Figure 8.

Golgi SNARE complex is destabilized in Cog3p- and Cog7p-depleted cells. (A) HeLa cells stably expressing GFP-GS15 were treated with scrambled (control) and COG3 siRNA for 72 h. GFP-GS15–containing SNARE complexes were precipitated from the 1% Triton X-100–solubilized cell lysates using anti-GFP antibody. 10% of the cell lysate (S20) and 100% of immunoprecipitate were immunoblotted with antibodies as indicated. Note that polyclonal anti-Syntaxin5 antibodies recognize both short (Syntaxin5a) and long (Syntaxin5b) isoforms of Syntaxin5 proteins. The results were plotted in a bar graph. (B) HeLa cells were treated with scrambled (control) and COG7 siRNA for 72 h. GS28-containing SNARE complexes were precipitated from the 1% Triton X-100–solubilized cell lysates using anti-GS28 antibody. 10% of the cell lysates (S20) and 100% of the immunoprecipitate (IP) were immunoblotted with antibodies as indicated. Error bars represent mean intensity of protein bands, calculated with ImageJ software, from three independent experiments