BACKGROUND: HIV-1 must package its genome to gen- erate infectious particles. We have shown that HIV-1 encap- sidates its genome with high ef ciency and most virions contain two viral RNAs. Hence, HIV-1 genome packaging is a highly regulated process; however, the mechanism that achieves this regulation is currently unknown. METHODS: The viral components of individual particles were determined by uorescence microscopy (single-virion analysis). Viral genomes were detected by uorescently tagged RNA-binding proteins recognizing sequences engi- neered into the viral genomes. The coassembly of Gag pro- teins was detected by tagging Gag from each virus with a different uorescent protein. RESULTS: To determine how HIV-1 regulates genome encapsidation, we examined viral RNAs much larger (≈17 kb) or much smaller (≈3 kb) than that of wild-type HIV-1 (≈9 kb) and found that two copies of the RNA were incorporated into each virion independent of their sizes. However, it is possible for the virus to package just one copy of the HIV-1 RNA if the genome contains a second packaging signal that allows for the formation of intramolecular dimers (self-dimers). These results indi- cate that HIV-1 genome encapsidation is regulated by the packaging of one dimer. We hypothesized that the recognition of the RNA dimer by Gag nucleates virus assembly, and the dynamics of assembly prevent the incorporation of additional RNA genomes. To test the ability of packaging signals to nucle- ate virus assembly, we generated an HIV-1 genome, termed HMPsi, that harbours a second packaging signal from murine leukaemia virus (MLV). The HMpsi RNA can be packaged by either HIV-1 or MLV Gag. Impor- tantly, the presence of the HMpsi RNA promoted the coassembly of HIV-1 Gag and MLV Gag, indicating that the two packaging signals are each nucleating assembly with their corresponding Gags. CONCLUSIONS: Taken together, our data support a model of HIV-1 assembly in which Gag recognizes dimeric RNA and this recognition serves to nucleate Gag polym- erization and particle assembly. This mechanism ensures that one RNA dimer, but not more, is packaged into each nascent virion. As an absolute requirement for infectivity, and a driver of virion assembly, the viral genome should not be overlooked as a potential target for therapeutics.
|Title of host publication||Antiviral Therapy|
|Publication status||Published - 2013|