Figure 3

The coiled-coil and caveolin-binding domains are required for oligomerization and PP2A binding but the calmodulin-binding domain is not. (A) HEK293 cells transfected with HA-tagged wild-type and mutant striatins were lysed and HA-striatin immune complexes were isolated, separated by SDS-PAGE, and proteins contained in the immunoprecipitates were detected by immunoblotting using antibodies that recognize the HA-epitope tag, PP2A C subunit, Mob3, SG2NA, and zinedin. Deletion of either the coiled-coil domain [Δ(70-116) striatin] or the caveolin-binding domain [Δ(53-66) striatin] of striatin resulted in loss of oligomerization and PP2A binding. (B) Relative binding of wild-type and deletion mutant striatins to PP2A C subunit, Mob3, SG2NA, and zinedin was measured as described in the legend to Figure 2B. (C) A similar experiment to (A) was conducted with wild-type striatin and Δ(148-166) striatin, which deletes the striatin calmodulin-binding motif. (D) Relative binding of wild-type and mutant striatins to PP2A C subunit, Mob3, SG2NA, and zinedin was measured as described in the legend to Figure 2B. The HA-striatins migrated at different positions due to their size differences but are shown side by side in panels A and C for comparison of their levels. The error bars in panels B and D represent the standard deviation of at least three independent experiments. *, p ≤ 0.05; **, p ≤ 0.01 relative to wild-type.