubMed] [Google Scholar]
  20. Graham D., Larder B. A., Inglis M. M. Evidence that the 'active centre' of the herpes simplex virus thymidine kinase involves an interaction between three distinct regions of the polypeptide. J Gen Virol. 1986 Apr;67(Pt 4):753–758. doi: 10.1099/0022-1317-67-4-753. [DOI] [PubMed] [Google Scholar]
  21. Hannink M., Donoghue D. J. Lysine residue 121 in the proposed ATP-binding site of the v-mos protein is required for transformation. Proc Natl Acad Sci U S A. 1985 Dec;82(23):7894–7898. doi: 10.1073/pnas.82.23.7894. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hill E. L., Hunter G. A., Ellis M. N. In vitro and in vivo characterization of herpes simplex virus clinical isolates recovered from patients infected with human immunodeficiency virus. Antimicrob Agents Chemother. 1991 Nov;35(11):2322–2328. doi: 10.1128/aac.35.11.2322. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jacobson M. A., Berger T. G., Fikrig S., Becherer P., Moohr J. W., Stanat S. C., Biron K. K. Acyclovir-resistant varicella zoster virus infection after chronic oral acyclovir therapy in patients with the acquired immunodeficiency syndrome (AIDS). Ann Intern Med. 1990 Feb 1;112(3):187–191. doi: 10.7326/0003-4819-112-3-187. [DOI] [PubMed] [Google Scholar]
  24. Kamps M. P., Sefton B. M. Neither arginine nor histidine can carry out the function of lysine-295 in the ATP-binding site of p60src. Mol Cell Biol. 1986 Mar;6(3):751–757. doi: 10.1128/mcb.6.3.751. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kit S., Sheppard M., Ichimura H., Nusinoff-Lehrman S., Ellis M. N., Fyfe J. A., Otsuka H. Nucleotide sequence changes in thymidine kinase gene of herpes simplex virus type 2 clones from an isolate of a patient treated with acyclovir. Antimicrob Agents Chemother. 1987 Oct;31(10):1483–1490. doi: 10.1128/aac.31.10.1483. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kit S. Thymidine kinase. Microbiol Sci. 1985 Dec;2(12):369–375. [PubMed] [Google Scholar]
  27. Lacey S. F., Suzutani T., Powell K. L., Purifoy D. J., Honess R. W. Analysis of mutations in the thymidine kinase genes of drug-resistant varicella-zoster virus populations using the polymerase chain reaction. J Gen Virol. 1991 Mar;72(Pt 3):623–630. doi: 10.1099/0022-1317-72-3-623. [DOI] [PubMed] [Google Scholar]
  28. Larder B. A., Cheng Y. C., Darby G. Characterization of abnormal thymidine kinases induced by drug-resistant strains of herpes simplex virus type 1. J Gen Virol. 1983 Mar;64(Pt 3):523–532. doi: 10.1099/0022-1317-64-3-523. [DOI] [PubMed] [Google Scholar]
  29. Larder B. A., Derse D., Cheng Y. C., Darby G. Properties of purified enzymes induced by pathogenic drug-resistant mutants of herpes simplex virus. Evidence for virus variants expressing normal DNA polymerase and altered thymidine kinase. J Biol Chem. 1983 Feb 10;258(3):2027–2033. [PubMed] [Google Scholar]
  30. Larder B. A., Kemp S. D., Darby G. Related functional domains in virus DNA polymerases. EMBO J. 1987 Jan;6(1):169–175. doi: 10.1002/j.1460-2075.1987.tb04735.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Liu Q. Y., Summers W. C. Site-directed mutagenesis of a nucleotide-binding domain in HSV-1 thymidine kinase: effects on catalytic activity. Virology. 1988 Apr;163(2):638–642. doi: 10.1016/0042-6822(88)90308-x. [DOI] [PubMed] [Google Scholar]
  32. Lopetegui P., Matsunaga Y., Okuno T., Ogino T., Yamanishi K. Expression of varicella-zoster virus-related antigens in biochemically transformed cells. J Gen Virol. 1983 May;64(Pt 5):1181–1186. doi: 10.1099/0022-1317-64-5-1181. [DOI] [PubMed] [Google Scholar]
  33. Machida H., Sakata S. In vitro and in vivo antiviral activity of 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil (BV-araU) and related compounds. Antiviral Res. 1984 Jun;4(3):135–141. doi: 10.1016/0166-3542(84)90013-5. [DOI] [PubMed] [Google Scholar]
  34. Machida H., Watanabe Y. Inhibition of DNA synthesis in varicella-zoster virus-infected cells by BV-araU. Microbiol Immunol. 1991;35(2):139–145. doi: 10.1111/j.1348-0421.1991.tb01541.x. [DOI] [PubMed] [Google Scholar]
  35. Martin J. L., Ellis M. N., Keller P. M., Biron K. K., Lehrman S. N., Barry D. W., Furman P. A. Plaque autoradiography assay for the detection and quantitation of thymidine kinase-deficient and thymidine kinase-altered mutants of herpes simplex virus in clinical isolates. Antimicrob Agents Chemother. 1985 Aug;28(2):181–187. doi: 10.1128/aac.28.2.181. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Mitchell C. D., Bean B., Gentry S. R., Groth K. E., Boen J. R., Balfour H. H., Jr Acyclovir therapy for mucocutaneous herpes simplex infections in immunocompromised patients. Lancet. 1981 Jun 27;1(8235):1389–1392. doi: 10.1016/s0140-6736(81)92569-1. [DOI] [PubMed] [Google Scholar]
