Figure 2
CnAα interacts with NIK through its phosphatase domain.
A. Co-immunoprecipitation of CnAα and its mutants (ΔN and ΔCA) with NIK. CnAα and its mutants expressed in cells are indicated at the top of panels. Control indicates the Myc-tagged expression vector. The upper left panel (Co-IP) shows western blotting of immunoprecipitates using the anti-Flag antibody to detect co-immunoprecipitation of Myc-tagged CnAα and its mutants. The lower left panel shows western blotting of immunoprecipitates using the anti-Flag antibody to detect Flag-tagged NIK. The upper right panel shows western blotting of total cell lysates using the anti-Myc antibody. The lower right panels show western blotting of immunoprecipitates using the anti-Flag antibody to detect Flag-tagged NIK. Arrows indicate bands of IgG chains used for immunoprecipitation. Results of one representative experiment of three are shown. Blots are cropped for clarity. Full-length blots of key data are presented in Supplementary Figure 2. B. Schematics of CnAα and its deletion mutants used in this study. “Phosphatase” indicates the phosphatase domain containing the catalytic domain and regulatory subunit-binding domain. “CaM” indicates a potential calmodulin-binding domain. “AI” indicates the auto-inhibitory domain. The Flag tag (abbreviated in this figure) was connected to the N-terminus of the wild-type protein and mutants. The binding ability of each protein for NIK, as determined in Fig. 2A, is indicated at the right of each structure. “+” indicates positive for binding and “−” indicates negative for binding.
