Effects of reduced gag cleavage efficiency on HIV-1 Gag-Pol package

BACKGROUND: HIV-1 pol, which encodes enzymes required for virus replication, is initially translated as a Gag-Pol fusion protein. Gag-Pol is incorporated into virions via interactions with Gag precursor Pr55(gag). Protease (PR) embedded in Gag-Pol mediates the proteolytic processing of both Pr55gag and Gag-Pol during or soon after virus particle release from cells. Since efficient Gag-Pol viral incorporation depends on interaction with Pr55(gag) via its N-terminal Gag domain, the prevention of premature Gag cleavage may alleviate Gag-Pol packaging deficiencies associated with cleavage enhancement from PR. RESULTS: We engineered PR cleavage-blocking Gag mutations with the potential to significantly reduce Gag processing efficiency. Such mutations may mitigate the negative effects of enhanced PR activation on virus assembly and Gag-Pol packaging due to an RT dimerization enhancer or leucine zipper dimerization motif. When co-expressed with Pr55(gag), we noted that enhanced PR activation resulted in reduced Gag-Pol cis or trans incorporation into Pr55(gag) particles, regardless of whether or not Gag cleavage sites within Gag-Pol were blocked. CONCLUSIONS: Our data suggest that the amount of HIV-1 Gag-Pol or Pol viral incorporation is largely dependent on virus particle production, and that cleavage blocking in the Gag-Pol N-terminal Gag domain does not exert significant impacts on Pol packaging. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12866-022-02503-3.