Arl2 regulates centrosomal localization of D-TACC and Msps through dynein. (A) Metaphase NBs of control, Arl2T30N, arl2Δ156/+, and Arl2Q70L labeled for Venus-Ctp, CNN, α-tubulin, and DNA. Arrowheads indicate Venus-Ctp observed on spindle microtubule. (B and C) NBs of control and ctpexc6; insc-Gal4; Venus-CtpCAAX labeled for D-TACC, γ-tubulin, GFP, PH3, and DNA (B) and Msps, γ-tubulin, GFP, PH3, and DNA (C). (D and E) Interphase NBs of control and dhc64C4-19 clones labeled for D-TACC, γ-tubulin, GFP, PH3, and DNA (D) and Msps, γ-tubulin, GFP, PH3, and DNA (E). (F) Larval brains expressing full-length Msps (Msps-FL), Arl2T30N, arl2Δ156/+ (Arl2T30N, arl2Δ156/+), and Msps-FL with Arl2T30N, arl2Δ156/+ stained with Dpn. Central brain is to the left of the white dotted line. (G) Quantification of NB number per brain hemisphere (with SD) in F. Msps-FL, CD8-GFP: 96.8 ± 5.8 (n = 20); CD8-GFP, Arl2T30N, arl2Δ156/+: 401.8 ± 79.8 (n = 20); Msps-FL, Arl2T30N, arl2Δ156/+: 100.0 ± 26.1 (n = 20). ***, P < 0.001. (H) Metaphase NBs of Msps-FL, Arl2T30N, arl2Δ156/+, and Msps-FL with Arl2T30N, arl2Δ156/+ labeled for Insc, α-tubulin, and DNA. Quantification of spindle orientation is shown in the right panels. (I) NBs of control, Msps-FL, Arl2T30N, and Arl2T30N, and Msps-FL coexpression labeled with α-tubulin and DNA. (J) Metaphase NBs of control, Msps RNAi, and Msps RNAi in arl2Δ156/+ background and coexpression of Msps RNAi with Arl2T30N labeled for aPKC and DNA. (K) Quantification of aPKC localization for J. aPKC crescents were seen in all control metaphase NBs (n = 38). In Msps RNAi, 55% of metaphase NBs still displayed aPKC crescents (n = 51), whereas only 28% (n = 54) of Msps RNAi, arl2Δ156/+, and 25% (n = 40) of Msps RNAi, Arl2T30N localized aPKC properly. NB outlines are indicated by white dotted circles in A–E and I. Arrows indicate centrosomes and spindle poles. Bars: (F) 20 µm; (A–E and H–J) 5 µm.
