PubMed Health⌕ Search

Biomedical subjects

A Panet

Publications and source records attributed to A Panet.

At least 73 records · Page 4Linked to original sources
PubMed1989

Cytocidal effect caused by the envelope glycoprotein of a newly isolated avian hemangioma-inducing retrovirus.

We isolated a field strain of avian hemangioma retrovirus (AHV) which induces a cytopathic effect (CPE) on cultured avian and mammalian cells shortly after infection. The kinetics of cell killing were dependent on the multiplicity of infection. The CPE on avian and mammalian cells was independent of virus replication, because UV-irradiated virus led to cell death as well. Biochemical and genetic experiments indicated that AHV env gene products were responsible for the CPE. Partially purified AHV envelope glycoproteins (gp85), but not those of the Rous sarcoma virus Prague C strain, induced a CPE. Rous-associated virus type 1, in which the env region was replaced by the AHV gp85 region, induced a CPE on avian and mammalian cultured cells. Therefore, we suggest that CPE is induced by AHV via interaction between viral gp85 and the cell membrane. This mode of CPE is unique among avian sarcoma-leukemia viruses.

Alpharetrovirus↗
PubMed1989

Translation of gag, pro, and pol gene products of human T-cell leukemia virus type 2.

Sequence analysis of human T-cell leukemia proviral DNA revealed three open reading frames arranged at a -1 position relative to one another. On the basis of homology to other retroviruses, these open reading frames were assigned to the gag, pro, and pol genes. To characterize the primary protein products of these genes and their modes of synthesis, a DNA clone of human T-cell leukemia virus type 2 was transcribed and translated in vitro. Analysis of the viral proteins revealed three polyproteins with molecular masses of 58, 75, and 112 kilodaltons at relative frequencies of 100:13:0.9, respectively. These proteins were mapped on the viral genome by both internal deletions and 3'-end truncations at gag, pro, and pol, respectively. The results indicate that translation of the pol gene requires two independent frameshift events, and the readthrough frequencies at the two frameshift sites appeared to be similar.

Chromosome Mapping↗
PubMed1988

Isolation and characterization of interferon-resistant variants from S49 mouse lymphoma.

S49 mouse lymphoma cells were found to be extremely sensitive to the antiproliferative activity of interferon. These characteristics were studied to select for IFN-resistant cell variants. Some 0.6% of the parental S49 cell population were resistant to the antiproliferative and cytotoxic activities of IFN. The resistant cells were cloned and analyzed for their responses to several of the activities of IFN, namely, inhibition of encephalomyocarditis (EMC) virus, murine leukemia virus (MuLV) replications, and the induction of (2'-5') oligoadenylate synthetase. Among the clones selected some were highly resistant while others demonstrated only partial responsiveness to IFN. S49 cells demonstrate tubular structures in the cytoplasm. These structures were previously reported to be antigenically related to mouse mammary tumor virus (MMTV). We report here that IFN treatment decreases the expression of these cytoplasmic viral structures as revealed by electron microscopy. To correlate this novel antiviral activity to the more established functions of IFN we utilized the above mentioned S49 IFN-resistant variants. The anti-MMTV activity of IFN correlated with the other effects of IFN in both the highly resistant and partially responsive S49 clones. Our findings indicate that a relatively high proportion of S49 cells vary in their response to IFN. The defect in the resistant cells appears to affect a primary response to IFN which is common to its diverse activities. Furthermore, the effect of IFN on MMTV-related structures involves the usual pathway of IFN action.

2',5'-Oligoadenylate Synthetase↗
PubMed1988

Inhibition of transcription of herpes simplex virus immediate early genes in interferon-treated human cells.

