1887

Journal of General Virology header logo

Volume 95, Issue 4

Roles of conserved residues within the pre-NH-terminal domain of herpes simplex virus 1 DNA polymerase in replication and latency in mice Free

Abstract

The catalytic subunit of the herpes simplex virus 1 DNA polymerase (HSV-1 Pol) is essential for viral DNA synthesis and production of infectious virus in cell culture. While mutations that affect 5′–3′ polymerase activity have been evaluated in animal models of HSV-1 infection, mutations that affect other functions of HSV-1 Pol have not. In a previous report, we utilized bacterial artificial chromosome technology to generate defined HSV-1 mutants with lesions in the previously uncharacterized pre-NH-terminal domain. We found that the extreme N-terminal 42 residues (deletion mutant ΔN43) were dispensable for replication in cell culture, while residues 44–49 (alanine-substitution mutant A) were required for efficient viral DNA synthesis and production of infectious virus. In this study, we sought to address the importance of these conserved elements in viral replication in a mouse corneal infection model. Mutant virus ΔN43 exhibited no meaningful defect in acute or latent infection despite strong conservation of residues 1–42 with HSV-2 Pol. The A mutation caused a modest defect in replication at the site of inoculation, and was severely impaired for ganglionic replication, even at high inocula that permitted efficient corneal replication. Additionally, the A mutation resulted in reduced latency establishment and subsequent reactivation. Moreover, we found that the A replication defect in cultured cells was exacerbated in resting cells as compared to dividing cells. These results reveal an important role for the conserved motif at residues 44–49 of HSV-1 Pol for ganglionic viral replication.

  • Received:
  • Accepted:
  • Published Online:
© 2014 SGM
Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.061903-0
2014-04-01
2024-04-16
Loading full text...

Full text loading...

/deliver/fulltext/jgv/95/4/940.html?itemId=/content/journal/jgv/10.1099/vir.0.061903-0&mimeType=html&fmt=ahah

