Gag-Pol supplied in trans is efficiently packaged and supports viral function in human immunodeficiency virus type 1

The intracellular trafficking and subsequent incorporation of Gag-Pol into human immunodeficiency virus type 1 (HIV-1) remains poorly defined. Gag-Pol is encoded by the same mRNA as Gag and is generated by ribosomal frameshif ting. The multimerization of Gag and Gag-Pol is an essential step in the fo rmation of infectious viral particles. In this study, we examined whether t he interaction between Gag and Gag-Pol is initiated during protein translat ion in order to facilitate the trafficking and subsequent packaging of Gag- Pol into the virion. A conditional cotransfection system was developed in w hich virion formation required the coexpression of two HIV-1-based plasmids , one that produces both Gag and Gag-Pol and one that only produces Gag-Pol . The Gag-Pol proteins were either immunotagged with a His epitope or funct ionally tagged with a mutation (K65R) in reverse transcriptase that is asso ciated with drug resistance. Gag-Pol packaging was assessed to determine wh ether the Gag-Pol incorporated into the virion was preferentially packaged from the plasmid that expressed both Gag and Gag-Pol or whether it could be packaged from either plasmid. Our data show that translation of Gag and Ga g-Pol from the same mRNA is not critical for virion packaging of the Gag-Po l polyprotein or for viral function.