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)

The X. laevis tRNA((Ser)Sec) gene is different from the other tRNA genes in that its promoter contains two external elements, a PSE and a TATA box functionally equivalent to those of the U6 snRNA gene. Of the two internal promoters governing classical tRNA gene transcription, only subsists the internal B box. In this report, we show that the tRNA((Ser)Sec) contains in addition an activator element (AE) which we have mapped by extensive mutagenesis. Activation is only dependent on a 15 bp fragment residing between -209 and -195 and containing an SPH motif. In vitro, this element forms a complex with a nuclear protein which is different from the TEF-1 transcriptional activator that binds the SV40 Sph motifs. This AE is versatile since it shows capacity of activating a variety of genes in vivo, including U1 and U6 snRNAs and HSV thymidine kinase. Unexpectedly for an snRNA-related gene, the tRNA((Ser)Sec) is deprived of octamer or octamer-like motifs. The X.laevis tRNA((Ser)Sec) gene represents the first example of a Pol III snRNA-type gene whose activation of transcription is completely octamer-independent.
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PMID:Optimal tRNA((Ser)Sec) gene activity requires an upstream SPH motif. 131 Oct 68

We have derived Vero cell lines containing the herpes simplex virus DNA polymerase (pol) gene that complement temperature-sensitive pol mutants. These cell lines were used to recover viruses containing new mutations at the pol locus. Two spontaneously arising host-range mutants, 6C4 and 7E4, were isolated. These mutants did not grow efficiently on Vero cells or synthesize late polypeptides but formed plaques on a cell line containing the pol gene (DP6 cells). Whereas mutant 6C4 specified a wild-type-size Pol protein, we detected no full-length Pol protein in 7E4-infected cell extracts. Complementation studies demonstrated that 6C4 and 7E4 contain different mutations and indicated that 6C4 is in a complementation group different from that of pol temperature-sensitive mutant tsC7 or tsD9. A mutant in which 2.2 kilobases of pol sequences were replaced with the Escherichia coli lacZ gene under the control of the herpes simplex virus thymidine kinase promoter was constructed. This mutant formed blue plaques on DP6 cells in the presence of 5-bromo-4-chloro-3-indolyl-beta-D-galactoside. Using this virus in marker rescue experiments, we engineered three mutants containing deletions in the pol coding region which grew efficiently on DP6 cells but not on Vero cells and which differed in their synthesis of Pol polypeptides. The lacZ insertion virus was also used to introduce a deletion in the region upstream of the pol long open reading frame, which removes a short open reading frame that could encode a 10-amino-acid peptide. This mutant grew to similar titers on Vero and DP6 cells, indicating that these sequences are not essential for growth of the virus in tissue culture.
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PMID:Isolation and characterization of herpes simplex virus mutants containing engineered mutations at the DNA polymerase locus. 215 81

A subline of Ehrlich ascites carcinoma (EAC) cells resistant to 5-fluoro-2'-deoxy-uridine (FdUrd) was developed by continuous exposure to progressively increasing concentrations of the drug (35-75 mg/kg per day) during 15 passages through mice. Since then, the EAC cells have been retransplanted more than 80 times through drug-untreated mice and continue to be resistant. After adaptation to growth in suspension culture the drug-adapted cells were 1000 times more resistant to FdUrd in comparison with parental ones, and remained near-tetraploid with doubling time longer than in parental line. The activity of thymidine kinase was deeply depressed (100-fold) whereas that of thymidylate synthetase several-fold increased in the resistant EAC cells, both grown in vivo and in vitro.
Acta Biochim Pol 1983
PMID:Development and characteristics of a subline of Ehrlich ascites carcinoma cells persistently resistant to 5-fluoro-2'-deoxyuridine. 622 75