The effect of interferon (IFN) treatment on the early stages of herpes simplex virus type 1 (HSV-1) replication in three types of human cells was investigated. Interferon pretreatment was shown to reduce the steady state levels of both total and polysomebound HSV-1 immediate early alpha mRNAs. Using the nuclear run-off transcription assay, we showed that IFN selectively inhibited transcription of the HSV-1 genes, with no effect on transcription of total cellular RNA or that of the beta-tubulin RNA. Thus, IFN appears to inhibit the initiation of HSV-1 alpha gene transcription rather than affect the stability of the respective mRNAs. IFN did not prevent the HSV-1-induced early shut-off of host cellular protein synthesis caused by a structural protein of infecting virus. This observation indicated that the IFN-mediated inhibition of HSV-1 replication is at a stage beyond viral penetration into the cytoplasm. These results suggested that IFN blocked HSV-1 replication primarily at a very early stage, during the onset of alpha mRNA transcription.

Cell Line↗
PubMed1987

Characterization of endonuclease activities in Moloney murine leukemia virus and its replication-defective mutants.

To study Moloney murine leukemia virus (M-MulV) proteins associated with the integration of proviral DNA into the host chromosome, we isolated endonuclease activities from purified virion preparations of the wild type and two of its replication mutants. A major endonuclease activity was identified in virions of M-MuLV; the enzyme catalyzed nicks in double-stranded DNA in the presence of either Mn2+ or Mg2+ and was stimulated by ATP. The endonuclease nicked DNA adjacent to all four nucleotides with some preference for G and C. The same enzyme, and in comparable amounts, was isolated from two virus replication mutants: dl2905, deficient in the processing of Pr65gag and Pr200gag-pol, and dl50401, deficient for the virus integration function. In the process of these experiments, the residual reverse transcriptase in mutant dl2905 was shown to be the mature size, implying that the uncleaved precursor lacks enzymatic activity. It appears that the major endonuclease activity found in virions of M-MuLV is not encoded by either the gag or pol genes.

Defective Viruses↗
PubMed1986

Use of reconstituted Sendai virus envelopes for fusion-mediated microinjection of double-stranded RNA: inhibition of protein synthesis in interferon-treated cells.

Poly(I).poly(C) molecules were trapped with reconstituted Sendai virus envelopes when added to the reconstitution system. A quantitative estimation indicated that about 10% of the added poly(I).poly(C) remained associated with the fusogenic viral envelopes. About 50% of the associated poly(I).poly(C) were found to be RNAase A resistant, enclosed within the viral envelopes. Incubation of loaded viral envelopes with HeLa or L-cells resulted in strong inhibition of protein synthesis, indicating fusion-mediated microinjection of the enclosed poly(I).poly(C). Introduction of poly(I).poly(C) into cultured cells by the use of reconstituted Sendai virus envelopes was as efficient as the introduction of these polynucleotides using the calcium phosphate coprecipitation technique. The inhibition of protein synthesis in L-cells but not in HeLa cells was dependent upon pretreatment with interferon. Incubation of poly(I).poly(C)-loaded viral envelopes with interferon-treated variant cells of the NIH 3T3 line, which possess a very low amount of RNAase L, resulted in only 25% inhibition of protein synthesis, compared to 85% inhibition observed in L-cells.

Animals↗
PubMed1986

Synthesis of herpes simplex virus proteins and nucleic acids in interferon-treated macrophages.

Mouse macrophages grown from spleen cells were found to be very sensitive to the interferon (IFN) activity against herpes simplex virus type 1 (HSV-1). Therefore we have used these cells to investigate the level at which IFN blocks the replication of HSV-1. IFN treatment resulted in a strong inhibition of the induction of HSV DNA polymerase and other beta proteins. RNA hybridization experiments revealed that the amount of mRNA for the beta protein thymidine kinase was strongly reduced in IFN treated HSV-1 infected cells. Analysis of the effect of IFN on expression of the alpha genes indicated a strong inhibition of alpha protein synthesis. In contrast the synthesis of mRNA of the alpha protein ICP 4 was only moderately inhibited. The results indicate that IFN primarily acts on the translation of HSV alpha proteins.

Animals↗
PubMed1986

Glutamine starvation of murine leukaemia virus-infected cells inhibits the readthrough of the gag-pol genes and proteolytic processing of the gag polyprotein.

