Fig. 6

Figure 6. A Dvl-independent interaction mechanism between mammalian GPR124 receptors and Frizzled (A) Distribution of Dvl-GFP co-expressed with full-length (FL) zebrafish (z), mouse (m), or human (h) Gpr124 or their ΔICD (ICD1–80 and corresponding) variants in FZ1–10−/− HEK293T cells. The arrowhead points to Dvl-GFP signal at the plasma membrane. (B) GFP-Trap co-immunoprecipitation assays in total lysates of FZ1–10−/− HEK293T cells co-expressing Dvl-GFP and N-terminally FLAG-tagged zebrafish, mouse, or human FL Gpr124 and their ΔICD variants; n = 3. (C) Distribution of individually expressed TagRFP-fused zebrafish or mouse FL Gpr124, or their ΔICD variants, as well as Fz4-GFP in zebrafish blastula deep-layer cells. (D) Representative images of zebrafish blastula deep-layer cells co-expressing Fz4-GFP and zebrafish FL or ΔICD Gpr124-TagRFP, with or without Dvl; n = 3. (E) Quantification of the enrichment level of Fz4-GFP in the Gpr124-TagRFP+ membranes in the presence or absence of Dvl. (F) Same as (D) with corresponding mouse GPR124 constructs. For (C), (D), and (F), the cells annotated with an asterisk are magnified, and the pixel intensity of the green and magenta channels along a virtual clockwise path following the cell cortex from a to b is plotted. (G) Same as (E) for mouse GPR124 constructs. (H) Distribution of mouse Fz4-GFP in zebrafish blastula deep-layer cells in the presence or absence of Dvl. (I) Bimolecular complementation assay strategy. Illustration created with BioRender.com. (J) Representative images of zebrafish blastula deep-layer cells co-expressing the indicated bimolecular complementation constructs, with or without Dvl. Venus junctional signal is indicated with arrowheads; n = 3. Data are represented as mean ± SD. Scale bars, 10 μm. Related to Figure S4.