Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.1.21 (thymidine kinase)
7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pseudorabies virus (PRV) is a herpesvirus of pigs. Homologous recombination with plasmids offers a method to engineer precise changes in the PRV genome to produce advantageous live vaccines. Safety can be ensured by using a non-reverting deletion to inactivate the thymidine kinase gene. One particularly important feature of new PRV vaccines is deletion of an antigen, so that vaccinated pigs are serologically distinguishable from infected pigs. We have constructed a live vaccine strain with deletions in the thymidine kinase gene and in the gene for a glycoprotein, gX. Molecular engineering techniques made it possible to choose deletion of gX, which has no known immunological significance, over deletion of other glycoproteins that contribute to protective immunity. Extensive experiments in pigs with isogenic virus pairs show that deletion of gX does not compromise efficacy of a vaccine as gI deletions do. Deletion of gX also suggests a site for replacement with antigens from other pathogens. In addition to molecular engineering of a live vaccine strain, research on PRV glycoproteins has led to the discovery that expression of the glycoprotein gp50 makes cells resistant to PRV infection. Perhaps this observation could be extrapolated to the level of a whole animal to allow engineering of pigs to become an alternative to engineered vaccines.
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PMID:Genetic engineering of the pseudorabies virus genome to construct live vaccines. 217 Jun 41

The rate of transcription of the beta 2-adrenergic receptor gene is increased in response to beta-adrenergic agonist stimulation of the receptor at the cell surface. This effect is mediated by stimulation of adenylyl cyclase and elevation of intracellular cAMP levels. We have previously shown that this responsiveness to cAMP resides in the 5'-flanking region of the human beta 2-adrenergic receptor gene (Collins, S., Bouvier, M., Bolanowski, M. A., Caron, M. G., and Lefkowitz, R. J. (1989) Proc. Natl. Acad. Sci. U. S. A. 86, 4853-4857). A 34-base pair sequence derived from the beta 2-adrenergic receptor promoter region (-70 to -37 base pairs), containing the sequence GTACGTCA, confers responsiveness to cAMP when present in either orientation 5' to the thymidine kinase promoter on the chloramphenicol acetyltransferase reporter gene. Overexpression of the catalytic subunit of protein kinase A fully substituted for forskolin in inducing expression through this sequence, indicating that the cAMP induction is mediated through this kinase. Mutations within the GTACGTCA sequence completely abolished the stimulation. A 43-kDa transcription factor (cAMP response element-binding protein) confers cAMP responsiveness through binding to specific sequences. In gel mobility shift assays, purified cAMP response element-binding protein bound to the 34-base pair oligonucleotide from the beta 2-adrenergic receptor gene with an affinity similar to that for the well-characterized cAMP response element from the human glycoprotein hormone alpha-subunit gene, and failed to bind to mutated elements. Thus, positive autoregulation of the beta 2-adrenergic receptor gene appears to occur through receptor-mediated stimulation of adenylyl cyclase, with consequent activation of cAMP response element-binding protein and stimulation of beta 2-adrenergic receptor gene transcription. These results demonstrate a novel mechanism by which a receptor (beta 2-adrenergic receptor) stimulatory for adenylyl cyclase can exert positive feedback regulation on its own expression.
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PMID:A cAMP response element in the beta 2-adrenergic receptor gene confers transcriptional autoregulation by cAMP. 217 52

The safety of an Aujeszky's disease virus vaccine based on strain 783, a deletion mutant which does not express glycoprotein I and thymidine kinase, was assessed in pigs, calves and sheep. Four-day-old piglets which were inoculated intranasally and intramuscularly with 10(7) plaque forming units (PFU) developed only slight depression and fever. The virus was transmitted to a sentinel piglet. Six weeks after inoculation, the pigs were injected with high doses of corticosteroids in an attempt to reactivate the vaccine virus. The pigs did not shed Aujeszky's disease virus, did not develop a rise in virus neutralising antibody titres and sentinel pigs remained seronegative to Aujeszky's disease virus. Strain 783 was passaged in two series of three- to five-day old piglets, but after the third and fourth passages virus could no longer be recovered. Pregnant sows were inoculated with 10(7) PFU of virus strain 783 around day 35 or on day 85 of pregnancy, and their fetuses and piglets were assayed for Aujeszky's disease virus and antibodies against Aujeszky's disease virus. No evidence was found for transplacental transmission of the virus. Calves and sheep were given 10(7) PFU of virus strain 783 intranasally or intramuscularly; they survived and did not develop clinical signs of Aujeszky's disease. All the sheep and the calves inoculated intramuscularly developed neutralising antibodies to Aujeszky's disease virus.
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PMID:Safety of an Aujeszky's disease vaccine based on deletion mutant strain 783 which does not express thymidine kinase and glycoprotein I. 217 88

