Type-C retrovirus maturation and assembly: Post-translational cleavage of the gag-gene coded precursor polypeptide occurs at the cell membrane

Temperature-sensitive (ts) replication mutants of the Rauscher (R-) strain of murine leukemia virus (MuLV) defective in post-translational processing of the 65,000 molecular weight gag-gene coded precursor (Pr65) have been used to study the initial stages in type-C virus assembly. In is mutant infected cells grown at the nonpermissive but not the permissive temperature, dense material was found below the cell membrane by electron microscopy. Within 30 sec following shiftdown of is mutant infected cells to the permissive temperature (31°) a decrease in the amount of this dense material was observed, coincident with the initiation of virus assembly. Virus assembly and the release of mature virions were completed within 2–4 min. Immunofluorescent studies using monospecific antibodies to several highly purified viral structural proteins revealed a high concentration of these proteins below the cell membrane which also diminished concurrently with virus release. From the evidence presented we propose that the submembranous dense material observed by electron microscopy represents a pool of the polypeptide precursor Pr65, which is cleaved and then incorporated into the developing virus within seconds following shiftdown of the cells to the permissive temperature.