Figure S4.

Inside the nucleus HIV-1 capsids rupture and release their dense content, related to Figures 2, 3, 4, and 5

(A) Related to Figure S3. Summary of total number of cones, cone-tubes, tubes and fragments captured by CLEM-ET or cryo-ET in the cytosol, inside NPCs or within the nucleus (see also Figure S3). (B) Quantification of densities observed inside of capsids captured in resin sections of infected SupT1-R5 cells by CLEM-ET. Each capsid structure was segmented. The voxel intensity observed within their interior was quantified and normalized to the average voxel intensity measured in the respective surrounding. See STAR methods for detail. Median values are indicated by black lines. Only complete structures that were fully captured within the resin section were analyzed (see Figure S3). Statistical significance was calculated using an unpaired two-tailed t test. WT CA: n.s., not significant; A77V CA: ^∗∗p = 0.0032; WT CA upon CPSF6 knock-down: ^∗∗∗p = 0.0008. Cy, cytosol; Nu, nucleus. The difference between WT CA / Cy and WT CA / Nu is not significant as the data contains two very dense structures (red dots), likely because it was collected from cells at an early time post-infection (3 h). (C–L) SupT1-R5 cells were infected with IN.mScarlet carrying NNHIV-A77V CA mutant for 15 h at 37°C, prior to high pressure freezing (for CLEM) or plunge freezing (for cryo-ET). Slices through tomographic reconstructions highlighting the morphology of A77V capsids or capsid-related structures visualized by CLEM-ET (C–F) or cryo-ET (G–L). Shown are examples of NNHIV-A77V structures visualized in the cytosol (C, C’, G), during docking (H, I), NPC penetration (D) and after translocation through the nuclear pores (E, F, J–L). Capsids in panels (C, C’) and (G, K) were identified in the same electron tomograms. Black, white and green arrowheads indicate capsids (or capsid related structures), microtubule cross sections and NPCs, respectively. Cy, cytosol; Nu, nucleus; NE, nuclear envelope.

See also Figure S3.