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:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The interaction between herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) and two neural cell lines, mouse neuroblastoma (N1E-115) and rat glioma (C6-BU-1), was investigated. N1E-115 cells were permissive to both types of HSV. In C6-BU-1 cells, on the other hand, all the HSV-1 strains tested so far showed persistent infection, and the infectious virus of HSV-2 strains disappeared spontaneously. The HSV-2-infected C6-BU-1 cells were positive for HSV-2-specific DNA sequences, virus-specific RNA, HSV-2-specific antigens and thymidine kinase activity, when no infectious virus was detected. The HSV-2 was reactivated from those C6-BU-1 cells by superinfection with murine cytomegalovirus (MCMV), but not with UV-irradiated MCMV or human cytomegalovirus. The reactivated HSV-2 was identical to the parental virus, when examined by restriction endonuclease cleavage analysis.
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PMID:Interaction of herpes simplex virus type 2 with a rat glioma cell line. 285 Apr 49

Five thymidine kinase (TK)-deficient mutants of varicella-zoster virus derived from the Oka vaccine strain were examined to determine the state of transcription of the TK gene, and the DNA sequences of two of the mutants and the parent strain were investigated. Restriction endonuclease analysis revealed no major alteration of whole genome organization, but did reveal a minor change of the cleaved fragment in size. Northern blot analysis showed that the transcripts from the TK gene of all five mutants were of the same size as that of the parent strain (1.6 kb). DNA sequences of the TK gene of the parent strain and those of two mutants were determined. One had a base substitution (C to T) in the coding region of TK, resulting in the termination of amino acid elongation in the 68th amino acid of 341. The other had a two-base deletion (AT) in the coding region, resulting in a frame shift from the 126th amino acid and termination at the 162nd amino acid. Both mutants had premature stop codons which might result in the synthesis of nonfunctional TK polypeptides. Thus, the molecular basis of TK deficiency was determined.
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PMID:Molecular analysis of the thymidine kinase gene of thymidine kinase-deficient mutants of varicella-zoster virus. 285 53

The morphologically normal rat fibroblast cell line Rat-2 was used as a target cell type to test the transforming ability of a human papillomavirus (HPV-1a). To this end, molecularly cloned HPV-1a genomes were introduced into cultured Rat-2 cells in cotransfection experiments using a cloned herpes simplex virus thymidine kinase gene as a selectable phenotypic maker. In each of 13 HPV-1a-positive cell clones examined the papillomavirus DNA sequences were associated with the high molecular weight fraction of the cellular DNA, and restriction endonuclease plus Southern blotting analyses revealed patterns of hybridization which were consistent with integration of the viral genomes. Even Rat-2 clones containing multiple copies of the entire HPV-1a genome retained the normal, i.e. flat, cell morphology and were unable to grow in soft agar. De novo methylation of the HPV-1a sequences in many Rat-2 cell clones was evidenced by resistance of the viral DNA to complete cleavage with the HpaII endonuclease. Two out of three cell lines harbouring multiple copies of the HPV-1a genome contained detectable levels of HPV-1a transcripts, whereas no transcripts were detected in the third such cell line in which the viral HpaII sites were methylated virtually to completion. These results are consistent with the notion that HPV-1a genes are expressed inefficiently in Rat-2 cells; consequently integration of the viral DNA occurs, and there is no effect of the virus on the growth properties of this cell type. It is possible that methylation of the HPV-1a sequences is responsible for the low levels of expression of the viral genome.
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PMID:Introduction of cloned human papillomavirus 1a DNA into rat fibroblasts: integration, de novo methylation and absence of cellular morphological transformation. 298 96

