Figure 1.
FBP17b, FBP17, and CIP4 interact with AKAP350. (A) Genomic organization of human FBP17. The 15 exons are denoted, with the dashed boxes indicating the exons that are spliced out in the FBP17B transcript. The splicing does not interrupt the exons comprising the MBD, the cdc42 interacting domain (CID), or the SH3 domain. (B) Diagram showing identity (top numbers) and similarity (bottom numbers) between FBP17 exons 1-15, and CIP4 exons 1-14. (C) FBP17, FBP17b, CIP4, and the CIP4 deletion constructs of the MBD or the C-terminal SH3 domain were cloned in pBD-Gal4 vector. The interaction of these proteins with AKAP350 was assessed in yeast two-hybrid binaries assays with pAD-AKAP350(1076-2143). The results confirm FBP17b interaction and indicate that FBP17 and CIP4 also interact with AKAP350(1076-2143) and that the MBD, but not the SH3 domain, in CIP4 is necessary for the interaction. (D) In vitro confirmation of AKAP350/CIP4 interaction. (His)6-tagged CIP4 was attached to Ni beads. A 100,000 × g gastric mucosal supernatant (SM) was incubated with His-CIP4 Ni beads (CIP4) or Ni beads alone (blank). After incubation, samples were washed and eluted with sample buffer. Samples were subjected to electrophoresis in 3-10% SDS-PAGE followed by Western blotting with anti-AKAP350 monoclonal antibody (14G2). Bar, 5 μm.