FIGURE 7.

Basolateral sorting motif in syntaxin 4 is independent of munc18c binding sequence. A, FLAG-munc18c cells were nucleofected with constructs for expression of EGFP or syntaxin 4 WT-EGFP or different syntaxin 4 mutants (syntaxin 4-L8K-EGFP, syntaxin 4-L8A-EGFP, and syntaxin 4-Δ10-EGFP). EGFP and the different syntaxin 4 proteins were immunoprecipitated with a GFP antibody and co-immunoprecipitated (IP) FLAG-munc18c was detected with a FLAG antibody. B, FLAG-munc18c cells in the presence or absence of dox were nucleofected with syntaxin 4-WT-EGFP, syntaxin 4-L8K-EGFP, and EGFP. EGFP and syntaxin 4-EGFP constructs were immunoprecipitated (IP) and co-immunoprecipitated FLAG-munc18c was visualized. Star indicates residual FLAG-munc18c that was not stripped off. IgG bands have been indicated. C, syntaxin 4 proteins were immunoprecipitated with a GFP antibody and co-immunoprecipitated endogenous munc18c (cmunc18c) was detected using a canine-specific munc18c antibody. D, quantification of co-immunoprecipitated munc18c, normalized for the amount of immunoprecipitated syntaxin 4. The results are the mean ± S.D. of three independent experiments.