Functional antagonism between Patronin and KLP10A. (A–C) Excess KLP10A-GFP inhibited anaphase B spindle elongation. Stills (A) and pole–pole distance versus time (B) for a representative embryo microinjected with purified KLP10A-GFP, which formed a visible concentration gradient and localized normally (A, bottom). Close to the injection site (arrowheads, t = 0 s), KLP10A depolymerase activity destabilized metaphase spindles, whereas more distal metaphase spindles were stable but displayed no anaphase B spindle elongation (spindles 1 and 2 [s1 and s2]). Mitosis was normal at very distal sites (e.g., spindle 4 [s4]). Repeated in >20 embryos. (C) SDS-PAGE of microinjected KLP10A-GFP. M, protein markers. (D and E) Co-depletion of Patronin (pat) and KLP10A caused persistent spindle elongation from prometaphase on, just like KLP10A inhibition alone. (D) Pole–pole distance versus time from NEB (0 s) in injected embryos. Error bars show SDs. (E) Time-lapse images of embryos injected with anti-KLP10A (left) or with both anti-Patronin and anti-KLP10A (middle and right). Right images show that Patronin dissociated from spindles to form cytoplasmic immunoprecipitates. 0 s = NEB. Bar, 10 µm.