We used partially purified RNA polymerase II from uninfected (Pol II) and from herpes simplex virus type 1 (HSV-1) infected HEp-2 cells (Pol II-H) to transcribe HSV-1 DNA in vitro. Gel electrophoretic analysis of the products produced from native HSV-1 DNA yielded weight average chain lengths of 4.0 and 3.5 kb for the Pol II and Pol II-H products, respectively. Blot hybridization analyses of the HSV DNA transcripts showed that both enzymes transcribed RNA from essentially all regions of the genome. However, Pol II preferentially transcribed regions coding for the immediate-early or alpha mRNAs, whereas Pol II-H preferentially copied regions coding for the early (beta) and late (gamma) gene products. Transcriptional analyses of the cloned HSV-1 Bam HI-Q fragment (containing the thymidine kinase (TK) gene) and its subfragments showed that (1) the major transcripts produced by Pol II-H were distinctly different from those produced by Pol II; (2) Pol II and Pol II-H utilized different promoters for the synthesis of major transcripts; (3) both enzymes produced three minor transcripts that were partially overlapping and in opposite direction to the TK gene; and (4) only Pol II-H initiated transcription from the TK promoter. In contrast, both Pol II and Pol II-H generated an identical set of transcripts from an adenovirus 2 early region DNA fragment. The sizes of the products suggest that RNA processing may be occurring in vitro. These results show that HSV-1 infection alters the in vitro transcriptional specificity of RNA polymerase II and demonstrate that this system should be useful for studying in vitro the regulation of gene transcription.
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PMID:Regulation of herpes simplex virus gene transcription in vitro. 629 54

Human immunodeficiency virus (HIV) reverse transcriptase substitutes for temperature-sensitive DNA polymerase I (Pol Its) in Escherichia coli, providing a screen for anti-HIV reverse transcriptase nucleoside analogs in bacteria. Since phosphorylation of nucleosides in E. coli is limited to thymidine and its derivatives, we coexpressed herpes simplex virus thymidine kinase, an enzyme that phosphorylates a wide variety of nucleoside analogs, together with HIV reverse transcriptase. Coexpression of herpes simplex virus thymidine kinase and HIV reverse transcriptase rendered Pol Its cells sensitive to dideoxycytidine. Studies with different nucleoside analogs indicate that this bacterial screening system is able to select and identify nucleoside analogs that specifically target HIV reverse transcriptase.
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PMID:A screen in Escherichia coli for nucleoside analogs that target human immunodeficiency virus (HIV) reverse transcriptase: coexpression of HIV reverse transcriptase and herpes simplex virus thymidine kinase. 754 49

The mutation spectrum attributable to an antimutator DNA polymerase in a eukaryotic cell was examined. Drug-resistant thymidine kinase (tk) mutants derived from both a wild-type (wt) strain, KOS and an antimutator DNA polymerase (pol) mutant, PAAr5, of herpes simplex virus (HSV), were isolated, and the mutated tk genes were characterized at the sequence level. While both transition and frameshift mutations were found in the mutated tk genes derived from the wt KOS Pol, all the PAAr5-mediated mutants analyzed were frameshift mutations. These results imply that the wt HSV Pol is less faithful than the antimutator enzyme, at least in part, because of its propensity to mediate transition mutations.
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PMID:An altered spectrum of herpes simplex virus mutations mediated by an antimutator DNA polymerase. 783 98

The mammalian deoxyribonucleoside kinases thymidine kinase 1 and 2, deoxycytidine kinase and deoxyguanosine kinase phosphorylate deoxyribonucleosides and provide an alternative to de novo synthesis of DNA precursors. Their activities are essential for activation of several chemotherapeutically important nucleoside analogs. These four salvage kinase enzymes exhibit distinct substrate specificities for nucleoside analogs modified in the base and glycon moieties. In this review their. structure-activity relationships are discussed. Alternative routes for phosphorylation of nucleoside analogs are also reviewed, such as the phosphotransfer capacity of 5'-nucleotidase and protein kinases.
Acta Biochim Pol 1996
PMID:Structure-activity relationships for phosphorylation of nucleoside analogs to monophosphates by nucleoside kinases. 879 Jul 20

