Recreating a critical step in HIV formation
Viruses are masters of using cells for their own purposes. We have synthetically recreated how HIV attracts so called ESCRT proteins - normally involved in cell division - to the sites where new virus particles are released from an infected cell.
HFSP Long-Term Fellow Lars-Anders Carlson and colleaguesauthored on Mon, 12 November 2012
How can viruses complete their intricate replication cycles with a very small number of genes? One answer is that they use cellular proteins for their own purposes. A prime example of this is the release of new virus particles from an infected cell. Most viruses that are released by budding through the cell membrane (as opposed to just breaking it) achieve this by hijacking the cellular ESCRT proteins (endosomal sorting complex required for transport). The ESCRT proteins are required for many membrane budding events in normal cell function, including cell division.
The initial interactions between HIV’s Gag protein and the ESCRT proteins have been well characterised, but the subsequent assembly of the entire ESCRT machinery at HIV release sites is less well understood. In this study, we have taken a novel approach to understand how HIV directs the assembly of the ESCRT machinery at sites of virus release. We have biochemically isolated the HIV-1 Gag protein and the individual ESCRT proteins, and used giant unilamellar vesicles as a synthetic model of the cell membrane to study how these proteins come together. Fluorescent labelling of the membrane, HIV-1 Gag, and the ESCRT proteins enabled us to delineate the interdependencies of these factors at HIV assembly sites. This defined the minimal protein set required for ESCRT assembly at HIV release sites, and put the role of the ESCRT-II complex in a new light.
Figure: Confocal fluorescence microscopy image of a giant unilamellar vesicle with bound HIV-1 Gag and ESCRT protein. The left panel shows how the Gag protein of HIV-1 (white) forms clusters on the surface of a giant unilamellar vesicle (red). The right panel shows how the ESCRT protein ALIX (green) is recruited to these membrane-bound Gag clusters. The scale bar in the left panel represents 10 micrometres.
In vitro reconstitution of the ordered assembly of the endosomal sorting complex required for transport at membrane-bound HIV-1 Gag clusters. Lars-Anders Carlson and James H Hurley. Proc Natl Acad Sci U S A 109: 16928-16933. (16 October 2012).