  37. Mori H., Shiraki K., Kato T., Hayakawa Y., Yamanishi K., Takahashi M. Molecular analysis of the thymidine kinase gene of thymidine kinase-deficient mutants of varicella-zoster virus. Intervirology. 1988;29(6):301–310. doi: 10.1159/000150060. [DOI] [PubMed] [Google Scholar]
  38. Pahwa S., Biron K., Lim W., Swenson P., Kaplan M. H., Sadick N., Pahwa R. Continuous varicella-zoster infection associated with acyclovir resistance in a child with AIDS. JAMA. 1988 Nov 18;260(19):2879–2882. [PubMed] [Google Scholar]
  39. Reardon J. E., Spector T. Herpes simplex virus type 1 DNA polymerase. Mechanism of inhibition by acyclovir triphosphate. J Biol Chem. 1989 May 5;264(13):7405–7411. [PubMed] [Google Scholar]
  40. Roberts G. B., Fyfe J. A., Gaillard R. K., Short S. A. Mutant varicella-zoster virus thymidine kinase: correlation of clinical resistance and enzyme impairment. J Virol. 1991 Dec;65(12):6407–6413. doi: 10.1128/jvi.65.12.6407-6413.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Robertson G. R., Whalley J. M. Evolution of the herpes thymidine kinase: identification and comparison of the equine herpesvirus 1 thymidine kinase gene reveals similarity to a cell-encoded thymidylate kinase. Nucleic Acids Res. 1988 Dec 9;16(23):11303–11317. doi: 10.1093/nar/16.23.11303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Ruth J. L., Cheng Y. C. Nucleoside analogues with clinical potential in antivirus chemotherapy. The effect of several thymidine and 2'-deoxycytidine analogue 5'-triphosphates on purified human (alpha, beta) and herpes simplex virus (types 1, 2) DNA polymerases. Mol Pharmacol. 1981 Sep;20(2):415–422. [PubMed] [Google Scholar]
  43. Sacks S. L., Wanklin R. J., Reece D. E., Hicks K. A., Tyler K. L., Coen D. M. Progressive esophagitis from acyclovir-resistant herpes simplex. Clinical roles for DNA polymerase mutants and viral heterogeneity? Ann Intern Med. 1989 Dec 1;111(11):893–899. doi: 10.7326/0003-4819-111-11-893. [DOI] [PubMed] [Google Scholar]
  44. Safrin S., Berger T. G., Gilson I., Wolfe P. R., Wofsy C. B., Mills J., Biron K. K. Foscarnet therapy in five patients with AIDS and acyclovir-resistant varicella-zoster virus infection. Ann Intern Med. 1991 Jul 1;115(1):19–21. doi: 10.7326/0003-4819-115-1-19. [DOI] [PubMed] [Google Scholar]
  45. Safrin S., Crumpacker C., Chatis P., Davis R., Hafner R., Rush J., Kessler H. A., Landry B., Mills J. A controlled trial comparing foscarnet with vidarabine for acyclovir-resistant mucocutaneous herpes simplex in the acquired immunodeficiency syndrome. The AIDS Clinical Trials Group. N Engl J Med. 1991 Aug 22;325(8):551–555. doi: 10.1056/NEJM199108223250805. [DOI] [PubMed] [Google Scholar]
  46. Sawyer M. H., Inchauspe G., Biron K. K., Waters D. J., Straus S. E., Ostrove J. M. Molecular analysis of the pyrimidine deoxyribonucleoside kinase gene of wild-type and acyclovir-resistant strains of varicella-zoster virus. J Gen Virol. 1988 Oct;69(Pt 10):2585–2593. doi: 10.1099/0022-1317-69-10-2585. [DOI] [PubMed] [Google Scholar]
  47. Schnipper L. E., Crumpacker C. S. Resistance of herpes simplex virus to acycloguanosine: role of viral thymidine kinase and DNA polymerase loci. Proc Natl Acad Sci U S A. 1980 Apr;77(4):2270–2273. doi: 10.1073/pnas.77.4.2270. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Shiraki K., Ogino T., Yamanishi K., Takahashi M. Immunochemical characterization of pyrimidine kinase induced by varicella-zoster virus. J Gen Virol. 1985 Feb;66(Pt 2):221–229. doi: 10.1099/0022-1317-66-2-221. [DOI] [PubMed] [Google Scholar]
  49. Straus S. E., Smith H. A., Brickman C., de Miranda P., McLaren C., Keeney R. E. Acyclovir for chronic mucocutaneous herpes simplex virus infection in immunosuppressed patients. Ann Intern Med. 1982 Mar;96(3):270–277. doi: 10.7326/0003-4819-96-3-270. [DOI] [PubMed] [Google Scholar]
  50. Suzutani T., Machida H., Sakuma T., Azuma M. Effects of various nucleosides on antiviral activity and metabolism of 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil against herpes simplex virus types 1 and 2. Antimicrob Agents Chemother. 1988 Oct;32(10):1547–1551. doi: 10.1128/aac.32.10.1547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Wade J. C., Newton B., McLaren C., Flournoy N., Keeney R. E., Meyers J. D. Intravenous acyclovir to treat mucocutaneous herpes simplex virus infection after marrow transplantation: a double-blind trial. Ann Intern Med. 1982 Mar;96(3):265–269. doi: 10.7326/0003-4819-96-3-265. [DOI] [PubMed] [Google Scholar]
  52. Weinmaster G., Zoller M. J., Pawson T. A lysine in the ATP-binding site of P130gag-fps is essential for protein-tyrosine kinase activity. EMBO J. 1986 Jan;5(1):69–76. doi: 10.1002/j.1460-2075.1986.tb04179.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES