Both the amino terminal and DH domains of XGef interact with CPEB. (A) Schematic diagram of wild-type and deletion mutant versions of HA-XGef. The HA-XGef wild-type or mutant name includes either the amino acids retained in the clone, or the Δ prefix to indicate which amino acids were removed. Interaction of the HA-XGef version with GST-CPEB is indicated by a plus sign, no interaction by a minus sign and very reduced interaction by (-). PH, pleckstrin homology domain; CSLL, amino terminal CAAX box; black squiggle, putative coiled-coil domain. (B) Oocyte extracts expressing various versions of HA-XGef were analyzed by SDS-PAGE and immunoblotting with HA antibody. HA-XGef(235–465) and HA-XGef(65–234) are not shown. (C–E) Representative interaction assays. Oocyte extract containing equivalent amounts of HA-XGef WT or a mutant HA-XGef version were combined with extract containing GST-CPEB or GST. Glutathioneagarose captured proteins were analyzed by SDS-PAGE and immunoblotting with anti-GST or anti-HA antibodies (IB: HA or IB: GST). The different combinations of fusion proteins combined into individual reactions are indicated in the grid above the picture. Immunoblots with only the most informative binding assays are shown.