The organization of eukaryotic chromosomes into topological domains has led to the assumption that DNA topology and perhaps supercoiling are involved in eukaryotic nuclear processes. Xenopus oocytes provide a model system for studying the role of DNA topology in transcription. Linear plasmid templates for RNA polymerases (Pols) I and II are not transcribed in Xenopus oocytes, while circular templates are transcriptionally active. Here we show that circularity is not required for transcription of Pol I or Pol II promoters if the linear template is sufficiently long (> 17 to 19 kb). The Xenopus rRNA (Pol I) promoter is active in central positions on a long linear template but is not transcribed when located near an end. Because supercoils generated by transcription could be retained by viscous drag against the long template, these results are consistent with a supercoiling requirement for this promoter. Surprisingly, the herpes simplex virus thymidine kinase (Pol II) promoter is active even 100 bp from the end of the long template, indicating that template length fulfills a critical parameter for transcription that is not consistent with a supercoiling requirement. These results show that DNA length has unrecognized importance for transcription in vivo.
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PMID:DNA length is a critical parameter for eukaryotic transcription in vivo. 881 96

In the majority of cases, the mechanism underlying the resistance to acyclovir (ACV) of herpes simplex viruses (HSVs) is thymidine kinase (TK) deficiency. Plaque isolates from eight ACV-resistant (ACVr) clinical isolates from AIDS patients, of which five reactivated, were sequenced to determine the genetic lesion within the tk gene conferring resistance and whether this may have correlated with reactivation potential. Mutations were clustered within two homopolymer nucleotide stretches. Three plaque isolates (1737-14, 90-150-3, and 89-650-5) had insertion mutations within a stretch of 7 guanosines, while two isolates (89-063-1 and 89-353-1) had frameshift mutations within a stretch of 6 cytosines (a deletion and an insertion, respectively). Mutations resulted in premature termination codons, and the predicted 28- and 32-kDa truncated TK products were detected by Western blot analysis of virus-infected cell extracts. The repair of one homopolymer frameshift mutation (in isolate 1737-14) restored TK activity, demonstrating that this mutation is the basis of TK deficiency. Of the five reactivated isolates, four were TK deficient and contained frameshift mutations while the fifth retained TK activity because of its altered-TK or Pol- phenotype. These data demonstrate that the majority of ACVr clinical isolates contain frameshift mutations within two long homopolymer nucleotide stretches which function as hot spots within the HSV tk gene and produce nonfunctional, truncated TK proteins.
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PMID:Homopolymer mutational hot spots mediate herpes simplex virus resistance to acyclovir. 909 63

We have isolated and characterized a cDNA encoding a transcription activating factor for the mouse selenocysteine tRNA (tRNAsec) gene from mouse mammary gland. The full-length cDNA, designated m-Staf, has a 1878-base pair open reading frame encoding 626 amino acids. The predicted amino acid sequence of m-Staf is highly homologous to that of Staf, another selenocysteine tRNA gene transcription activating factor of Xenopus laevis. Like Staf, m-Staf contains seven tandemly repeated zinc fingers and four repeated motifs. Gel shift assays indicated that the recombinant m-Staf specifically bound to the activator element region in the mouse tRNAsec gene. Transient co-transfection experiments in Drosophila Schneider cells, which lack endogenous Staf-like binding activity, showed that m-Staf increased the mouse tRNAsec gene transcription about 15-fold, whereas it stimulated Pol II-dependent thymidine kinase promoter only 2-fold. Northern blot analysis detected the presence of a 3.4-kilobase pair m-Staf transcript, which was widely but differentially expressed in various murine tissues. The binding activity of m-Staf in mouse mammary gland was undetectable during virgin and postlactating periods but increased markedly in parallel with the increase of tRNAsec transcript during the periods of pregnancy and lactation, when the gland undergoes growth and development. These results indicate that m-Staf is a transcriptional activator of the mouse tRNAsec gene and that its binding activity in the mammary gland undergoes developmental alterations.
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PMID:Molecular cloning and characterization of the murine staf cDNA encoding a transcription activating factor for the selenocysteine tRNA gene in mouse mammary gland. 953 33


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