KLP10A inhibition is sufficient to induce anaphase B spindle elongation in preanaphase B–arrested spindles. (A and B) KLP10A inhibition in spindles arrested at metaphase (A; Video 5) or anaphase A (B; Videos 6 and 7). (left) Control spindles injected only with cyclin B (see also Fig. S1, E and F). (right) Spindles sequentially injected (inj) with cyclin B (CycB) and anti-KLP10A (see also Fig. S2 A). Top two rows show representative micrographs, and the bottom row shows corresponding kymographs. Time is in seconds from prometaphase (A) or anaphase A onset (B). Green, tubulin; red, histone. Bars: (horizontal) 5 µm; (vertical) 100 s. (C) In embryos sequentially injected with cyclin B and anti-KLP10A, the extent (P = 0.8609) and rate (P = 0.7766) of elongation of spindles arrested in metaphase (meta; 21 spindles in 9 embryos) or anaphase A (ana; 21 spindles in 7 embryos) were indistinguishable from wild-type anaphase B (22 spindles in 5 embryos). Error bars show SDs. (D) Anaphase A–arrested spindles (52 spindles in 11 embryos) were classified as (I) spindle elongation followed anaphase A and led to complete chromosome segregation (19%); (II) spindle elongation occurred after anaphase A, but lagging chromosomes or chromosome bridges formed (29%); (III) spindle elongation accompanied anaphase A, and bridges were present on chromosomes whose leading edges reached the poles (27%); and (IV) spindle elongation occurred with incomplete anaphase A, and the leading edges of chromosomes did not reach the poles (25%). Bar, 10 µm. 0 s = anti-KLP10A injection time. See also Fig. S2 B. (E) Spindle pole dynamics from NEB (0 s) in (a) an untreated embryo (wild type [WT]), (b) a representative embryo injected only with cyclin B before NEB (cyclin B [cycB] only), and (c) a representative embryo injected with cyclin B before NEB and anti-KLP10A at the indicated time (cyclin B + anti-KLP10A [anti-10A]). Repeated in >25 embryos. See also Fig. S2 A.