Fig. 4. PDZ1 is required for localizing INAD at the membrane of CHO cells. A–C, Wild-type and mutantinaD constructs containing point mutations (see Materials and Methods) in PDZ1, PDZ2, PDZ3, PDZ4, or PDZ5 were transfected into CHO cells. Shown are confocal images of immunofluorescently stained CHO cells transfected with wild-typeinaD (INAD; A),inaDPDZ1 (PDZ1;B), and inaDPDZ2(PDZ2; C). Left, Anti-INAD staining. Right, Anti-INAD (green) superimposed with rhodamine-conjugated phalloidin staining (red). INAD shows membrane-associated staining, as seen in A and E, in 72.7% of transfected cells (n = 414), whereas PDZ1 shows membrane-associated staining in only 19.9% of transfected cells (n = 272). In most cells PDZ1 (B, F) is expressed diffusely throughout the cell. The percentage of transfected cells displaying membrane-associated localization was 63.8% for PDZ2 (n = 315), 74.8% for PDZ3 (n = 302), 42.4% for PDZ4 (n = 85), and 40.6% for PDZ5 (n = 256). D–G, Wild-type and mutant INADs redistribute eye-PKC when cotransfected. Shown is immunofluorescent staining of CHO cells cotransfected withinaD (INAD),inaDPDZ1 (PDZ1), orinaDPDZ2 (PDZ2) with eye-PKC. Left, Anti-INAD staining. Right, Anti-PKC staining. PKC transfected alone displays a punctate, perinuclear expression pattern (D). When cotransfected with inaD (E),inaDPDZ1(F), orinaDPDZ2 (G), PKC is redistributed into an expression pattern like that of INAD, PDZ1, or PDZ2, respectively.