Assembly and composition of intracellular particles formed by Moloney murine leukemia virus

Assembly of type C retroviruses such as Moloney murine leukemia virus (M-MuLV) ordinarily occurs at the plasma membranes of infected cells and absolutely requires the particle core precursor protein, Pr65^gag. Previously we have shown that Pr65^gag is membrane associated and that at least a portion of intracellular Pr65^gag protein appears to be routed to the plasma membrane by a vesicular transport pathway. Here we show that intracellular particle formation can occur in M-MuLV-infected cells. M-MuLV immature particles were observed by electron microscopy budding into and within rough endoplasmic reticulum, Golgi, and vacuolar compartments. Biochemical fractionation studies indicated that intracellular Pr65^gag was present in nonionic detergent-resistant complexes of greater than 150S. Additionally, viral RNA and polymerase functions appeared to be associated with intracellular particles, as were Gag-β-galactosidase fusion proteins which have the capacity to be incorporated into virions. Immature intracellular particles in postnuclear lysates could be proteolytically processed in vitro to mature forms, while extracellular immature M-MuLV particles remained immature as long as 10 h during incubations. The occurrence of M-MuLV-derived intracellular particles demonstrates that Pr65^gag can associate with intracellular membranes and indicates that if a plasma membrane Pr65^gag receptor exists, it also can be found in other membrane compartments. These results support the hypothesis that intracellular particles may serve as a virus reservior during in vivo infections.