Bovine respiratory syncytial (BRS) virus causes a severe lower respiratory tract disease in calves similar to the disease in children caused by human respiratory syncytial (HRS) virus. While there is antigenic cross-reactivity among the other major viral structural proteins, the major glycoprotein, G, of BRS virus and that of HRS virus are antigenically distinct. The G glycoprotein has been implicated as the attachment protein for HRS virus. We have carried out a molecular comparison of the glycoprotein G of BRS virus with the HRS virus counterparts. cDNA clones corresponding to the BRS virus G glycoprotein mRNA were isolated and analyzed by dideoxynucleotide sequencing. The BRS virus G mRNA contained 838 nucleotides exclusive of poly(A) and had a major open reading frame coding for a polypeptide of 257 amino acid residues. The deduced amino acid sequence of the BRS virus G polypeptide showed only 29 to 30% amino acid identity with the G protein of either the subgroup A or B HRS virus. However, despite this low level of identity, there were strong similarities in the predicted hydropathy profiles of the BRS virus and HRS virus G proteins. A cDNA molecule containing the complete BRS virus G major open reading frame was inserted into the thymidine kinase gene of vaccinia virus by homologous recombination, and a recombinant virus containing the BRS virus G protein gene was isolated. This recombinant virus expressed the BRS virus G protein, as demonstrated by Western immunoblot analysis and immunofluorescence of infected cells. The BRS virus G protein expressed from the recombinant vector was transported to and expressed on the surface of infected cells. Antisera to the BRS virus G protein made by using the recombinant vector to immunize animals recognized the BRS virus attachment protein but not the HRS virus G protein and vice versa, confirming the lack of antigenic cross-reactivity between the BRS and HRS virus attachment proteins. On the basis of the data presented here, we conclude that BRS virus should be classified within the genus Pneumovirus in a group separate from HRS virus and that it is no more closely related to HRS virus subgroup A than it is to HRS virus subgroup B.
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PMID:Nucleotide sequence analysis and expression from recombinant vectors demonstrate that the attachment protein G of bovine respiratory syncytial virus is distinct from that of human respiratory syncytial virus. 221 24

We have determined the genomic location and nucleotide sequence of the equine herpesvirus 4 thymidine kinase (TK) gene. The gene is positioned at approximately 0.48 map units within the long unique component of the genome and is flanked by genes encoding a herpes simplex virus 1 (HSV-1) UL24 homologue and glycoprotein H. The predicted protein is composed of 352 amino acids, has an Mr of 38,800 and exhibits 36% identity to the predicted TK of HSV-1.
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PMID:The nucleotide sequence of the equine herpesvirus 4 thymidine kinase gene. 239

We constructed a recombinant herpes simplex virus (HSV) containing the transcribed coding and non-coding sequences of HSV-1 strain F glycoprotein B (gB) gene, a gamma 1 gene, fused to the promoter-regulatory sequences of the HSV-1 alpha 4 gene and inserted into the thymidine kinase gene of RH1G44, an HSV-1 x HSV-2 recombinant that contains an HSV-2 gB gene at the natural locus. Phenotypic analyses of the insertion mutant, R3145, showed that the alpha gB gene was transcribed in the presence of cycloheximide but underwent partial conversion to the HSV-2 form. Nucleotide sequencing of the gene indicated that the 5' crossover occurred between nucleotides 107 and 117 upstream from the translation initiation site and that the 3' crossover occurred between the sequences specifying amino acids 402 and 412 of the HSV-1 gB. The chimeric protein consisted of an N-terminal 405 to 415 amino acids encoded by the HSV-2 gene and a C-terminal 462 to 472 amino acids encoded by the HSV-1 gene. Comparison of the reactivity of the parental and recombinant gB with type-specific monoclonal antibodies indicated that the chimeric gB lost reactivity with four HSV-1-specific antibodies but gained reactivity with three HSV-2-specific antibodies.
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PMID:A subset of type-specific epitopes map in the amino terminus of herpes simplex virus type 1 glycoprotein B. 247 97

Previous studies demonstrated that the rabbit beta-globin gene is transcribed from its own promoter and regulated as a herpes simplex virus (HSV) early gene following insertion into the early HSV thymidine kinase gene in the intact viral genome (J. R. Smiley, C. Smibert, and R. D. Everrett, J. Virol. 61:2368-2377, 1987). We report here that the beta-globin promoter remained under early control after insertion into the late HSV gene encoding glycoprotein C. On the basis of these findings, we concluded that the beta-globin promoter is functionally equivalent to an HSV early-control region. We found that a transduced human alpha-globin gene was also regulated as an early HSV gene, while two linked Alu elements mimicked the behavior of HSV late genes. These results demonstrate that certain aspects of HSV temporal regulation can be duplicated by cellular elements and provide strong support for the hypothesis that the regulation of HSV gene expression can occur through mechanisms that do not rely on recognition of virus-specific temporal control signals.
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PMID:Regulation of cellular genes transduced by herpes simplex virus. 253 95

