Human immunodeficiency virus type 1 integrase protein promotes reverse transcription through specific interactions with the nucleoprotein reverse transcription complex

The human immunodeficiency virus type 1 (HIV-1) integrase protein (IN) is e ssential for integration of the viral DNA into host cell chromosomes. Since IN is expressed and assembled into virions as part of the 160-kDa Gag-Pol precursor polyprotein and catalyzes integration of the provirus in infected cells as a mature 32-kDa protein, mutations in IN are pleiotropic and may affect virus replication at different stages of the virus life cycle in add ition to integration. Several different phenotypes have been observed for I N mutant viruses, including defects in virion morphology, protein compositi on, reverse transcription, nuclear import, and integration. Because the eff ects of mutations in the IN domain of Gag-Pol can not always be distinguish ed from those of mutations in the mature IN protein, there remains a signif icant gap in our understanding of IN function in vivo. To directly analyze the function of the mature IN protein itself, in the context of a replicati ng virus but independently from that of Gag-Pol, we used an approach develo ped in our laboratory for incorporating proteins into HIV virions by their expression in trans as fusion partners of either Vpr or Vpx, By providing I N in trans as a Vpr-IN fusion protein, our analysis revealed, for the first time, that the mature IN protein is essential for the efficient initiation of reverse transcription in infected cells and that this function does not require the IN protein to be enzymatically (integration) active. Our findi ngs of a direct physical interaction between IN and reverse transcriptase a nd the failure of heterologous HIV-2 IN protein to efficiently support reve rse transcription indicate that this novel function occurs through specific interactions with other viral components of the reverse transcription init iation complex. Studies involving complementation between integration- and DNA synthesis-defective IN mutants further support this conclusion and reve al that the highly conserved HHCC motif of LN is important for both activit ies. These findings provide important new insights into IN function and rev erse transcription in the context of the nucleoprotein reverse transcriptio n complex within the infected cell. Moreover, they validate a novel approac h that obviates the need to mutate Gag-Pol in order to study the role of it s individual mature components at the virus replication level.