References

  1. Aron G. M., Purifoy D. J., Schaffer P. A. 1975; DNA synthesis and DNA polymerase activity of herpes simplex virus type 1 temperature-sensitive mutants. J Virol 16:498–507[PubMed]
     [Google Scholar]
  2. Balliet J. W., Kushnir A. S., Schaffer P. A. 2007; Construction and characterization of a herpes simplex virus type I recombinant expressing green fluorescent protein: acute phase replication and reactivation in mice. Virology 361:372–383 [View Article][PubMed]
     [Google Scholar]
  3. Bernad A., Blanco L., Lázaro J. M., Martín G., Salas M. 1989; A conserved 3′→5′ exonuclease active site in prokaryotic and eukaryotic DNA polymerases. Cell 59:219–228 [View Article][PubMed]
     [Google Scholar]
  4. Bolovan C. A., Sawtell N. M., Thompson R. L. 1994; ICP34.5 mutants of herpes simplex virus type 1 strain 17syn+ are attenuated for neurovirulence in mice and for replication in confluent primary mouse embryo cell cultures. J Virol 68:48–55[PubMed]
     [Google Scholar]
  5. Brown S. M., Harland J., MacLean A. R., Podlech J., Clements J. B. 1994; Cell type and cell state determine differential in vitro growth of non-neurovirulent ICP34.5-negative herpes simplex virus types 1 and 2. J Gen Virol 75:2367–2377 [View Article][PubMed]
     [Google Scholar]
  6. Cai W., Astor T. L., Liptak L. M., Cho C., Coen D. M., Schaffer P. A. 1993; The herpes simplex virus type 1 regulatory protein ICP0 enhances virus replication during acute infection and reactivation from latency. J Virol 67:7501–7512[PubMed]
     [Google Scholar]
  7. Chen S. H., Pearson A., Coen D. M., Chen S. H. 2004; Failure of thymidine kinase-negative herpes simplex virus to reactivate from latency following efficient establishment. J Virol 78:520–523 [View Article][PubMed]
     [Google Scholar]
  8. Coen D. M., Kosz-Vnenchak M., Jacobson J. G., Leib D. A., Bogard C. L., Schaffer P. A., Tyler K. L., Knipe D. M. 1989; Thymidine kinase-negative herpes simplex virus mutants establish latency in mouse trigeminal ganglia but do not reactivate. Proc Natl Acad Sci U S A 86:4736–4740 [View Article][PubMed]
     [Google Scholar]
  9. Darby G., Churcher M. J., Larder B. A. 1984; Cooperative effects between two acyclovir resistance loci in herpes simplex virus. J Virol 50:838–846[PubMed]
     [Google Scholar]
  10. Di Tommaso P., Moretti S., Xenarios I., Orobitg M., Montanyola A., Chang J. M., Taly J. F., Notredame C. 2011; T-Coffee: a web server for the multiple sequence alignment of protein and RNA sequences using structural information and homology extension. Nucleic Acids Res 39:Suppl 2W13–W17 [View Article][PubMed]
     [Google Scholar]
  11. Efstathiou S., Preston C. M. 2005; Towards an understanding of the molecular basis of herpes simplex virus latency. Virus Res 111:108–119 [View Article][PubMed]
     [Google Scholar]
  12. Efstathiou S., Kemp S., Darby G., Minson A. C. 1989; The role of herpes simplex virus type 1 thymidine kinase in pathogenesis. J Gen Virol 70:869–879 [View Article][PubMed]
     [Google Scholar]
  13. Ehmann G. L., McLean T. I., Bachenheimer S. L. 2000; Herpes simplex virus type 1 infection imposes a G(1)/S block in asynchronously growing cells and prevents G(1) entry in quiescent cells. J Virol 267:335–349 [CrossRef]
     [Google Scholar]
  14. Elmore D., Eberle R. 2008; Monkey B virus (Cercopithecine herpesvirus 1). Comp Med 58:11–21[PubMed]
     [Google Scholar]
  15. Field H. J., Coen D. M. 1986; Pathogenicity of herpes simplex virus mutants containing drug resistance mutations in the viral DNA polymerase gene. J Virol 60:286–289[PubMed]
     [Google Scholar]
  16. Field H. J., Wildy P. 1978; The pathogenicity of thymidine kinase-deficient mutants of herpes simplex virus in mice. J Hyg (Lond) 81:267–277 [View Article][PubMed]
     [Google Scholar]
  17. Goldstein D. J., Weller S. K. 1988a; Factor(s) present in herpes simplex virus type 1-infected cells can compensate for the loss of the large subunit of the viral ribonucleotide reductase: characterization of an ICP6 deletion mutant. Virology 166:41–51 [View Article][PubMed]
     [Google Scholar]
  18. Goldstein D. J., Weller S. K. 1988b; Herpes simplex virus type 1-induced ribonucleotide reductase activity is dispensable for virus growth and DNA synthesis: isolation and characterization of an ICP6 lacZ insertion mutant. J Virol 62:196–205[PubMed]
     [Google Scholar]
  19. Hwang C. B., Ruffner K. L., Coen D. M. 1992; A point mutation within a distinct conserved region of the herpes simplex virus DNA polymerase gene confers drug resistance. J Virol 66:1774–1776[PubMed]
     [Google Scholar]
  20. Hwang Y. T., Liu B. Y., Coen D. M., Hwang C. B. 1997; Effects of mutations in the Exo III motif of the herpes simplex virus DNA polymerase gene on enzyme activities, viral replication, and replication fidelity. J Virol 71:7791–7798[PubMed]
     [Google Scholar]
  21. Jacobson J. G., Leib D. A., Goldstein D. J., Bogard C. L., Schaffer P. A., Weller S. K., Coen D. M. 1989; A herpes simplex virus ribonucleotide reductase deletion mutant is defective for productive acute and reactivatable latent infections of mice and for replication in mouse cells. Virology 173:276–283 [View Article][PubMed]
     [Google Scholar]
  22. Jamieson A. T., Gentry G. A., Subak-Sharpe J. H. 1974; Induction of both thymidine and deoxycytidine kinase activity by herpes viruses. J Gen Virol 24:465–480 [View Article][PubMed]
     [Google Scholar]
  23. Katz J. P., Bodin E. T., Coen D. M. 1990; Quantitative polymerase chain reaction analysis of herpes simplex virus DNA in ganglia of mice infected with replication-incompetent mutants. J Virol 64:4288–4295[PubMed]
     [Google Scholar]