Two of the major glycoproteins of bovine herpesvirus 1 (BHV-1) are gI, a polypeptide complex with apparent molecular weights of 130,000, 74,000, and 55,000, and gIII (a 91,000-molecular-weight [91K] glycoprotein), which also exists as a 180K dimer. Vaccinia virus (VAC) recombinants were constructed which carry full-length gI (VAC-I) or gIII (VAC-III) genes. The genes for gI and gIII were each placed under the control of the early VAC 7.5K gene promoter and inserted within the VAC gene for thymidine kinase. The recombinant viruses VAC-I and VAC-III retained infectivity and expressed both precursor and mature forms of glycoproteins gI and gIII. The polypeptide backbones, partially glycosylated precursors, and mature gI and gIII glycoproteins were indistinguishable from those produced in BHV-1-infected cells. Consequently, they were apparently cleaved, glycosylated, and transported in a manner similar to that seen during authentic BHV-1 infection, although the processing efficiencies of both gI and gIII were generally higher in recombinant-infected cells than in BHV-1-infected cells. Immunofluorescence studies further demonstrated that the mature gI and gIII glycoproteins were transported to and expressed on the surface of cells infected with the respective recombinants. Immunization of cattle with recombinant viruses VAC-I and VAC-III resulted in the induction of neutralizing antibodies to BHV-1, which were reactive with authentic gI and gIII. These data demonstrate the immunogenicity of VAC-expressed gI and gIII and indicate the potential of these recombinant glycoproteins as a vaccine against BHV-1.
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PMID:Synthesis, cellular location, and immunogenicity of bovine herpesvirus 1 glycoproteins gI and gIII expressed by recombinant vaccinia virus. 253 9

We have constructed recombinant human adenovirus (Ad) vectors containing the glycoprotein gene of vesicular stomatitis virus (VSV). The structural gene of the VSV glycoprotein was modified by the addition of promoter and poly(A) addition sequences from the herpes simplex virus type 1 thymidine kinase (TK) gene and inserted, in either orientation, into early region 3 (E3) of human Ad type 5. The recombinant vectors were fully infectious and replicated in HeLa cells in culture. The TK promoter was functional in both insert orientations and responsive to trans-activation by herpes virus infection; however production of VSV glycoprotein in readily detectable amounts was only obtained with the vector having an insert in the E3 parallel orientation (AdG12), and depended principally on transcripts initiating within upstream Ad sequences. The onset of expression of the glycoprotein in AdG12-infected cells was detectable at about the same time as the Ad 72K DNA-binding protein encoded by E2, and its synthesis was not prevented by blocking viral DNA synthesis. The VSV glycoprotein produced by AdG12 was fully processed and could function to direct low pH-induced fusion of infected cells. These Ad vectors have considerable potential utility for the expression of antigens in cell culture and for the immunization of animals in studies of immunity and protection.
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PMID:Expression of the glycoprotein of vesicular stomatitis virus by infectious adenovirus vectors. 254 46

The spread of herpes simplex virus type 1 (HSV-1) strain KOS, and two less neurovirulent mutants of the strain was studied in female DBA/2 mice during the 1- to 5-day postinoculation period after intracerebral inoculation. Immunohistopathology showed that wild-type KOS virus first infected the meninges and ependymal cells but did not infect cells at the inoculation sites. The virus continued to spread to some cells directly adjacent to ventricles; however, the most extensive and severe lesions were found in the pyriform lobes and other structures associated with the limbic system. The pattern of spread suggested that direct cell to cell viral spread is important but that retrograde axonal transport to distant sites probably accounts for the more severe lesions associated with the limbic system. Both less neurovirulent mutant viruses multiplied to a much lesser degree in the brain and spread less extensively than the wild type virus when equivalent doses were given; however, when a large dose of the least neurovirulent mar C10.1 mutant virus was inoculated, infection spread rapidly to the same regions of the brain affected by KOS. Studies of mar C10.1 showed that thymidine kinase deficiency, rather than a mutation in the gene coding for glycoprotein C, probably accounted for the decreased neurovirulence of this mutant. This mouse model of HSV-1 virus-induced encephalitis, in combination with appropriate studies of the molecular biology of the HSV-1 KOS strain, should be useful for the study of neurovirulence factors contributing to the pathogenesis of HSV-1.
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PMID:Characterization of encephalitis in adult mice induced by intracerebral inoculation of herpes simplex virus type 1 (KOS) and comparison with mutants showing decreased virulence. 254 66


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