Fig. 5. The interaction between ADAM22 and 14-3-3 isoforms is dependent on the phosphorylation of ADAM22 but not the phosphorylation of 14-3-3 proteins. (A) Lysates of HEK 293T cells expressing either full-length FLAG-tagged ADAM22 or Myc-tagged or HA-tagged 14-3-3 isoforms were treated with alkaline phosphatase (ap) as indicated. Alkaline phosphatase-treated samples were subsequently treated with sodium vanadate prior to incubation with the corresponding cell lysates. Pooled lysates were then immunoprecipitated with anti-Myc-tag or anti-HA-tag antibodies and immunoblots probed with anti-FLAG antibody to detect ADAM22 precursor protein. Immunoblots were re-probed with anti-Myc-tag or anti-HA-tag antibodies as control for 14-3-3 protein expression. (B) Lysates of HEK 293T cells expressing full length FLAG-tagged ADAM22 were immunoblotted with anti-FLAG antibody as control for protein expression levels following alkaline phosphatase treatment. (C) Lysates of HEK 293T cells expressing full length FLAG-tagged ADAM22 were immunoblotted with anti-phosphoserine (pSer) antibody and re-probed with anti-FLAG antibody.