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) organizes 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-like membrane microdomains that we have termed "barges." Barges have a higher density than standard rafts, most likely due to the presence of oligomeric Gag-Gag assembly complexes. The regions of the Gag protein responsible for barge targeting were mapped by examining the flotation behavior of wild-type and mutant proteins on Optiprep density gradients. N-myristoylation of Gag was necessary for association with barges. Removal of the NC and p6 domains 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. The functional significance of barge association was revealed by several lines of evidence. First, Gag isolated from virus-like particles was almost entirely localized in barges. Moreover, a comparison of wild-type Gag with Fyn(10)Gag, a chimeric protein containing the N-terminal sequence of Fyn, revealed that Fyn(10)Gag exhibited increased affinity for barges and a two- to fourfold increase in particle production. These results imply that association of Gag with raft-like barge membrane microdomains plays an important role in the HIV-1 assembly process.
Lindwasser
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OW
Cell Biology Program, Memorial Sloan-Kettering Cancer Center, and Graduate Program in Cell Biology and Genetics, Weill Graduate School of Medical Sciences of Cornell University, New York, New York 10021, USA.
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D000818:Animals; D001688:Biological Products / Q000378:metabolism; D019556:COS Cells; D002522:Chlorocebus aethiops; D015967:Gene Expression Regulation, Viral; D015683:Gene Products, gag / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015497:HIV-1 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D021962:Membrane Microdomains / Q000737:chemistry / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011498:Protein Precursors / Q000378:metabolism; D014162:Transfection; D019065:Virus Assembly
D001688:Biological Products; D015683:Gene Products, gag; D011498:Protein Precursors; C059767:p55 gag precursor protein, Human immunodeficiency virus 1; C105085:viron