Regulation of epithelial integrity and organ growth by Tctp and Coracle in Drosophila
Fig 1
Cora interacts with Tctp and is required for Tctp maintenance.
(A) GST-pull down of Cora and Tctp. GST-Cora directly binds to MBP-Tctp. The upper blot shows MBP-Tctp pulled down by GST-Cora and stained with an anti-MBP antibody. The lower blot shows GST tagged proteins used for GST-pulldown. (B) Co-immunoprecipitation of Cora and Tctp using S2 cells. 5% input was used as a positive control for western blotting. Myc-Tctp is co-immunoprecipitated by Flag-Cora. (C) An endogenous Cora isoform of about 200 kDa is co-immunoprecipitated with Tctp from wild-type embryos. (D) Western blot analysis of Tctp and Cora levels in the embryo. Tctp RNAi using actin-Gal4 does not affect the Cora protein level. In contrast, cora RNAi reduces the level of Tctp. The graph shows the quantification of three western blot results. (E-F”) Embryos stained for Cora and Tctp. In hindgut, Cora (green) is localized to the basolateral membrane (E, arrows) while Tctp (red) is ubiquitously distributed (E’). Dashed lines indicate the position of the lumen of the hindgut. In the epidermis, Cora and Tctp show overlapping localization at the basolateral membrane (F-F”, arrows). (G-H”) Effects of cora or Tctp mutation on the level of Cora and Tctp, respectively. cora4 and Tctph59 were balanced with CyO-GFP and TM3-GFP, respectively, to identify the genotypes of progeny. Distribution of Cora (shown as white staining) is normal in Tctph59 mutant embryo (G”’, arrows). Tctp (shown in red) is strongly reduced in cora4 mutant embryo (H”) at stage 14. The embryonic stage was determined based on the segmented pattern of the epidermis and the shape of the midgut (see S1C–S1D”’ Fig). GFP-positive embryos are heterozygotes for Tctph59 (G) or cora4 (H). Scale bars, 50 μm.