Figure 7.

NEM induces the redistribution of SNAP23. (A) NEM treatment of human fibroblasts leads to a decrease of the Triton X-100 insoluble (TX-100 insol.) pool of SNAP23 and an increased recovery of syntaxin 4 (Stx4) in SNAP23 immunoprecipitates. Immunoprecipitation was carried out from the Triton X-100 soluble (TX-100 sol.) fraction with sheep anti-rabbit magnetic beads coated with control rabbit immunoglobulins (Ctrl.) or anti-SNAP23 antibodies. The amounts loaded in each condition correspond to an equivalent cell number. Control rabbit immunoglobulins did not immunoprecipitate SNAP23 or Stx4. (Immunoglobulin heavy chains, IgG HC.) (B) SNAP23 localizes to the plasma membrane following NEM treatment. Human fibroblasts were treated (NEM) or not (Ctrl.) with NEM and double-stained for SNAP23 and vimentin. (C) NEM treatment decreases the vimentin-associated pool of SNAP23. Human fibroblasts were homogenized with a cell cracker and vimentin filaments were immunoisolated. Bars represent the average of three independent experiments. In each experiment, the intensity of the vimentin and SNAP23 bands was quantified. The amount of SNAP23 was expressed as a ratio of the amount of vimentin recovered in each sample. In each experiment, the SNAP23/vimentin ratio was set to 100 arbitrary units for control cells.