Fig. 2.

HLTF degradation induced by HIV-1 Vpr precedes G2/M arrest. (A) Kinetic of HLTF disappearance. HeLa cells were transduced with empty VLP or WT Vpr VLP for 2 h. Cells were harvested 6, 12, 24, and 48 h after VLP treatment, and whole cell lysates were analyzed by Western blot. Quantification was performed: ratios between the HLTF and the GAPDH signals were calculated relative to a 100% reference indicated in red. The Western blot is representative of two independent experiments. (B) Kinetic of Vpr-mediated G2 arrest. Cell cycle analysis after VLP treatment, same time points as in A. (C) Short time kinetic of HLTF disappearance. HeLa cells were transduced with empty VLP (R−), VLP containing WT Vpr (R+), or Vpr mutants K27M or S79A. Cells were harvested at time 0 min, 30 min, 2 h, and 4 h after the 2-h VLP treatment, and whole cell lysates were analyzed by Western blot as in A. Quantification was performed as in A. (D) HLTF degradation precedes Vpr-mediated cell cycle arrest. Cell cycle analysis performed after VLP treatment at the indicated time points (same as in C) for the 6-h time point: 6(−) indicates with no MG132 treatment and 6(+) with MG132 all along the kinetic.