Mapping of the Fbw7 dimerization domain (1). A: Fbw7α can form dimers. Flag- and Myc-tagged versions of Fbw7α as well as the Flag-tagged WD40 domain were transfected in 293A cells as indicated and analyzed for their interaction by immunoprecipitation with Flag antibody. Co-precipitated Myc-tagged Fbw7α was detected with 9E10 antibody (upper panel). The membrane was subsequently re-probed with Flag antibody (lower panel). Flag-Fbw7α corresponds to the lower band, Myc-Fbw7α to the upper band. The asterisk indicates the heavy chain of the Flag antibody used for immunoprecipitation. B: Analysis of N-terminal truncation mutants. Myc-tagged Fbw7α was co-expressed with Flag-tagged Fbw7α or various N-terminal truncation mutants as well as a point mutant of Fbw7α that no longer interacts with CPDs. Lysates were immunoprecipitated with Flag antibody and probed as indicated. ΔN refers to an N-terminal-less Fbw7 construct corresponding to the common region of Fbw7. All amino acid numbers reflect positions within the common region. The truncation mutants are depicted at the bottom. C: Fine-tuning of the dimerization domain beginning. The assay is similar as in A. The asterisk indicates the heavy chain of the Flag antibody used for immunoprecipitation.