Ikbkap deficiency causes defects in oocyte maturation, spindle organization and chromosome alignment. (A–C) Fully-grown oocytes isolated from Ctrl and CKO mice were cultured for 16 h to evaluate the rate of polar body extrusion (PBE) (B) and abnormal division rate (C). (A) Photomicrographs of representative Ctrl and CKO oocytes. Arrowheads indicate oocytes that extruded a large-sized polar body; the arrow indicates an oocyte with apparent symmetrical division. Scale bar, 100 μm. (B,C) Quantitative analysis of Pb1 extrusion rate (B) and abnormal division rate (C) in Ctrl (n = 91) and CKO (n = 76) oocytes. Data are presented as mean ± SEM of three independent experiments. **p < 0.01 vs. Ctrl. (D) Ctrl and CKO oocytes were stained with acetylated α-tubulin antibody, H4K20me3 antibody, and DAPI to visualize the spindle (green), pericentric chromosomal regions (red), and chromosomes (blue), respectively. Ctrl MII oocytes presented a typical barrel-shaped spindle and well-aligned chromosomes on the metaphase plate. Apolar spindles, multipolar spindles (arrows) and chromosome misalignment were frequently observed in CKO MII oocytes. Scale bars, 5 µm. (E) Quantification of Ctrl and CKO oocytes with abnormal spindle or chromosome organization. Graph shows mean ± SEM of results obtained from at least six independent experiments in which ≥190 oocytes were analyzed. ***p < 0.001. (F) Western blot analysis showing the decreased acetylation of α-tubulin in CKO oocytes compared to the Ctrl group. The level of α-tubulin was not changed significantly in CKO oocytes compared to the Ctrl oocytes. β-actin served as a loading control.