Fig. 4. Colocalization of Par-6 complex with Crb complex and direct binding of Dlt to Par-6. (A-D) Both Par-6 (A, green) and aPKC (B, green) colocalize with Dlt (red, A-C) at the apical domain of photoreceptor cluster. aPKC not only colocalizes with Dlt in the apical membrane but also overlaps with AJs (B). Baz does not co-localize with Dlt (C). Baz colocalizes with Arm (D) at the AJ of photoreceptors. (E-G) In eye discs of the third instar larvae, Par-6 colocalizes with Dlt (E) and Baz colocalizes with Arm (G), but not with Dlt (F). (H) Direct binding of Dlt to Par-6, but not to aPKC or Baz. MBP-Dlt fusion protein was attached to amylose bead and used for the binding to GST-Par-6, GST-aPKC or GST-Baz. The blot was probed with anti-GST antibody. (I,J) Direct binding of Dlt by its PDZ3 region with Par-6. Each of GST constructs fused with Dlt deletion mutant proteins was attached to glutathione-agarose bead and used for the binding test with MBP-Par-6. The blots were probed with anti-MBP antibody. (K,L) Deletion of C-terminal PBM motif of Par-6 did not affect the binding of Par-6 to Dlt (K). GST or GST-Dlt was attached to beads and used for binding of MBP-Par-6 or MBP-Par-6C. The blot was probed with anti-MBP antibody (K). The N-terminal region of Par-6 is required for the binding to Dlt, but the PDZ domain is not (L). Each deletion mutant protein of Par-6 was attached to beads and used for binding with MBP-Dlt. The blot was probed with anti-MBP (L). (M,N) Schematic of molecular interaction between Dlt and Par-6. Binding is indicated by two-headed arrow. AID and CRIB are atypical PKC interaction domain and Cdc42-Rac1 interaction domain, respectively.

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