Figure 2.

The E295A mutation disrupts the intermolecular interaction of Ephexin4. (a,b) 293T cells were transfected with the indicated plasmids and lysed 2 days later. Cell lysates were incubated with FLAG-conjugated agarose beads. Bead-bound proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), transferred to a nitrocellulose membrane, and detected by immunoblotting. IP, immunoprecipitation. n = 3. (c,d) 293T cells were transfected with Ephexin4 (c) or Ephexin4^E295A (d). Two days later, cells were harvested, resuspended in PBS, and incubated with 0.1 mM DSS for 30 min. Thereafter, cells were lysed and proteins were detected by immunoblotting. n = 4. (e) 293T cells were transfected with the indicated plasmids. Two days later, cells were lysed and cell lysates were incubated with FLAG-conjugated agarose beads. Bead-bound proteins were detected by immunoblotting. n = 3. Images shown are representative of at least three independent experiments.