The production of Moloney murine leukaemia virus from chronically infected cells was inhibited after starvation of glutamine. While the rate of synthesis of the precursor of the core proteins, Pr65gag, was not affected in the starved cells, its proteolytic processing was blocked. Pulse-chase experiments indicated that glutamine was required during the synthesis of Pr65gag to facilitate its subsequent processing. In addition, the synthesis of Pr200gag-pol, the precursor of the protease, reverse transcriptase and endonuclease, was inhibited in the glutamine-starved cells. Starvation for other essential amino acids such as tyrosine and isoleucine affected neither the synthesis nor the processing of the virus proteins. These results suggest that the readthrough mechanism which enables synthesis of the Pr200gag-pol polyprotein is modulated in the chronically infected cells by glutamine levels. Since the viral protease is part of the pol gene, its synthesis may be inhibited in the glutamine-starved cells and Pr65gag is therefore not processed.

Animals↗
PubMed1985

Control of K+ influx in 3T3 cells transformed by a conditional mutant of Rous sarcoma virus.

Mouse 3T3 cells transformed by a conditional mutant of Rous sarcoma virus (LA90) can assume either a normal or a transformed phenotype, depending on the temperature of cultivation. These cells (LA90) were arrested at the G0/G1 phase of the cell cycle by starvation for serum growth factors at the nonpermissive temperature (39 degrees C). Release from the G0/G1 phase by serum growth factors resulted in a rapid stimulation of Rb+ influx. To investigate whether the stimulation of Rb+ influx is obligatory for cell proliferation, the cultures were released from the G0/G1 phase by a temperature decrease in the absence of serum. A temperature decrease from 39 to 32 degrees C activated the viral pp60src gene mitogenic activity. Under these conditions, no rapid stimulation of Rb+ influx was observed. These results suggest that the rapid stimulation of Rb+ influx induced by serum growth factors is not an essential signal for cell release from the G0/G1 phase. However, a delayed increase in Rb+ influx concomitant with an increase in the cell content of K+ was observed in the cultures released from the G0/G1 phase by temperature decrease in the absence of serum growth factors. We found that the LA90 cells incubated at the permissive temperature (32 degrees C) secreted a mitogenic activity into the medium. Moreover, the conditioned medium from cultures incubated at 32 degrees C, but not at 39 degrees C, stimulate Rb+ influx in G0/G1 cells. These results indicate that Rous sarcoma virus pp60src induces a slow autocrine secretion of a mitogenic activity. This mitogenic activity slowly modulates the K+ content. Therefore, the slow elevation in cellular content of K+ is proposed to be an obligatory event for proliferation in normal and transformed cells.

Animals↗
PubMed1985

Independent regulation of ppp(A2'p)nA-dependent RNase in NIH 3T3, clone 1 cells by growth arrest and interferon treatment.

The regulation of ppp(A2'p)nA-(2-5A)-dependent RNase (RNase L or RNase F) was investigated in NIH 3T3, clone 1 cells using 2-5A-binding and nuclease activity assays. Minimal levels of 2-5A-dependent RNase were detected in actively dividing clone 1 cells; these levels were independently induced by growth arrest or interferon treatment. Accordingly, levels of the RNase were enhanced during growth arrest by confluency regardless of the presence or absence of interferon or antibody to interferon in the media. Measurement of 2-5A-dependent RNase was unaffected by the addition of any of six different proteinase inhibitors to the cells prior to extraction. The expression of 2-5A-dependent RNase in growth-arrested, interferon-treated cells was still relatively low (about one-third to one-half of that found in similarly treated murine Ehrlich ascites tumor cells). Although this amount of 2-5A-dependent RNase could not be detected by 2-5A-mediated ribosomal RNA cleavage, the activity was identified using a more sensitive novel assay for 2-5A-dependent RNase. In addition, introduction of 2-5A or poly(I) X poly(C) into growth-arrested, interferon-treated cells resulted in some inhibition of protein synthesis. The results indicated that the expression of 2-5A-dependent RNase in NIH 3T3, clone 1 cells is regulated under different physiological conditions and that low levels of 2-5A-dependent RNase were insufficient to significantly inhibit encephalomyocarditis virus replication.

