A replication-defective variant of Moloney murine leukemia virus. I. Biological characterization

Abstract

We have studied the virus produced by a clone, termed 8A, that was isolated from a culture of murine sarcoma virus-transformed mouse cells after superinfection with Moloney murine leukemia virus (MuLV-M). Clone 8A produced high levels of type C virus particles, but only a low titer of infectious murine sarcoma virus and almost no infectious MuLV. When fresh cultures of mouse cells were infected with undiluted clone 8A culture fluids, they released no detectable progeny virus for several weeks after infection. Fully infectious MuLV was then produced in these cultures. This virus was indistinguishable from MuLV-M by nucleic acid hybridization tests and in its insensitivity to Fv-1 restriction. It also induced thymic lymphomas in BALB/c mice. To explain these results, we propose that clone 8A is infected with a replication-defective variant of MuLV-M. Particles produced by clone 8A, containing this defective genome, can establish an infection in fresh cells but cannot produce progeny virus at detectable levels. Several weeks after infection, the defect in the viral genome is corrected by back-mutation or by recombination with endogenous viral genomes, resulting in the formation of fully infectious progeny MuLV. The progeny MuLV's that arose in two different experiments were found to be genetically different from each other. This is consistent with the hypothesis that, in each experiment, the progeny virus is formed as a result of an independent genetic event. DNA was isolated from clone 8A cells and assayed for infectivity by the calcium phosphate transfection technique. No detectable MuLV was produced by cells treated with this DNA. This finding, along with positive results obtained in control experiments, indicates that clone 8A cells do not contain a normal MuLV provirus.

Mutant of B-tropic murine leukemia virus synthesizing an altered polymerase molecule

“…Cells and virus stocks. All cells used in these studies have been described previously (33)(34)(35). Cells were grown in McCoy 5a medium with 10% heatinactivated fetal calf serum, penicillin, and streptomycin.…”

“…Nonconditional mutants of mammalian RNA tumor viruses will provide valuable tools for the analysis of steps in viral infection and for the study of cell-virus interactions such as Fv-1 restriction (1) which are unique to mammalian cells. Reports from several laboratories indicate that the generation of replication-defective variants of mammalian RNA tumor viruses occurs with high frequency (16,35,38,39). To date, the number of extensively characterized mutants is quite small.…”

Requirements for different components of the host cell cytoskeleton distinguish ecotropic murine leukemia virus entry via endocytosis from entry via surface fusion

“…In newly infected mouse cells, ecotropic virus particles were seen both inside endocytic vesicles (40) and at the surface undergoing uncoating (31), although it is not clear if one or both observations represent productive virus infection because the particle to infectious unit ratio is very high (38,39). Additional disparate results revealing the complexity of ecotropic virus penetration emerged from studies of the effects of pH on ecotropic virus entry.…”

Cell fusion for genetic analysis of two nonconditional Rous sarcoma virus replication mutants

“…We have also been able to demonstrate complementation by fusing pairs of permissive cells transformed by nonconditional replication-defective viruses (Steimer and Boettiger, manuscript in preparation). This approach might also be used to analyze other retrovirus mutants such as the recently isolated Moloney murine leukemia virus replication defectives (27,33).…”