  24. Kramer M. F., Jurak I., Pesola J. M., Boissel S., Knipe D. M., Coen D. M. 2011; Herpes simplex virus 1 microRNAs expressed abundantly during latent infection are not essential for latency in mouse trigeminal ganglia. Virology 417:239–247 [View Article][PubMed]
     [Google Scholar]
  25. Larder B. A., Darby G. 1985; Selection and characterisation of acyclovir-resistant herpes simplex virus type 1 mutants inducing altered DNA polymerase activities. Virology 146:262–271 [View Article][PubMed]
     [Google Scholar]
  26. Leib D. A., Bogard C. L., Kosz-Vnenchak M., Hicks K. A., Coen D. M., Knipe D. M., Schaffer P. A. 1989; A deletion mutant of the latency-associated transcript of herpes simplex virus type 1 reactivates from the latent state with reduced frequency. J Virol 63:2893–2900[PubMed]
     [Google Scholar]
  27. Leib D. A., Nadeau K. C., Rundle S. A., Schaffer P. A. 1991; The promoter of the latency-associated transcripts of herpes simplex virus type 1 contains a functional cAMP-response element: role of the latency-associated transcripts and cAMP in reactivation of viral latency. Proc Natl Acad Sci U S A 88:48–52 [View Article][PubMed]
     [Google Scholar]
  28. Liu S., Knafels J. D., Chang J. S., Waszak G. A., Baldwin E. T., Deibel M. R. Jr, Thomsen D. R., Homa F. L., Wells P. A. other authors 2006; Crystal structure of the herpes simplex virus 1 DNA polymerase. J Biol Chem 281:18193–18200 [View Article][PubMed]
     [Google Scholar]
  29. Marcy A. I., Olivo P. D., Challberg M. D., Coen D. M. 1990; Enzymatic activities of overexpressed herpes simplex virus DNA polymerase purified from recombinant baculovirus-infected insect cells. Nucleic Acids Res 18:1207–1215 [View Article][PubMed]
     [Google Scholar]
  30. McGeoch D. J., Dolan A., Ralph A. C. 2000; Toward a comprehensive phylogeny for mammalian and avian herpesviruses. J Virol 74:10401–10406 [View Article][PubMed]
     [Google Scholar]
  31. Notredame C., Higgins D. G., Heringa J. 2000; T-Coffee: A novel method for fast and accurate multiple sequence alignment. J Mol Biol 302:205–217 [View Article][PubMed]
     [Google Scholar]
  32. Nutter L. M., Grill S. P., Cheng Y. C. 1985; Can ribonucleotide reductase be considered as an effective target for developing antiherpes simplex virus type II (HSV-2) compounds?. Biochem Pharmacol 34:777–780 [View Article][PubMed]
     [Google Scholar]
  33. Pelosi E., Mulamba G. B., Coen D. M. 1998; Penciclovir and pathogenesis phenotypes of drug-resistant Herpes simplex virus mutants. Antiviral Res 37:17–28 [View Article][PubMed]
     [Google Scholar]
  34. Perng G. C., Ghiasi H., Slanina S. M., Nesburn A. B., Wechsler S. L. 1996; High-dose ocular infection with a herpes simplex virus type 1 ICP34.5 deletion mutant produces no corneal disease or neurovirulence yet results in wild-type levels of spontaneous reactivation. J Virol 70:2883–2893[PubMed]
     [Google Scholar]
  35. Pesola J. M., Zhu J., Knipe D. M., Coen D. M. 2005; Herpes simplex virus 1 immediate-early and early gene expression during reactivation from latency under conditions that prevent infectious virus production. J Virol 79:14516–14525 [View Article][PubMed]
     [Google Scholar]
  36. Preston V. G., Darling A. J., McDougall I. M. 1988; The herpes simplex virus type 1 temperature-sensitive mutant ts1222 has a single base pair deletion in the small subunit of ribonucleotide reductase. Virology 167:458–467[PubMed]
     [Google Scholar]
  37. Purifoy D. J., Lewis R. B., Powell K. L. 1977; Identification of the herpes simplex virus DNA polymerase gene. Nature 269:621–623 [View Article][PubMed]
     [Google Scholar]
  38. Song B., Liu J. J., Yeh K. C., Knipe D. M. 2000; Herpes simplex virus infection blocks events in the G1 phase of the cell cycle. Virology 267:326–334 [CrossRef]
     [Google Scholar]
  39. Tenser R. B., Hay K. A., Edris W. A. 1989; Latency-associated transcript but not reactivatable virus is present in sensory ganglion neurons after inoculation of thymidine kinase-negative mutants of herpes simplex virus type 1. J Virol 63:2861–2865[PubMed]
     [Google Scholar]
  40. Terrell S. L., Coen D. M. 2012; The pre-NH(2)-terminal domain of the herpes simplex virus 1 DNA polymerase catalytic subunit is required for efficient viral replication. J Virol 86:11057–11065 [View Article][PubMed]
     [Google Scholar]
  41. Thompson R. L., Sawtell N. M. 2000; Replication of herpes simplex virus type 1 within trigeminal ganglia is required for high frequency but not high viral genome copy number latency. J Virol 74:965–974 [View Article][PubMed]
     [Google Scholar]
  42. Thompson R. L., Preston C. M., Sawtell N. M. 2009; De novo synthesis of VP16 coordinates the exit from HSV latency in vivo. PLoS Pathog 5:e1000352 [View Article][PubMed]
     [Google Scholar]
  43. Wagner E. K., Bloom D. C. 1997; Experimental investigation of herpes simplex virus latency. Clin Microbiol Rev 10:419–443[PubMed]
     [Google Scholar]
  44. Wang T. S., Wong S. W., Korn D. 1989; Human DNA polymerase alpha: predicted functional domains and relationships with viral DNA polymerases. FASEB J 3:14–21[PubMed]
     [Google Scholar]
  45. Zhang J., Chung D. W., Tan C. K., Downey K. M., Davie E. W., So A. G. 1991; Primary structure of the catalytic subunit of calf thymus DNA polymerase delta: sequence similarities with other DNA polymerases. Biochemistry 30:11742–11750 [View Article][PubMed]
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.061903-0
Loading
Roles of conserved residues within the pre-NH2-terminal domain of herpes simplex virus 1 DNA polymerase in replication and latency in mice
J. Gen. Virol. 95, 940 (2014); https://doi.org/10.1099/vir.0.061903-0
/content/journal/jgv/10.1099/vir.0.061903-0
/content/journal/jgv/10.1099/vir.0.061903-0
Loading

Data & Media loading...

Most cited Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error