Abstract
The HIV-1 Gag p6 protein regulates the final abscission step of nascent virions from the cell membrane by the action of its two late (l-) domains, which recruit Tsg101 and ALIX, components of the ESCRT system. Even though p6 consists of only 52 amino acids, it is encoded by one of the most polymorphic regions of the HIV-1 gag gene and undergoes various posttranslational modifications including sumoylation, ubiquitination, and phosphorylation. In addition, it mediates the incorporation of the HIV-1 accessory protein Vpr into budding virions. Despite its small size, p6 exhibits an unusually high charge density. In this study, we show that mutation of the conserved glutamic acids within p6 increases the membrane association of Pr55 Gag followed by enhanced polyubiquitination and MHC-I antigen presentation of Gag-derived epitopes, possibly due to prolonged exposure to membrane bound E3 ligases. The replication capacity of the total glutamic acid mutant E0A was almost completely impaired, which was accompanied by defective virus release that could not be rescued by ALIX overexpression. Altogether, our data indicate that the glutamic acids within p6 contribute to the late steps of viral replication and may contribute to the interaction of Gag with the plasma membrane.
Highlights
The HIV-1 Gag polyprotein Pr55 is necessary and sufficient for the generation of virus like particles (VLPs) [1,2]
Even though the coding region of p6 is highly polymorphic within the HIV-1 viral genome, it contains motifs, some of which are highly conserved and enable interaction with several cellular (ERK-2, aPKC, ALIX, Tsg101, SUMO-1, ubiquitin) and viral (Vpr) proteins [8,9,10,12,21,22,26,34,35,36,37]
We demonstrate that mutation of these Glu residues within p6 to Ala impairs Gag processing and virus release and enhances membrane association of Gag which is accompanied by increased polyubiquitination and entry of Gag into the MHC-I antigen presentation pathway
Summary
The HIV-1 Gag polyprotein Pr55 is necessary and sufficient for the generation of virus like particles (VLPs) [1,2]. The multifunctional p6 protein undergoes various post-translational modifications, including sumoylation at Lys-27 and mono-ubiquitination at Lys-27 and -33 [21,22,23] It is the predominant phosphoprotein of HIV-1 particles [24]. Even though the coding region of p6 is highly polymorphic within the HIV-1 viral genome, it contains motifs, some of which are highly conserved and enable interaction with several cellular (ERK-2, aPKC, ALIX, Tsg101, SUMO-1, ubiquitin) and viral (Vpr) proteins [8,9,10,12,21,22,26,34,35,36,37]. We demonstrate that mutation of these Glu residues within p6 to Ala impairs Gag processing and virus release and enhances membrane association of Gag which is accompanied by increased polyubiquitination and entry of Gag into the MHC-I antigen presentation pathway. Gag-membrane-interaction and to the late functions of p6 in virus budding
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