IF was used to visualize the nuclear-cytoplasmic distribution of the wild-type and mutant Hlc. IF was performed as previously described (15 (link),30 (link),31 (link)). In brief, after treatment with fixative solution (75% methanol (v/v) and 25% glacial acetic acid (v/v)), fixed cells were permeabilized with 0.1% Triton X-100 (v/v) in TBST buffer (Tris-Buffered Saline Tween-20 pH 7.6: 0.242% Tris–HCl (w/v), 0.8% NaCl (w/v) and 0.05% Tween-20 (v/v)) at room temperature for 10 min and incubated with the indicated primary antibody in the presence of 5% bovine serum albumin (w/v; Beyotime, ST025) at 4°C overnight. After incubation with the fluorophore-conjugated secondary antibody, 1 μg/ml DAPI (Beyotime, C1002) was used to stain nuclei for 5 min before confocal microscopy analyses. Fluorescence signals were detected using the Leica TCS SP8 system and quantified using Image J. Line profiles were analyzed using Image-Pro Plus 6.0. These antibodies were used in IF: anti-Hlc (raised against amino acids 545–560 of the endogenous Drosophila Hlc), anti-FLAG (Beyotime, AF519), Alexa Fluor 555-labeled anti-mouse IgG (Beyotime, A0460) and Alexa Fluor 555-labeled anti-rabbit IgG (Beyotime, A0453).