Adenine Nucleotides↗
PubMed1985

Production and characterization of interferon from endothelial cells.

The capacity of cultured bovine aortic, capillary, and corneal endothelial cells as well as of human umbilical cord endothelial cells to produce interferon (IFN) was investigated. The endothelial cells of the two species produced significant amounts of IFN in response to various viruses and poly (I) poly (C). The IFN produced by human umbilical cord endothelial cells was a mixture of alpha- and beta-IFN, as determined by neutralization with antibodies directed against these two types of IFNs as well as by measuring the antiviral activity on heterologous cells. Bovine endothelial cells also produced a mixture of at least two IFN subspecies, one of them labile at pH 2 and active on human cells and the other stable at pH 2 and inactive on human cells.

Animals↗
PubMed1985

The antiproliferative effect of interferon and the mitogenic activity of growth factors are independent cell cycle events. Studies with vascular smooth muscle cells and endothelial cells.

We studied the antagonistic effects of interferon (IFN) and growth factors in G0/G1-arrested normal bovine aortic smooth muscle cells (SMC) which were stimulated by serum, or purified platelet derived growth factor (PDGF), supplemented with plasma-derived serum (PDS). The growth response, measured as [3H]thymidine incorporation into DNA, was dependent on the concentration of the mitogen. Human IFN alpha, recombinant human IFN alpha 2, or a crude bovine-IFN preparation prepared from virus-infected bovine aortic endothelial cells, inhibited SMC growth induced by either serum or PDGF with PDS. The extent of IFN inhibition was inversely related to the concentration of the mitogenic stimulus. We also investigated whether IFN inhibited the early events in G1 phase, stimulated by the competence factor PDGF, or the progression of the cell into the S phase induced by PDS. The results indicated that IFN inhibited these two stages of the G1 phase independently. In addition, we investigated the antiproliferative effect of IFN on bovine aortic endothelial cells (BAEC), which do not respond to PDGF but to the mitogenic activity of fibroblast growth factor (FGF). IFN inhibited the mitogenic activity of FGF in a dose-dependent manner. The results indicate that the anti-proliferative activity of IFN and the mitogenic effects of different growth factors are independent.

Animals↗
PubMed1985

Tubular structures in S49 mouse lymphoma are regulated through in vivo host-cell interaction and in vitro interferon treatment.

Malignant S49 mouse lymphoma cells that grow in suspension culture demonstrate in their cytoplasm characteristic tubular structures. These structures also appear in immunogenic, substrate-adherent variants of S49 cells that grow in culture. Upon transfer of both cell types into nude mice, the tubular structures of the adherent variants (and not the suspension-growing cells) undergo a profound alteration whereby their tubular components disappear and clusters of viruslike particles appear. These very closely resemble, on morphological grounds, precursors of B-type retroviruses. This specific in vivo interaction between the host and the S49 variant can be mimicked in culture by treatment of these cells for 24 h with 500 U/ml of mouse interferon. The suspension-growing S49 cells are unresponsive to interferon in this respect. Immunohistochemical analysis reveals that both tubular structures and the viruslike particles represent stages in the morphogenesis of mouse mammary tumor virus. A working hypothesis is advanced relating the regulation of the tubular system to the impaired tumorigenic potential of adherent S49 cells in syngeneic Balb/c hosts.

Animals↗
PubMed1985

Interferon inhibits prostaglandin E2 synthesis and stimulates (2'-5')oligoadenylate synthetase activity in peripheral blood mononuclear cells of inflammatory bowel disease patients.

