Transmission electron microscopy reveals an optimal HIV-1 nucleocapsid aggregation with single-stranded nucleic acids and the mature HIV-1 nucleocapsid protein

HIV-1 nucleocapsid protein (NCp7) condenses the viral RNA within the mature capsid. In a capsid-free system, NCp7 promotes an efficient mechanism of aggregation with both RNA and DNA. Here, we show an analysis of these macromolecular complexes by dark-field imaging using transmission electron microscopy. Thousands of mature NCp7 proteins co-aggregate with hundreds of single-stranded circular DNA molecules (ssDNA) within minutes, as observed with poly(rA). These co-aggregates are highly stable but dynamic structures, as they dissociate under harsh conditions, and after addition of potent ssDNA or NCp7 competitive ligands. The N-terminal domain and zinc fingers of NCp7 are both required for efficient association. Addition of magnesium slightly. increases the avidity of NCp7 for ssDNA, while it strongly inhibits coaggregation with relaxed circular double-stranded DNA (dsDNA). This DNA selectivity is restricted to mature NCp7, compared to its precursors NCp15 and NCp9. Moreover, for NCp15, the linkage of NCp7 with the Gag C-terminal p6-peptide provokes a deficiency in ssDNA aggregation, but results in DNA spreading similar to prototypical SSB proteins. Finally, this co-aggregation is discussed in a dynamic architectural context with regard to the mature HIV-1 nucleocapsid. On the basis of the present data, we propose that condensation of encapsidated RNA requires the C-terminal processing of NCp. Subsequently, disassembly of the nucleocapsid should be favoured once dsDNA is produced by HIV-1 reverse transcriptase. Published by Elsevier Ltd.