Fig. 7.

Drosophila Rab1 interacts with the COG complex. (A) Recombinant GST–Rab1 protein, immobilized on glutathione beads and loaded with either GDP-β-S (GDP) or GMP–PNP (GTP), was incubated with larval brain extracts expressing GFP–Cog7. GST–Rab1, but not GST, precipitated both GFP–Cog7 and Gio from brain protein extracts. Ponceau staining is shown as a loading control. 2% of the input and 25% of the pull-downs were loaded and probed with the indicated antibody. Molecular mass is given in kilodaltons. The graph represents quantification of the amount of GFP–Cog7 that was pulled down from each form of GST–Rab1 in western blotting analysis. Protein band intensities were obtained from three independent experiments. (B) Yeast two-hybrid assay was used to test Rab1 interaction with Cog5, Cog6 and Cog7 proteins. In the presence of Rab1 bait, only Cog6 induces LacZ expression (blue color indicates positive interaction). Graph shows quantification of LacZ reporter expression (graph) induced with different combinations of bait and prey plasmids. See Materials and Methods for further details. Error bars indicate s.e.m. *P<0.05; **P<0.01 (unpaired Student's t-test).