Figure 4

Contribution of the different domains of MxB to capsid binding and restriction of HIV-1. (A) Wild type human MxB protein is depicted in the top figure. The numbers of the amino acid residues at the boundaries of the MxB domains are indicated. The different MxB variants are shown. (B) The ability of wild type and mutant MxB proteins to bind in vitro assembled HIV-1 CA-NC complexes was measured, as described in Methods. Similar results were obtained in three independent experiments, and the standard deviation for bound fractions relative to input are shown. (C) Cf2Th cells stably expressing wild type and the indicated MxB variant were analyzed by Western blotting using anti-FLAG antibodies. As loading control, cell lysates were Western blotted using anti-GAPDH antibodies. (D) Cf2Th cells stably expressing wild type and the indicated MxB variants were challenged with increasing amounts of HIV-1 viruses expressing GFP as a reporter for infection (HIV-1-GFP). Forty-eight hours post-infection the percentage of GFP-positive cells was determined by flow cytometry. As a control, Cf2Th cells stably transduced with the empty vector LPCX were challenged with increasing amounts of HIV-1-GFP. Similar results were obtained in three independent experiments and a representative experiment is shown.