Figure 5
A C-terminal KKVY-motif of Wg mediates its Golgi-to-ER retrieval.
(a-a’) UAS-vps35RNAi was driven by hhGal4 carrying sqh-eYFP-Golgi transgene. Wg was accumulated in the posterior wg-expressing cells (arrowhead in a). The colocalization of unsecreted Wg with Golgi marker was mildly enhanced (arrows in a’) compared with the wild-type control. (b-b”’) UAS-vps35RNAi was expressed using hhGal4. The unsecreted Wg shows increasing of colocalization with the ER marker, KDEL (arrows in b’,b”). (b”) shows the side-view of Wg-expressing cells. Z-section images were taken from apical to basal of (b). (c) Cross sections were cut along the Wg-expressing region for colocalization analysis. The Pearson’s Correlation (PC) value represents the colocalization of Wg with KDEL. The colcalization of unsecreted Wg with KDEL was significantly increased compared with the wild-type control. Data represent the Mean ± SEM (t-test, n = 10, ****P < 0.0001). (d-e”) Ectopic expression of Sens can be induced by either UAS-wg or UAS-wg(K334K > R334R) driven by dppGal4. (f-g’) UAS-wg or UAS-wg(K334K > R334R) was co-expressed with UAS-wlsRNAi using dppGal4, respectively. The genetic crosses and immunostaining were performed under the same condition. Images were taken with same laser setting on the confocal microscope. The accumulation of wild-type Wg in its expressing cells induces remarkable ectopic Bip expression (arrowheads in f). Whereas, the K-to-R mutant form of Wg does not show obvious induction of ectopic Bip (arrowheads in g). (h-i) UAS-wg or UAS-wg(K334K > R334R) was co-expressed with UAS-wlsRNAi and UAS-GFP-KDEL using dppGal4, respectively. Z-stack sections were taken with same laser setting on the confocal microscope. The subcellular distribution of Wg or Wg(K > R) was shown in the lower panel. The colocalization of Wg or Wg(K > R) with GFP-KDEL was shown in the right panel. (j) Cross sections were cut along the dppGal4-expressing region for colocalization analysis. Comparing with the wild-type Wg, the K-to-R mutant form of Wg shows weaker colocalization with GFP-KDEL. Data represent the Mean ± SEM (t-test, n = 15, ****P < 0.0001). The Wg dilution used in this Figure was 1:20. Scale bars in (a,b,h,i) 10 μm; (d–g) 50 μm.
