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M Esteban

Publications and source records attributed to M Esteban.

At least 163 records · Page 9Linked to original sources
PubMed1984

Expression of cloned vaccinia virus DNA sequences introduced into animal cells.

Individual cloned HindIII fragments of vaccinia virus DNA were introduced into cells by DNA-mediated gene transfer. The presence of individual fragments in the different transformants was confirmed by Southern blot hybridization analysis. Several of the transformants were found to express viral sequences at various levels. The sizes of the transcripts containing vaccinia virus sequences were highly heterogeneous, with no discrete species of RNA. Positive clones contained vaccinia virus sequences in both the poly(A)+ and poly(A)- RNA fractions, although the prevalence of these sequences was variable in the two fractions. The S1 nuclease map of the 5' end of the transcripts from transformants containing the HindIII-J fragment revealed a unique 5' end, similar to RNA from virus-infected cells. In contrast, analysis of the 3' end of RNAs from these transformants showed a high degree of heterogeneity, which might explain the heterogeneity found in Northern blot patterns. In this report, it is shown for the first time that in cells transformed with vaccinia virus DNA there is proper initiation for, at least, the viral thymidine kinase gene.

Cells, Cultured↗
PubMed1984

Indiscriminate degradation of RNAs in interferon-treated, vaccinia virus-infected mouse L cells.

In this report we used Northern blot hybridization analysis to characterize the fate of several species of viral RNA transcribed from internal and terminal regions of vaccinia DNA in interferon-treated, infected mouse L cells grown in suspension. All species of viral RNAs were expressed but were reduced in amount. Larger-sized RNAs were reduced more than smaller-sized RNAs. This reduction appears to be related to the activation of the interferon-mediated double-stranded RNA-dependent 2-5A synthetase-endoribonuclease system, as the rRNA cleavage pattern characteristic of this system was observed early in infection and in cell extracts in response to exogenous 2-5A. Thus, in interferon-treated, vaccinia-infected mouse L cells in suspension, there is indiscriminate degradation of viral and cellular RNAs, and this RNA breakdown might play a role in the interferon-mediated inhibition of protein synthesis.

Animals↗
PubMed1983

The relationship between the antiviral action of interferon and prostaglandins in virus-infected murine cells.

The relationship between prostaglandins (PG) and interferon (IFN) was investigated. IFN induced the synthesis of immunoreactive PGE and PGA at early and late stages, respectively, of vaccinia virus infection in mouse L fibroblasts. Only species-specific IFN possessed this activity and PG synthesis was stimulated in virus-infected cells, while normal L cells were not affected. The vaccinia virus infection did not significantly alter PG synthesis in the absence of IFN. Indomethacin increased the rate of vaccinia virus replication and partially inhibited the IFN-induced protection of L cells. The addition of exogenous PGA1 only partially reversed this effect. Finally, short-term PGA treatment induced the synthesis of two enzymes (protein kinase and 2,5A synthetase) thought to be partially responsible for the antiviral action of interferon. These findings suggest that a prostaglandin or PG-related compound seems to mediate at least one aspect of IFN action.

2',5'-Oligoadenylate Synthetase↗
PubMed1983

Thymidine kinase genes and the induction of anti-viral responses by interferon.

A mouse fibroblast cell-line deficient in thymidine kinase (Ltk(-) aprt(-)) fails to show an anti-viral response when treated with interferon. After introduction of a viral tk gene into these cells the resultant clones showed normal responses to interferon. However, one such tk-containing clone (C6) spontaneously lost its ability to respond to interferon by inducing an antiviral state although it retained its ability to induce the enzyme oligo(2'-5' A)-synthetase. This sub-clone (6A) still expressed thymidine kinase activity but restriction endonuclease analysis indicated an alteration in the sequences flanking the exogenous viral tk gene. Our results suggest that a modification in the exogenous viral DNA sequences led to a loss of interferon sensitivity.

2',5'-Oligoadenylate Synthetase↗
PubMed1983

Electron microscopic studies of transcriptional complexes released from vaccinia cores during RNA-synthesis in vitro: methods for fractionation of transcriptional complexes.

