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. 2004 Oct 1;101(41):14800–14805. doi: 10.1073/pnas.0406451101

Fig. 4.

Fig. 4.

Cup interacts with eIF4E (4E) through an eIF4E-binding motif. (A) Cup deletions tested in a yeast two-hybrid assay. Growth on selective media containing 4 mM 3-aminotriazole (+) is indicated on the right. Amino acids 342–453 (Cup100) represent the minimal region of Cup interacting with eIF4E (black box). The striped box indicates the Nos-binding domain (amino acids 593–962) (4). (B) Cup100 interacts with eIF4E in a GST pull-down assay. Retained proteins were eluted and subjected to SDS/PAGE. (C) Coimmunoprecipitation assay using an α-HA antibody of 35S-Met-labeled Cup100 and HA-tagged eIF4E produced in reticulocyte lysates. Cup100 coimmunoprecipitates with HA-eIF4E (Cup100 + 4E). (Right) Control immunoprecipitations with α-HA antibody show specific immunoprecipitation of HA-eIF4E but not of Cup100. (D) Yeast two-hybrid assay using Cup100 (wild-type) and Cup100-MEBS (mutant). Yeast transformants were streaked on selective media containing 4 mM 3-aminotriazole. AD, GAL4 AD; DBD, GAL4 DBD.