Figure 3

Localization of Alix and CHMP4 at the viral assembly compartment. Monocyte-derived-macrophages infected with HIV-1 NLAD8 for 14 days were processed for cryosectioning as described [65]. (A) Two examples of virus-containing compartments that were triple labeled for p17/p55 Gag with protein A coupled to gold particles of 5 nm or PAG5, for Alix with PAG10, and for CD63 with PAG15. Alix labeling was found on the virions and at the limiting membrane of the viral assembly compartment (black arrowheads). Note the labeled mitochondria nearby (small arrow). (B) Cryosections were triple labeled for p17/p55 Gag with PAG5, for CHMP4B with PAG10, and for CD63 with PAG15. CHMP4B was present in many virions (black arrowheads). In panels (A) and (B), CD63 was at the limiting membrane of the compartment, in small internal vesicles or incorporated in the membrane of virus particles. (B') Two examples of viral compartments double labeled for CHMP4B with PAG10, and p17/p55 Gag with PAG15. CHMP4B was associated with a thick molecular coat present at the limiting membrane of the assembly compartments (black arrowheads). Bars, 100 nm.