The herpes simplex virus (HSV) type 1 dUTPase gene was inactivated by insertion of HindIII oligonucleotide linker sequences into the KpnI site within the coding region of the cloned gene. The mutated gene was introduced into wild type herpes simplex virus by marker rescue and the recombinants were identified by the acquisition of a HindIII site within genome map coordinates 0.69 to 0.70 and the failure to induce virus-specific dUTPase activity. A spontaneous dUTPase deficient mutant, which had an identical restriction endonuclease DNA pattern to wild type virus, was also isolated from this transfection experiment. Both types of dUTPase-negative mutants failed to induce a virus-specific 39,000 mol wt polypeptide. Cells infected with the insertional mutant contained instead a novel polypeptide about 40,000 mol wt. No abnormal virus specific polypeptide was detected in cells infected with the spontaneous mutant. We conclude that the 39,000 mol wt polypeptide induced by wild type HSV-1 is the virus-coded dUTPase. Since both types of mutants grew well in exponentially growing and serum-starved tissue culture cells in the absence of wild type helper virus, the dUTPase is not required for virus replication under these conditions. Thymidine kinase deficient, dUTPase deficient double mutants were constructed by recombination of a thymidine kinase insertional mutation into dUTPase deficient virus. These mutants also grew as well as wild type virus both in normal tissue culture cells and cells lacking the cellular thymidine kinase.
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PMID:Isolation and characterisation of herpes simplex virus type 1 mutants which fail to induce dUTPase activity. 300 29

We have demonstrated the presence of an Epstein-Barr virus (EBV)-coded thymidine kinase (TK) by producing biochemically transformed, TK-positive mammalian cell lines using either microinjection of whole EBV virions or calcium phosphate-mediated transfection of the SalI-B restriction endonuclease fragment of EBV DNA. Analysis of these cell lines showed that: (i) EBV DNA was present in the cell lines, (ii) sequences from the SalI-B restriction endonuclease fragment of EBV were expressed, (iii) a TK activity was present and (iv) a protein with antigenic cross-reactivity with the herpes simplex virus (HSV) TK was produced. The identity of the EBV TK gene was determined by demonstrating that a recombinant plasmid, which expressed the protein product of the BXLF1 open reading frame as a fusion protein, could complement TK- strains of E. coli. A comparison of the predicted amino acid sequences of the TK proteins of EBV and HSV-1 revealed significant regions of homology.
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PMID:Identification of an Epstein-Barr virus-coded thymidine kinase. 301 75

HeLa cells generally do not respond well to interferon (IFN). We have used is-1, an IFN-sensitive mutant of mengovirus, to select a clone of IFN-responsive HeLa cells (F-H12). At moderate levels of human alpha/beta IFN, is-1 yields were fivefold lower in these cells than in similarly protected control cells. In contrast, wild-type mengovirus, vesicular stomatitis virus and a wild-type and thymidine kinase-negative strains of herpes simplex virus type 1 grew equally well in both cell lines. By a cell survival assay, the F-H12 line was up to 100 times more responsive to IFN than the parental line when challenged by is-1. 2'-5'-Oligo(A)-dependent endonuclease activity was the same in both lines. These observations cannot be accounted for by enhanced induction of IFN following infection.
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PMID:Selection and characterization of an interferon-responsive clonal cell line of HeLa cells. 302 28

Herpes simplex virus strains, isolated from three immunocompromised patients whose infections showed clinical resistance to acyclovir, were studied as treatment progressed. Virus isolated from two patients remained sensitive to acyclovir throughout. Isolates from the third patient, who had received a prolonged course of oral acyclovir, showed a sharp decrease in drug sensitivity which corresponded to loss of thymidine kinase activity. No changes in restriction endonuclease profiles were observed in isolates from the same patient as treatment with acyclovir progressed.
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PMID:Clinical resistance to acyclovir of herpes simplex virus infections in immunocompromised patients. 302 52