The effect of interferon (IFN) on prostaglandin E2 (PGE2) synthesis by peripheral blood mononuclear cells of normal subjects and patients with inflammatory bowel disease was studied. Exposure of peripheral blood mononuclear cells, isolated from normal subjects and patients with active ulcerative colitis and Crohn's disease, to IFN reduced the release of PGE2 in a dose-dependent manner. The inhibitory effect was observed for at least 24 h after the removal of IFN. To correlate the effect of IFN on PGE2 synthesis with a known activity of IFN, the induction of (2'-5')oligoadenylate synthetase (OAS) was also studied in the same mononuclear cells. Low basal levels of this enzyme were detected in mononuclear cells isolated from normal subjects and patients with active ulcerative colitis and Crohn's disease. Moreover, IFN treatment in vitro resulted in a similar induction of this enzyme in patients and controls. There results indicate that peripheral blood mononuclear cells of patients with active ulcerative colitis and Crohn's disease have normal response to IFN. In addition, our data do not suggest that IFN has an important role in the pathogenesis of these diseases.

2',5'-Oligoadenylate Synthetase↗
PubMed1985

Inhibition of replication of herpes simplex virus in mouse macrophages by interferons.

The replication of herpes simplex virus (HSV) type 1 in macrophages grown from spleen cells of mouse strains susceptible to HSV infection in vivo was very sensitive to interferon (IFN). Different types of mouse IFN (alpha, beta, gamma) exhibited similar antiviral activities. However, treatment of cells with IFN-gamma in combination with IFN-alpha or IFN-beta resulted in a synergistic inhibition of virus growth. As shown by assaying HSV DNA polymerase, IFN inhibited expression of the beta-genes. Inhibition of enzyme induction correlated well with the reduction of viral yield. Induction of HSV DNA polymerase was delayed by IFN in a dose-dependent manner. These results show that IFN inhibits HSV replication at an early step prior to or during the synthesis of beta-proteins.

Animals↗
PubMed1985

Generation of a uniform 3' end RNA of murine leukemia virus.

Using the S1 nuclease mapping technique, we demonstrated that the majority of Moloney murine leukemia RNA molecules, isolated either from the nucleus or cytoplasm of infected mouse cells, share a uniform 3' end located at the border of the R and U-5 regions of the long terminal repeat. When the long terminal repeat sequences were inserted in the pSV plasmid downstream of the simian virus 40 late promoter, the 3' end of the viral RNA was also generated close to the R region of the long terminal repeat. These results demonstrate that the long terminal repeat signals the generation of an authentic 3' end when situated downstream of an actively transcribed region.

Animals↗
PubMed1985

Regulation of Moloney murine leukemia virus replication in chronically infected cells arrested at the G0/G1 phase.

The replication of Moloney murine leukemia virus (MMuLV) in chronically infected mouse cells arrested at the G0/G1 phase of the cell cycle by different procedures was investigated. MMuLV production was inhibited in glutamine- and isoleucine (Gln-Ile)-deprived G0/G1 cells. In contrast, butyric acid treatment, which efficiently arrested the cells at the G0/G1 phase of the cell cycle, did not inhibit MMuLV production. Furthermore, the inhibition of MMuLV production caused by either Gln-Ile deprivation or by interferon (IFN) treatment was overcome by butyric acid treatment. Thus, the replication of MMuLV could be dissociated from cell proliferation. The inhibition of MMuLV production in Gln-Ile-deprived cell cultures was compared to the inhibitory effect of IFN, which is known to affect budding and release of the virus. Rates of MMuLV protein synthesis were not affected in both the IFN-treated and Gln-Ile-deprived cells. However, processing of the viral polyprotein Pre65gag into p30 was blocked in the Gln-Ile-deprived cells. Furthermore, whereas in IFN-treated cells, MMuLV accumulated on the cell surface and could be released upon treatment with trypsin, in Gln-Ile-deprived cells, no virions were released by such treatment. These results indicate that in cells arrested by Gln-Ile deprivation, MMuLV is inhibited at a posttranslation step. This step appears to precede the anti-MMuLV block induced by IFN.

Animals↗