Multimerization of human immunodeficiency virus type 1 gag promotes its localization to barges, raft-like membrane microdomains

The Gag polyprotein of human immunodeficiency virus type 1 (HIV-1) organize s the assembly of nascent virions at the plasma membrane of infected cells. Here we demonstrate that a population of Gag is present in distinct raft-l ike membrane microdomains that we have termed "barges." Barges have a highe r density than standard rafts, most likely due to the presence of oligomeri c Gag-Gag assembly complexes. The regions of the Gag protein responsible fo r barge targeting were mapped by examining the flotation behavior of wild-t ype and mutant proteins on Optiprep density gradients. N-myristoylation of Gag was necessary for association with barges. Removal of the NC and p6 dom ains shifted much of the Gag from barges into typical raft fractions. These data are consistent with a model in which multimerization of myristoylated Gag proteins drives association of Gag oligomers into raft-like barges. Th e functional significance of barge association was revealed by several line s of evidence. First, Gag isolated from virus-like particles was almost ent irely localized in barges. Moreover, a comparison of wild-type Gag with Fyn (10)Gag, a chimeric protein containing the N-terminal sequence of Fyn, reve aled that Fyn(10)Gag exhibited increased affinity for barges and a two- to fourfold increase in particle production. These results imply that associat ion of Gag with raft-like barge membrane microdomains plays an important ro le in the HIV-1 assembly process.