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. 2018 Mar 26;7:e33685. doi: 10.7554/eLife.33685

Figure 4. Dome is endocytosed via interactions with early/recycling endosome GTPases.

(A–C) Dome localization in nos-gal4 > UAS-YFP-rab5 GSCs at interphase (A), prophase (B), metaphase (C). Green: YFP. Red: Dome. (D) Co-immunoprecipitation of Dome with Rab5. Dome-GFP was pulled down from GSC extracts using an anti-GFP antibody and blotted using anti-Rab5 and anti-GFP antibodies. (E–K) Examples of spindles in control (E), nos-gal4 > UAS-YFP-rab4DN (F), nos-gal4 > UAS-YFP-rab5DN (G), nos-gal4 > UAS-YFP-rab7DN (H), nos-gal4 > UAS-YFP-rab8DN (I), nos-gal4 > UAS-YFP-rab11DN (J), nos-gal4 > UAS-YFP-rab35DN (K) GSCs (indicated by a white dotted line). Asterisk indicates the hub. Arrowheads indicate spindle poles. Green: Vasa and PH3. Red: Fas III and γ-Tubulin. White: YFP. Blue: DAPI. Bar: 5 µm. (L) Summary of GSC centrosome/spindle misorientation in the indicated genotypes. P value comparing control and the indicated genotype was calculated using two-tailed Student’s t-test. Error bars indicate the standard deviation. N = GSC number scored for centrosome orientation or N = mitotic GSC number scored for spindle orientation.

Figure 4.

Figure 4—figure supplement 1. Colocalization/immunoprecipitation of Dome with RabGTPases.

Figure 4—figure supplement 1.

(A–E) Dome localization in prophase of nos-gal4 > UAS-YFP-rab4 (A), nos-gal4 > UAS-YFP-rab7 (B), nos-gal4 > UAS-YFP-rab8 (C), nos-gal4 > UAS-YFP-rab11 (D), nos-gal4 > UAS-YFP-rab35 (E) GSCs. Green: YFP. Red: Dome. (F–I) Co-immunoprecipitations of Dome with Rab4 (F), Rab7 (G), Rab8 (H) and Rab11 (I). Dome-GFP was pulled down from GSC extracts using an anti-GFP antibody and blotted using anti-Rab4, 7, 8 or 11 and anti-GFP.
Figure 4—figure supplement 2. Early endosome GTPases rab4 and rab5 are required for correct Dome localization.

Figure 4—figure supplement 2.

(A–C) Examples of Dome staining in control interphase (A), prophase (B) and metaphase (C) GSCs. Asterisk indicates the hub. Green: Vasa. Red: Dome. (D–F) Examples of Dome staining in nos-gal4 > UAS-YFP-rab4DN interphase (D), prophase (E) and metaphase (F) GSCs. (G–I) Examples of Dome staining in nos-gal4 > UAS-YFP-rab5DN interphase (G), prophase (H) and metaphase (I) GSCs. Asterisk indicates the hub. Green: YFP. Red: Dome. Bar: 5 µm.