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The core of the retrovirus Murine leukemia virus (MLV) consists of the Gag precursor protein and viral RNA. It assembles at the cytoplasmic face of the cell membrane where, by an unclear mechanism, it collects viral envelope proteins embedded in the cell membrane and buds off. The C-terminal half of the short cytoplasmic tail of the envelope transmembrane protein (TM) is cleaved off to yield R-peptide and fusion-active TM. In Moloney MLV particles, R-peptide was found to bind to core particles. In cells, R-peptide and low amounts of uncleaved TM were found to be associated with small core-like complexes, i.e. mild detergent-insoluble, Gag-containing complexes with a density of 1.23 g ml(-1) and a size of 150-200 S. Our results suggest that TM associates with the assembling core particle through the R-peptide before budding and that this is the mechanism by which the budding virus acquires the envelope proteins.

Ten mutations were generated in the env gene of Moloney murine leukaemia virus DNA. The mutations were made by site-directed mutagenesis to alter basic amino acids (lysine or arginine) in the surface glycoprotein gp70. Mutants were investigated following transfection into NIH/3T3 cells. All 10 mutants released virion particles into the medium, suggesting that none of the mutations affected overall viral gene expression or virion budding. Two mutants were positive in XC plaque assay, reverse transcriptase assay and re-infection experiments, showing that these mutations occurred in parts of the molecule not essential for infection. Three mutants were negative in both the XC plaque assay and re-infection experiments, suggesting that they make non-infectious virus particles. The results indicate a defect in the early phase of infection, perhaps in receptor binding or in the fusion of virion and host membranes. The other mutations resulted in reduced infectivity of released virion particles.

Protein-protein interaction of Moloney murine leukaemia virus was studied by an assay where one protein preparation was coupled covalently to Sepharose, and binding of radiolabelled proteins to the protein-Sepharose was examined. It was found that the virus proteins gp70, p30, p15E and p15 in solution could associate weakly to disrupted virus particles and to p30. However, when the disrupted virus particles and p30 were coupled to Sepharose in the presence of Triton X-100, stronger binding of the four proteins was observed. Only low or no binding of p12 and p10 was observed to these protein-Sepharoses. The results are discussed with respect to the assembly and structure of the virus particle.

The effect of proteases on retrovirus-induced cell fusion was studied. Both 'fusion from within' (fusion between infected and uninfected cells) and 'fusion from without' (fusion of uninfected cells by virus addition) was enhanced after treatment with proteases. An enhancement of up to 10-fold, giving 30% fusion of the cells, was observed. Protease treatment of infected cells or of virus particles enhanced fusion, indicating that cleavage of the virus surface proteins is important for cell fusion. Cell fusion is believed to reflect the virus-cell membrane fusion which occurs during the entry of enveloped viruses into cells. It is therefore possible that proteolytic cleavage of virus surface components during entry is involved in virus infection.

The surface protein gp70 of an ecotropic murine retrovirus was followed during entry of [3H]glucosamine-labelled virions into SC-1 mouse fibroblasts. Upon entry, gp70 was cleaved into fragments with molecular weights 35K, 30K and 17K. The 35K and 17K fragments were also observed after trypsin or thermolysin cleavage of the virion, indicating that certain locations on the gp70 molecule are easily accessible from the outside of the virion. The conformation of gp70 on the membrane was shown to have a major effect on the cleavage. This protein is known to be important for early interactions with the cell (binding and membrane fusion). The results indicate that gp70 cleavage may be important for membrane fusion.