Electron microscopic (EM) and biochemical methods were employed to study the transcriptional complexes present in detergent lysates of vaccinia virus cores actively synthesizing RNA in vitro. When processed and examined in the EM, 14 'transcriptional sites' could be observed on full-length DNA templates. Fractionation of lysates by equilibrium density centrifugation in CsSO4, chromatography on hydroxyapatite columns or by sedimentation in sucrose gradients, allowed isolation of DNA templates associated with transcripts but these manipulations often resulted in fragmentation of the DNA template or promoted the release of transcripts from the template. It is suggested that RNA transcripts remain associated with the template in regions of supercoiling. These regions, in turn, may be maintained by DNA-protein interactions which are compromised as the transcriptional complexes are fractionated and purified.

Centrifugation, Density Gradient↗
PubMed1983

Induction of an antiviral response by interferon requires thymidine kinase.

Mouse fibroblastoid cells (Ltk-) that lack thymidine kinase (tk) activity are unable to respond to murine beta-interferon by establishing antiviral activity or inducing the double-stranded RNA-dependent enzymes, oligo[(2'-5')A] polymerase and Mr 68,000 protein kinase. In contrast, the parental L-929 cell line or clonal derivatives of Ltk- cells into which the herpes virus tk gene was introduced by DNA-mediated gene transfer respond normally to interferon in developing resistance to viral infection and in inducing double-stranded RNA-dependent enzymes. Further evidence for a role of tk in the response to interferon was obtained by isolating revertants of tk+ clones that lost the herpes virus tk gene during growth in BrdUrd-containing medium. In such revertant sublines both tk enzyme activity and viral tk genes were undetectable and treatment with interferon failed to produce an antiviral effect or induce synthesis of the double-stranded RNA-dependent enzymes. Our results indicate that the ability of mouse L cells to respond to beta-interferon is dependent upon the presence of a functional tk gene. We propose that the induction of antiviral responses by interferon stringently requires a metabolite, the level of which is determined by tk activity. The system described may provide a means for elucidating the mechanisms by which responses to interferon are induced.

Adenine Nucleotides↗
PubMed1983

Prostaglandin A inhibits the replication of vesicular stomatitis virus: effect on virus glycoprotein.

Prostaglandins of the A series were found to strongly suppress the replication of vesicular stomatitis virus (VSV) in mouse L fibroblasts. The highest non-toxic dose of PGA1, 4 micrograms/ml, decreased VSV production by 93.6%. At this dose, PGA1 did not alter DNA, RNA or protein synthesis in uninfected L cells for periods up to 24 h, whereas it further suppressed protein synthesis and slightly increased RNA synthesis in VSV-infected cells. The presence of PGA1 during virus adsorption, with no treatment after infection, reduced VSV yields by 63.6%. However, the presence of PGA1 during an early step of VSV replication was not essential for the antiviral action to occur (PGA1 treatment could be started 1 to 2 h post-infection). Apart from a slight overall inhibition of virus protein synthesis, PGA1 strongly suppressed the synthesis of the VSV glycoprotein G; moreover, it produced an alteration in the mobility of this protein in SDS-polyacrylamide gels. We propose that this slight decrease in molecular weight (about 4000) of the G protein in the presence of PGA1 could be due to an alteration in the glycosylation process.

Animals↗
PubMed1982

Antiviral effect of prostaglandins of the A series: inhibition of vaccinia virus replication in cultured cells.

Prostaglandins of the A series potently inhibited the production of vaccinia virus in mouse L fibroblasts. With the highest non-toxic dose of PGA1, 4 micrograms/ml, the replication of the virus was inhibited by 95 . 3%. The antiviral activity was dose-dependent and specific for the A series. At the dose used, PGA1 was not toxic to uninfected cells and did not alter cell metabolism as measured by DNA, RNA and protein synthesis. PGA1 did not influence the adsorption of the virus by the host cells and the antiviral activity was not dependent on the presence of PGA1 during the early stages of infection. PGA treatment delayed and partially inhibited virus DNA synthesis and, while it did not produce any change in the pattern of protein synthesis in uninfected cells, it altered both the rate and the pattern of virus protein synthesis. We conclude that PGA1 selectively inhibits one or more steps involved in the replication of vaccinia virus in mouse L fibroblasts.

Adsorption↗