Independent segregation of human immunodeficiency virus type 1 Gag protein complexes and lipid rafts.
Formation of human immunodeficiency virus type 1 (HIV-1) particles takes place at the plasma membrane of cells and is directed by the Pr55Gag polyprotein. A functional assembly domain (the M domain) within the N-terminal portion of Pr55Gag mediates the interaction of Gag with cellular membranes. However, the determinants that provide specificity for assembly on the plasma membrane, as opposed to intracellular membranes, have not been identified. Recently, it was reported that Pr55Gag interacts with lipid raft microdomains of the plasma membrane. We sought to identify the domains within Pr55Gag that contribute to lipid raft association of Gag. Here we demonstrate that the I domain is required for interaction with detergent-resistant membrane fractions (DRMs). Mutation of key I-domain residues or loss of myristylation abrogated the association of Gag with DRMs. Thus, the I domain and the M domain combine to mediate Gag-lipid raft interactions as defined by these biochemical criteria. However, Gag protein complexes defined by flotation studies were much denser than classical lipid rafts, failed to incorporate classical lipid raft marker proteins, and were not disrupted by cholesterol extraction. Large sheets of Gag protein were identified in DRM fractions upon examination by electron microscopy. These results indicate that HIV-1 Pr55Gag forms detergent-resistant complexes at the cellular periphery that are distinct from lipid raft microdomains.
Ding
Lingmei
L
Departments of Pediatrics, Microbiology, and Immunology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-2581, USA.
Derdowski
Aaron
A
Wang
Jaang-Jiun
JJ
Spearman
Paul
P
D000595:Amino Acid Sequence; D015683:Gene Products, gag / Q000378:metabolism*; D015497:HIV-1 / Q000502:physiology*; D021962:Membrane Microdomains / Q000378:metabolism*; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D019814:Myristic Acid / Q000378:metabolism; D017830:Octoxynol / Q000494:pharmacology; D011498:Protein Precursors / Q000378:metabolism*
D015683:Gene Products, gag; D011498:Protein Precursors; C059767:p55 gag precursor protein, Human immunodeficiency virus 1; D019814:Myristic Acid; D017830:Octoxynol