Figure 6: Interaction of PC1 cytoplasmic C-terminal tail with Rabep1.

(a) Yeast two-hybrid screening of a rat brain cDNA library using human PC1 215-amino-acid C-terminal tail as bait (PC1-CT) identified a clone (TH10-3) corresponding to amino acids 465 to 799 of Rabep1 with extensive coiled-coils⁶⁹. (b) Mapping of Rabep1 interaction region by yeast two-hybrid assay using a series of PC1-CT deletion constructs²². The coiled-coil domain is indicated. PC1-M7 contains proline at a and d positions of the heptad, disrupting α-helical structure of the coiled-coil domain. PC1-P89 contains a germline mutation R4227X^22,27. PC1-C15 contains a somatic mutation found in a renal cyst resulting in reading-frame shift^22,70. Positive interaction was scored by both growth on triple selective medium and positive β-gal activity. PC2 C-terminal tail (PC2-CT) or Fos served as negative controls. Interactions of the PC1 constructs with PC2-CT are shown as a comparison. (c) Co-immunoprecipitation of endogenous PC1 with Rabep1, but not with Arf4 in CD cells.