HIV-1 Capsid Assembly

HIV-1 Electron Microscopy:

HIV-1 capsids assembling at the inner aspect of the plasma membrane of Cos-1 cells. Transmission electron micrograph produced by Julia Dooher and Bobbie Schneider at the FHCRC EM facility.

HIV-1 particles containing assembled capsids budding and releasing from the plasma membrane. Transmission electron micrograph produced by Julia Dooher and Bobbie Schneider at the FHCRC EM facility.

Gag and HP68 are Co-localized:

Wild-type Gag, but not an assembly-defective Gag mutant, recruits the cellular protein ABCE1 into sites of assembly:

Cos-1 cells were transfected with a plasmid encoding the full-length, wild-type HIV-1 pBru genome with a deletion in Env (first two columns, a - f), or a pBru plasmid encoding a truncated form of HIV-1 Gag termed p41 that fails to associate with ABCE1 or assemble into capsids (third column, g - I). Immunostaining was performed for HIV-1 Gag (green; Cy-2) and endogenous ABCE1 (red; Cy-3). Each column shows one field of cells examined by immunofluorescence microscopy using filters that allow visualization of either ABCE1 (a, d, g) or Gag (b, e, h) or both (c, f, I). The first column shows two cells on the left that are not expressing Gag (b). In these cells, endogenous cellular ABCE1 is present diffusely (a). In contrast, the cell on the far right in the first column (b) and the cell in the center of the middle column (e) both express Gag in a coarsely punctate pattern than most likely represent sites of Gag assembly at the plasma membrane. In both cases endogenous ABCE1 takes on a coarse punctate appearance (a, d). Merged images reveal that ABCE1 is co-localized with Gag in these cells (c, f). In cells with low levels of Gag expression, a large pool of diffuse ABCE1 is still present in addition to a small pool of punctate ABCE1 (cell on right in a, c, e). In cells expressing higher levels of Gag, a larger percentage of endogenous ABCE1 has been recruited into punctate sites of assembly, and the residual pool of diffuse ABCE1 is much smaller. As expected, the p41 Gag mutant fails to form punctate sites of assembly (h) or to recruit HP68 from diffuse pools (g) as seen by failure of co-localization (i). (From Zimmerman et al., Nature 415(6867), 88-92 Fig. )

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