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. 1997 Apr;71(4):3039–3047. doi: 10.1128/jvi.71.4.3039-3047.1997

The product of the UL12.5 gene of herpes simplex virus type 1 is a capsid-associated nuclease.

J C Bronstein 1, S K Weller 1, P C Weber 1
PMCID: PMC191433  PMID: 9060664

Abstract

The UL12 open reading frame of herpes simplex virus type 1 (HSV-1) encodes a deoxyribonuclease that is frequently referred to as alkaline nuclease (AN) because of its high pH optimum. Recently, an alternate open reading frame designated UL12.5 was identified within the UL12 gene. UL12.5 and UL12 have the same translational stop codon, but the former utilizes an internal methionine codon of the latter gene to initiate translation of a 60-kDa amino-terminal truncated form of AN. Since the role of the UL12.5 protein in the HSV-1 life cycle has not yet been determined, its properties were investigated in this study. Unlike AN, which can be readily solubilized from infected cell lysates, the UL12.5 protein was found to be a highly insoluble species, even when isolated by high-salt detergent lysis. Since many of the structural polypeptides which constitute the HSV-1 virion are similarly insoluble, a potential association of UL12.5 protein with virus particles was examined. By using Western blot analysis, the UL12.5 protein could be readily detected in preparations of intact virions, isolated capsid classes, and even capsids that had been extracted with 2 M guanidine-HCl. In contrast, AN was either missing or present at only low levels in each of these structures. Since the inherent insolubility of the UL12.5 protein prevented its potential deoxyribonuclease activity from being assayed in infected-cell lysates, partially purified fractions of soluble UL12.5 protein were generated by selectively solubilizing either insoluble infected-cell proteins or isolated capsid proteins with urea and renaturing them by stepwise dialysis. Initial analysis of these preparations revealed that they did contain an enzymatic activity that was not present in comparable fractions from cells infected with a UL12.5 null mutant of HSV-1. Additional biochemical characterization revealed that UL12.5 protein was similar to AN with respect to pH optimum, ionic strength, and divalent cation requirements and possessed both exonucleolytic and endonucleolytic functions. The finding that the UL12.5 protein represents a capsid-associated form of AN which exhibits nucleolytic activity suggests that it may play some role in the processing of genomic DNA during encapsidation.

