The Nonmyristylated Pr1609a9P'1 Polyprotein of Human Immunodeficiency Virus Type 1 Interacts with pr55ga5 and Is Incorporated into Viruslike Particles

The expression of the pol gene of human immunodeficiency virus type 1 occurs via a ribosomal frameshift between the gag and pol genes. The resulting protein, a Gag-Pol polyprotein, is produced at a level 5 to 10%S of that of the Gag protein. The Gag-Pol polyprotein is incorporated into virions and provides viral protease, reverse transcriptase, and integrase, which are essential for infectivity. It is generally believed that the Gag-Pol polyprotein is incorporated into virions via interaction with the Gag protein, although the details of the mechanism are unknown. To further study this problem, we have constructed a human immunodeficiency virus type 1 proviral genome which overexpresses the Gag-Pol polyprotein (Pr1609`u°ol). Transfection of this proviral genome (pGPpr-) into COS-1 cells resulted in the expression of full-length Pr1606"'"9 polyprotein. Although the majority of the Pr160"" ' was confined to the cells, low levels of reverse transcriptase activity were detectable in the cell supernatants. The cotransfection of pGPpr- with a second plasmid which expresses only the Pr558"w precursor (pGAG) resulted in a significantly higher level of Pr1609'" " in the medium of transfected cells. Sedimentation analysis using sucrose density gradients demonstrated that most PrI60'9"P' was found in fractions corresponding to the density of virion particles, indicating that the Pr160"g'-Po' polyprotein was released in association with a Pr55mt viruslike particle. To further characterize the requirements for the release, a mutation was constructed to express an unmyristylated Pr1609"°" ' polyprotein