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Figure 2 | Retrovirology

Figure 2

From: BI-2 destabilizes HIV-1 cores during infection and Prevents Binding of CPSF6 to the HIV-1 Capsid

Figure 2

BI-2 destabilizes the HIV-1 Core during infection. (A) Cf2Th cells were challenged with HIV-1-GFP viruses in the presence of BI-2, and used to perform the fate of the capsid assay 12 hours post-infection, as described [9],[11]. Briefly, HIV-1 infected cells were used to prepare post-nuclear supernatants that were layered onto a 50% sucrose cushion to separate soluble from pelletable HIV-1 capsids. INPUT, SOLUBLE and PELLET fractions were analyzed by Western blotting using antibodies against HIV-1 CA p24. As control, we studied the fate of the HIV-1 capsid in the presence of PF74. The percentages of pelletable capsids relative to the infected control in the presence of DMSO are shown. Similar results were obtained in three independent experiments and standard deviations are shown. (B) Cf2Th cells were challenged with HIV-1-GFP viruses bearing the capsid change T107N in the presence of BI-2, and used to perform the fate of the capsid assay 12 hours post-infection. As a control, we performed experiments in cells stably expressing rhesus TRIM5α.The percentages of pelletable capsids relative to the infected control in the presence of DMSO are shown. Similar results were obtained in three independent experiments and standard deviations are shown. (C) Stability of in vitro assembled HIV-1 CA-NC complexes in destabilization buffer containing increasing concentrations of BI-2 (upper panel) or PF74 (lower panel) were measured as described [2]. Input and Pellet fractions were analyzed by Western blotting using antibodies against HIV-1 CA p24. As control, stability of in vitro assembled HIV-1 CA-NC complexes in stabilization buffer was measured. Similar results were obtained in three independent experiments. (D) Stability of wild type (upper panel) or T107N mutant (lower panel) in vitro assembled HIV-1 CA-NC complexes in stabilization buffer containing cell extracts at increasing concentrations of BI-2 was measured, as described [2]. Similar results were obtained in three independent experiments.

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