Mutant strains of herpes simplex deficient in thymidine kinase-inducing activity

Dubbs, D. R.; Kit, Saul

doi:10.1016/0042-6822(64)90070-4

Cited by 255 publications

(111 citation statements)

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“…Both deletion mutants completely lacked TK activity and reduced the TK activity of the infected cells below that of mock-infected controls. This effect on the host has been reported previously by Dubbs & Kit (1964) and Leiden et al (1976).…”

Section: Thymidine Kinase Assayssupporting

confidence: 60%

Thymidine Kinase Deletion Mutants of Herpes Simplex Virus Type 1

Sanders

Wilkie

Davison

1982

Journal of General Virology

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277 SUMMARYDeletions in the cloned thymidine kinase (TK) gene of herpes simplex virus type 1 (HSV-I), strain 17 syn ÷, were produced by two methods. Removal of a 506 base pair fragment from between the unique SstI and BglII restriction endonuclease sites of pTK1 (HSV-1 BamHI p cloned in pAT153) and subsequent transformation of Escherichia coli resulted in the isolation of 50 deleted plasmids. Sequential digestion of pTK1 with BgllI and nuclease BAL 31 followed by ligation and recleavage with BglII resulted in the isolation of 31 deleted plasmids. Three clones, pTK2, pTK3 and pTK4, obtained following BglII and SstI treatment of pTK 1 were recombined with wild-type (wt) HSV-1 (17) syn ÷ DNA in baby hamster kidney (BHK) cells to produce TKdeletion mutants HSV-1 (17)TK 1301, HSV-1 (17)TK 1302 and HSV-1 (17)TK 1303 respectively. 5-Bromo-2'-deoxyuridine, 5-bromo-2'-deoxycytidine and 9-(2-hydroxyethoxymethyl)guanine were used to reduce the background of TK + virus in heterogeneous recombinant stocks analysed for the presence of TK-recombinants. All recombinant clones isolated produced a small syncytial plaque morphology in BHK cells. The mutants HSV-1 (17) TK 1301 and HSV-I (17) TK 1302 were TK-, failed to produce polypeptides of molecular weights 43 000 and 19 000 found in wt-infected cells and demonstrated one-step growth curves different from wt virus and the TK-mutant HSV-I (17)dPyk -7. Superinfection studies with HSV-1 (17)TK 1301, HSV-1 (17)TK 1302, HSV-I (MDK) and HSV-1 (17) dPyk -7 indicated that all TK-mutants except dPyK -7 produce a tram-acting gene product which can switch on the transforming HSV-1 TK gene.

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Section: Thymidine Kinase Assayssupporting

confidence: 60%

Thymidine Kinase Deletion Mutants of Herpes Simplex Virus Type 1

Sanders

Wilkie

Davison

1982

Journal of General Virology

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“…This enzyme is induced if cells are about to synthesize DNA either in the normal cell cycle [13-i5], or following viral infection [16]. Cellular and viral mutants lacking thymidine kinase show however, that the enzyme is, not indispensable for DNA replication [17,18].…”

Section: Discussionmentioning

confidence: 99%

On the Synthesis of RNA in Lymphocytes Stimulated by Phytohemagglutinin. 1. Induction of Uridine-Kinase and the Conversion of Uridine to UTP

Hausen

Stein

1968

Eur J Biochem

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Conversion of uridine to UTP is enhanced in lymphocytes under the influence of phytohemagglutinin concomitant with the induction of uridine kinase. The uridine kinase activity in the induced cells decreases if the cells are treated with antibodies against phytohemagglutinin. The phosphorylation of uridine may be the limiting step in the incorporation of uridine into RNA. Uridine incorporation can thus not be taken as a direct measure of RNA synthesis.

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“…Vero cells were not fully permissive for the replication of HVS strains SMH 1 and HOT or HVA-73, but these viruses gave yields of about 1 p.f.u./cell (2 x 106 to 4 x 10 6 p.f.u. and their relevant properties are described elsewhere (Honess & Watson, 1977a;Honess et al, 1980;Dubbs & Kit, 1964;Pereira et al, 1976 Conditions for high multiplicity infections and labelling infected cultures with radioactive precursors. Monolayer cultures of OMK or Vero cells (2 x 10 6 to 4 × 106 cells per 25 cm 2 flask; 1 x 107 to 2 x 107 cells per 75 cm 2 flask) were inoculated with 2 to 10 p.f.u./cell of HVS or HSV strains in volumes of 1 (25 cm 2 flasks) or 5 to 10 ml (75 cm 2 flasks) and incubated at 37 °C (or 34 °C for ts viruses) with continuous agitation for 1 to 2 h. Thereafter, inocula were decanted and replaced with culture medium and the cultures reincubated.…”

Section: Introductionmentioning

confidence: 99%

Comparison of Thymidine Kinase Activities Induced in Cells Productively Infected with Herpesvirus Saimiri and Herpes Simplex Virus

Honess¹,

O’Hare²,

Young³

1982

Journal of General Virology

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SUMMARYThe replication of unselected strains of herpesvirus saimiri (HVS) was sensitive to bromodeoxyuridine and bromovinyldeoxyuridine (BVdU) but insensitive to acycloguanosine (ACG), in contrast to the growth of herpes simplex virus (HSV) which was sensitive to all three analogues. Mutants of HVS resistant to bromodeoxyuridine and BVdU could be selected by growth in the presence of these inhibitors. Productive infections of owl monkey kidney or Vero cell cultures by unselected strains of HVS resulted in increases in a thymidine kinase (TK) activity which was deficient in cells infected with bromodeoxyuridine-resistant mutants of the virus. Induction of the virus enzyme promoted a net increase in the uptake and incorporation of exogenous labelled thymidine in the face of the progressive inhibition of the overall incorporation of [35S]methionine and [3H]uridine into productively infected cells. The TK induced in cells infected with HVS differed from the major activity of uninfected cells and resembled that encoded by HSV in its capacity to phosphorylate iododeoxyuridine and in the sensitivity of all the thymidine phosphorylating activity to competition by BVdU. However, in contrast to the HSV TK, which phosphorylated deoxycytidine and iododeoxycytidine relatively efficiently and was sensitive to ACG, the HVS enzyme did not phosphorylate deoxycytidine or iododeoxycytidine and was insensitive to ACG. Whilst HVS, therefore, shares the characteristic of other members of the herpesvirus group of inducing a novel TK, the properties of the HVS-induced enzyme differ significantly from the enzyme of the prototype herpesvirus, HSV. The properties of the HVS TK are nonetheless sufficiently distinct from those of the uninfected cell to provide a possible basis for selective antiviral chemotherapy based on preferential phosphorylation of nucleoside analogues such as BVdU by infected cells.

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Mutant strains of herpes simplex deficient in thymidine kinase-inducing activity

Cited by 255 publications

References 12 publications

Thymidine Kinase Deletion Mutants of Herpes Simplex Virus Type 1

Thymidine Kinase Deletion Mutants of Herpes Simplex Virus Type 1

On the Synthesis of RNA in Lymphocytes Stimulated by Phytohemagglutinin. 1. Induction of Uridine-Kinase and the Conversion of Uridine to UTP

Comparison of Thymidine Kinase Activities Induced in Cells Productively Infected with Herpesvirus Saimiri and Herpes Simplex Virus

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