The production of new HIV-1 particles is initiated at the plasma membrane where the virus polyprotein Gag assembles into a budding site, and proceeds through release of an immature virion which is subsequently transformed to the infectious virion by proteolytic cleavage of Gag. Recently HIV-1 budding sites have been studied by cryo electron tomography. This technique allows three-dimensional structure determination of single objects at macromolecular resolution, and is uniquely suited to study variable structures such as HIV-1 particles and budding sites.
Using cryo electron tomography, researchers obtained three-dimensional images with unprecedented detail of the formation of HIV-1 particles. By analyzing these images they showed that the organization of released immature HIV-1 is determined at its intracellular assembly without major subsequent rearrangements. They were able to identify a lattice structure of the viral protein Gag present in budding sites that seem to lack the viral genome and thus cannot be precursors of infectious viruses. Some HIV-1 infected T-cells preferentially carry these budding sites, suggesting that they have lost a crucial control of the proteolytic maturation of the virus.
Cryo Electron Tomography of Native HIV-1 Budding Sites. (2010) PLoS Pathog 6(11): e1001173. doi:10.1371/journal.ppat.1001173
The structure of immature and mature HIV-1 particles has been analyzed in detail by cryo electron microscopy, while no such studies have been reported for cellular HIV-1 budding sites. Here, we established a system for studying HIV-1 virus-like particle assembly and release by cryo electron tomography of intact human cells. The lattice of the structural Gag protein in budding sites was indistinguishable from that of the released immature virion, suggesting that its organization is determined at the assembly site without major subsequent rearrangements. Besides the immature lattice, a previously not described Gag lattice was detected in some budding sites and released particles; this lattice was found at high frequencies in a subset of infected T-cells. It displays the same hexagonal symmetry and spacing in the MA-CA layer as the immature lattice, but lacks density corresponding to NC-RNA-p6. Buds and released particles carrying this lattice consistently lacked the viral ribonucleoprotein complex, suggesting that they correspond to aberrant products due to premature proteolytic activation. We hypothesize that cellular and/or viral factors normally control the onset of proteolytic maturation during assembly and release, and that this control has been lost in a subset of infected T-cells leading to formation of aberrant particles.