HIV-1 Matrix Dependent Membrane Targeting Is Regulated by Gag mRNA Trafficking
Figure 6
PRE-dependent HIV-1 Gag failed to associate with lipid raft in human cells.
(A) Both Rev-dependent and PRE-dependent HIV-1 Gag are membrane-associated in human cells. Postnuclear supernatants derived from 293T cells expressing Rev-dependent or PRE-dependent HIV-1 Gag, Rev-dependent or PRE-dependent G2A mutant were subjected to equilibrium flotation centrifugation. Pr55Gag, TfR and actin were detected by Western blotting. Membrane- and non-membrane-associated fractions were shown. (B) Rev-dependent HIV-1 Gag but not PRE-dependent HIV-1 Gag is associated with Triton X-100 insoluble lipid rafts in human cells. Postnuclear supernatants derived from 293T cells expressing Rev-dependent or PRE-dependent HIV-1 Gag were treated with 0.5% Triton X-100 on ice for 30 min prior to membrane flotation analysis. Pr55Gag, TfR and actin were detected by Western blotting. Raft and non-raft fractions are indicated. (C) Quantitation of Rev-dependent (left panel) and PRE-dependent (right panel) HIV-1 Gag in each fraction with (open circles) or without (closed squares) cold Triton extraction.