Effect of interferon on the penetration of murine leukemia virus and the binding of its genome RNA to polyribosomes at the early stage of infection

ABSTRACT The limited efficiency of in vivo gene transfer by replication-deficient retroviral vectors remains an obstacle to achieving effective gene therapy for solid tumors. One approach to circumvent this problem is the use of replication-competent retroviral vectors. However, the application of such vectors is at a comparatively early stage and the effects which virus strain, transgene cassette position, and target cell can exert on vector spread kinetics, genomic stability, and transgene expression levels remain to be fully elucidated. Thus, in this study a panel of vectors allowing the investigation of different design features on an otherwise genetically identical background were analyzed with respect to these readout parameters in cultures of both murine and human cells and in preformed tumors in nude mice. The obtained data revealed that (i) Moloney murine leukemia virus (Mo-MLV)-based vectors spread with faster kinetics, drive higher levels of transgene expression, and are more stable than equivalent Akv-MLV-based vectors; (ii) vectors containing the transgene cassette directly downstream of the envelope gene are genomically more stable than those containing it within the 3′-long terminal repeat U3 region; and (iii) the genomic stability of both strains seems to be cell line dependent.

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Packaging of Host mY RNAs by Murine Leukemia Virus May Occur Early in Y RNA Biogenesis

Journal of Virology ◽

10.1128/jvi.01219-09 ◽

2009 ◽

Vol 83 (23) ◽

pp. 12526-12534 ◽

Cited By ~ 33

Author(s):

Eric L. Garcia ◽

Adewunmi Onafuwa-Nuga ◽

Soyeong Sim ◽

Steven R. King ◽

Sandra L. Wolin ◽

...

Keyword(s):

Quality Control ◽

Murine Leukemia Virus ◽

Noncoding Rna ◽

Early Stage ◽

Leukemia Virus ◽

Rna Polymerase Iii ◽

Wild Type ◽

Rna Biogenesis ◽

Retroviral Assembly ◽

Murine Leukemia

ABSTRACT Moloney murine leukemia virus (MLV) selectively encapsidates host mY1 and mY3 RNAs. These noncoding RNA polymerase III transcripts are normally complexed with the Ro60 and La proteins, which are autoantigens associated with rheumatic disease that function in RNA biogenesis and quality control. Here, MLV replication and mY RNA packaging were analyzed using Ro60 knockout embryonic fibroblasts, which contain only ∼3% as much mY RNA as wild-type cells. Virus spread at the same rate in wild-type and Ro knockout cells. Surprisingly, MLV virions shed by Ro60 knockout cells continued to package high levels of mY1 and mY3 (about two copies of each) like those from wild-type cells, even though mY RNAs were barely detectable within producer cells. As a result, for MLV produced in Ro60 knockout cells, encapsidation selectivity from among all cell RNAs was even higher for mY RNAs than for the viral genome. Whereas mY RNAs are largely cytoplasmic in wild-type cells, fractionation of knockout cells revealed that the residual mY RNAs were relatively abundant in nuclei, likely reflecting the fact that most mY RNAs were degraded shortly after transcription in the absence of Ro60. Together, these data suggest that these small, labile host RNAs may be recruited at a very early stage of their biogenesis and may indicate an intersection of retroviral assembly and RNA quality control pathways.

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Recent Studies on a Murine Leukemia Virus Grown in Tissue Culture

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100060544 ◽

1967 ◽

Vol 25 ◽

pp. 106-107

Author(s):

L. Z. de Tkaczevski ◽

E. de Harven ◽

C. Friend

Keyword(s):

Tissue Culture ◽

Solid Tumors ◽

Murine Leukemia Virus ◽

Leukemia Virus ◽

Supernatant Fluid ◽

Virus Particles ◽

Friend Leukemia ◽

Type C ◽

Leukemia Viruses ◽

Murine Leukemia

Despite extensive studies, the correlation between the morphology and pathogenicity of murine leukemia viruses (MLV) has not yet been clarified. The virus particles found in the plasma of leukemic mice belong to 2 distinct groups, 1 or 2% of them being enveloped A particles and the vast majority being of type C. It is generally believed that these 2 types of particles represent different phases in the development of the same virus. Particles of type A have been thought to be an earlier form of type C particles. One of the tissue culture lines established from Friend leukemia solid tumors has provided the material for the present study. The supernatant fluid of the line designated C-1A contains an almost pure population of A particles as illustrated in Figure 1. The ratio is, therefore, the reverse of what is unvariably observed in the plasma of leukemic mice where C particles predominate.

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Immunoferritin Labelling of Murine Leukemia Virus Antigens in thin Sections

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s1431927600002221 ◽

1980 ◽

Vol 38 ◽

pp. 508-509

Author(s):

Ray A. Weigand ◽

Gregory C. Varjabedian

Keyword(s):

Murine Leukemia Virus ◽

Leukemia Virus ◽

Intracellular Localization ◽

Uranyl Acetate ◽

Antibody Technique ◽

Thin Sections ◽

Tumor Virus ◽

Labelling Procedure ◽

Unlabelled Antibody ◽

Murine Leukemia

We previously described the intracellular localization of murine mammary tumor virus (MuMTV) p28 protein in thin sections (1). In that study, MuMTV containing cells fixed in 3% paraformaldehyde plus 0.05% glutaraldehyde were labelled after thin sectioning using ferritin-antiferritin in an unlabelled antibody technique. We now describe the labelling of murine leukemia virus (MuLV) particles using the unlabelled antibody technique coupled to ferritin-Fab antiferritin. Cultures of R-MuLV in NIH/3T3 cells were grown to 90% confluence (2), fixed with 2% paraformaldehyde plus 0.5% glutaraldehyde in 0.1 M cacodylate at pH 7.2, postfixed with buffered 17 OsO4, dehydrated with a series of etha-nols, and embedded in Epon. Thin sections were collected on nickel grids, incubated in 107 H2O2, rinsed in HEPES buffered saline, and subjected to the immunoferritin labelling procedure. The procedure included preincubation in 27 egg albumin, a four hour incubation in goat antisera against purified gp69/71 of MuLV (3) (primary antibody), incubation in F(ab’)2 fragments of rabbit antisera to goat IgG (secondary antibody), incubation in apoferritin, incubation in ferritin-Fab ferritin, and a brief fixation with 2% glutaraldehyde. The sections were stained with uranyl acetate and examined in a Siemens IA electron microscope at an accelerating voltage of 60 KV.

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