Several different genital and non-genital HSV isolates were obtained from a patient with an acquired immune deficiency of unknown aetiology. The patient was initially treated with topical acyclovir (ACV) and later with topical and intravenous ACV. In spite of treatment with antiviral drugs the patient continued to shed virus and to have extensive genital ulcerations. Restriction endonuclease (RE) analyses of the viral DNA revealed that all the isolates had characteristic HSV-2 patterns and that there were three genetically distinct virus groups among the ten isolates tested. Three post-therapy isolates, with the same RE pattern, were found to be devoid of thymidine kinase activity (TKD), highly resistant to ACV in cell culture, but sensitive to vidarabine (ara-A), phosphonoacetate and phosphonoformate. Two of these TKD isolates were obtained during and after topical ACV therapy and before intravenous treatment. Mice inoculated intracerebrally with a lethal dose of each of the three TKD viruses were refractory to ACV, but responded to vidarabine or a combination of ACV and ara-A. Mice inoculated with the TK+ viruses (including the pre-therapy isolate) responded to ACV and/or ara-A treatment. The results indicate that: (i) TKD variants may be produced in humans after topical ACV therapy; (ii) different ACV-resistant or sensitive HSV-2 variants can establish latency at different body sites and reactivate; and (iii) when drug-resistant viruses are isolated from patients with multiple reactivations, the drug in question should not be discontinued, since the patients may also be shedding drug-sensitive virus at a different body site.
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PMID:Characterization of acyclovir-resistant and -sensitive herpes simplex viruses isolated from a patient with an acquired immune deficiency. 302 53

We have identified two regions of the herpes simplex virus type 1 (HSV-1) genome that inhibit DNA-mediated transformation of thymidine kinase-less L (Ltk-) cells by the cloned HSV-1 tk gene. When plasmids containing the EcoRI fragments EK or JK were mixed at 30 fmol/ml with the tk gene and transfected into Ltk- cells, the frequency of transformation was inhibited 80 to more than 90% relative to the control. Of the remaining 10 EcoRI fragments of the HSV-1 genome, 8 were inactive and 2 were weakly active. A 6.1-kilobase PstI subclone between 0.743 and 0.782 map units was isolated from pEK. This clone, pEK-P3P4, exhibited antitransformation activity toward HSV-1 tk and also the bacterial genes gpt and neo. pEK-P3P4 contains the alpha 27 gene, and restriction endonuclease inactivation and subcloning studies established that alpha 27 alone did not inhibit transformation. However, alpha 27 plus sequences both upstream and downstream of alpha 27 did inhibit transformation. In addition, alpha 0 or alpha 4 could substitute for alpha 27 in effecting antitransformation with these sequences. Therefore, an alpha gene and two additional loci in pEK-P3P4 are required for antitransformation. A second antitransforming locus in the reiterated sequences common to EK and JK and distinct from those in pEK-P3P4 was also identified but not characterized in detail. How antitransformation may be an expression of regulation of viral and host cell gene expression is discussed.
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PMID:Sequences of herpes simplex virus type 1 that inhibit formation of stable TK+ transformants. 304 Oct 18

An approach is devised for studying the role of DNA methylation in eukaryotic gene expression. The approach is based on the expression of site-specific bacterial methylase genes in animal cells. A model system using the cloned PaeR7 (an isoschizomer of Xho I) methylase gene was constructed to test the feasibility of this approach. Expression plasmids for the PaeR7 methylase gene were introduced into mouse Ltk- cells by cotransfection with the cloned chicken thymidine kinase (tk) gene. Several of the cell strains derived from Tk+ colonies were found to express the PaeR7 gene as judged by four criteria: the cellular DNA of these strains showed increased resistance to cleavage by Xho I; these strains contained cellular proteins that comigrated with pure PaeR7 methylase protein, as visualized by immunoblotting; PaeR7 methylase activity was found in vitro in crude extracts of total cellular protein from these strains; and murine adenovirus genomes grown on cells expressing PaeR7 methylase showed resistance to cleavage to PaeR7 endonuclease. The potential applications of this approach for the study of cellular and viral gene regulation, DNA repair, and restriction modification are discussed.
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PMID:Introduction and expression of the bacterial PaeR7 methylase gene in mammalian cells. 346 94


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