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Selected References

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  1. Banks L., Purifoy D. J., Hurst P. F., Killington R. A., Powell K. L. Herpes simplex virus non-structural proteins. IV. Purification of the virus-induced deoxyribonuclease and characterization of the enzyme using monoclonal antibodies. J Gen Virol. 1983 Oct;64(Pt 10):2249–2260. doi: 10.1099/0022-1317-64-10-2249. [DOI] [PubMed] [Google Scholar]
  2. Bronstein J. C., Weber P. C. Purification and characterization of herpes simplex virus type 1 alkaline exonuclease expressed in Escherichia coli. J Virol. 1996 Mar;70(3):2008–2013. doi: 10.1128/jvi.70.3.2008-2013.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Costa R. H., Draper K. G., Banks L., Powell K. L., Cohen G., Eisenberg R., Wagner E. K. High-resolution characterization of herpes simplex virus type 1 transcripts encoding alkaline exonuclease and a 50,000-dalton protein tentatively identified as a capsid protein. J Virol. 1983 Dec;48(3):591–603. doi: 10.1128/jvi.48.3.591-603.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Davison A. J., Davison M. D. Identification of structural proteins of channel catfish virus by mass spectrometry. Virology. 1995 Feb 1;206(2):1035–1043. doi: 10.1006/viro.1995.1026. [DOI] [PubMed] [Google Scholar]
  5. Deiss L. P., Chou J., Frenkel N. Functional domains within the a sequence involved in the cleavage-packaging of herpes simplex virus DNA. J Virol. 1986 Sep;59(3):605–618. doi: 10.1128/jvi.59.3.605-618.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Draper K. G., Devi-Rao G., Costa R. H., Blair E. D., Thompson R. L., Wagner E. K. Characterization of the genes encoding herpes simplex virus type 1 and type 2 alkaline exonucleases and overlapping proteins. J Virol. 1986 Mar;57(3):1023–1036. doi: 10.1128/jvi.57.3.1023-1036.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hoffmann P. J., Cheng Y. C. DNase induced after infection of KB cells by herpes simplex virus type 1 or type 2. II. Characterization of an associated endonuclease activity. J Virol. 1979 Nov;32(2):449–457. doi: 10.1128/jvi.32.2.449-457.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hoffmann P. J., Cheng Y. C. The deoxyribonuclease induced after infection of KB cells by herpes simplex virus type 1 or type 2. I. Purification and characterization of the enzyme. J Biol Chem. 1978 May 25;253(10):3557–3562. [PubMed] [Google Scholar]
  9. Kwong A. D., Frenkel N. Herpes simplex virus-infected cells contain a function(s) that destabilizes both host and viral mRNAs. Proc Natl Acad Sci U S A. 1987 Apr;84(7):1926–1930. doi: 10.1073/pnas.84.7.1926. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Martinez R., Sarisky R. T., Weber P. C., Weller S. K. Herpes simplex virus type 1 alkaline nuclease is required for efficient processing of viral DNA replication intermediates. J Virol. 1996 Apr;70(4):2075–2085. doi: 10.1128/jvi.70.4.2075-2085.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Martinez R., Shao L., Bronstein J. C., Weber P. C., Weller S. K. The product of a 1.9-kb mRNA which overlaps the HSV-1 alkaline nuclease gene (UL12) cannot relieve the growth defects of a null mutant. Virology. 1996 Jan 15;215(2):152–164. doi: 10.1006/viro.1996.0018. [DOI] [PubMed] [Google Scholar]
  12. McGeoch D. J., Dolan A., Frame M. C. DNA sequence of the region in the genome of herpes simplex virus type 1 containing the exonuclease gene and neighbouring genes. Nucleic Acids Res. 1986 Apr 25;14(8):3435–3448. doi: 10.1093/nar/14.8.3435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Newcomb W. W., Brown J. C. Induced extrusion of DNA from the capsid of herpes simplex virus type 1. J Virol. 1994 Jan;68(1):433–440. doi: 10.1128/jvi.68.1.433-440.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Oroskar A. A., Read G. S. A mutant of herpes simplex virus type 1 exhibits increased stability of immediate-early (alpha) mRNAs. J Virol. 1987 Feb;61(2):604–606. doi: 10.1128/jvi.61.2.604-606.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Poffenberger K. L., Roizman B. A noninverting genome of a viable herpes simplex virus 1: presence of head-to-tail linkages in packaged genomes and requirements for circularization after infection. J Virol. 1985 Feb;53(2):587–595. doi: 10.1128/jvi.53.2.587-595.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Shao L., Rapp L. M., Weller S. K. Herpes simplex virus 1 alkaline nuclease is required for efficient egress of capsids from the nucleus. Virology. 1993 Sep;196(1):146–162. doi: 10.1006/viro.1993.1463. [DOI] [PubMed] [Google Scholar]
  17. Strobel-Fidler M., Francke B. Alkaline deoxyribonuclease induced by herpes simplex virus type 1: composition and properties of the purified enzyme. Virology. 1980 Jun;103(2):493–501. doi: 10.1016/0042-6822(80)90206-8. [DOI] [PubMed] [Google Scholar]
  18. Szilágyi J. F., Cunningham C. Identification and characterization of a novel non-infectious herpes simplex virus-related particle. J Gen Virol. 1991 Mar;72(Pt 3):661–668. doi: 10.1099/0022-1317-72-3-661. [DOI] [PubMed] [Google Scholar]
  19. Thomas M. S., Gao M., Knipe D. M., Powell K. L. Association between the herpes simplex virus major DNA-binding protein and alkaline nuclease. J Virol. 1992 Feb;66(2):1152–1161. doi: 10.1128/jvi.66.2.1152-1161.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Varmuza S. L., Smiley J. R. Signals for site-specific cleavage of HSV DNA: maturation involves two separate cleavage events at sites distal to the recognition sequences. Cell. 1985 Jul;41(3):793–802. doi: 10.1016/s0092-8674(85)80060-x. [DOI] [PubMed] [Google Scholar]
  21. Vaughan P. J., Banks L. M., Purifoy D. J., Powell K. L. Interactions between herpes simplex virus DNA-binding proteins. J Gen Virol. 1984 Nov;65(Pt 11):2033–2041. doi: 10.1099/0022-1317-65-11-2033. [DOI] [PubMed] [Google Scholar]
  22. Weller S. K., Seghatoleslami M. R., Shao L., Rowse D., Carmichael E. P. The herpes simplex virus type 1 alkaline nuclease is not essential for viral DNA synthesis: isolation and characterization of a lacZ insertion mutant. J Gen Virol. 1990 Dec;71(Pt 12):2941–2952. doi: 10.1099/0022-1317-71-12-2941. [DOI] [PubMed] [Google Scholar]

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