Fig. 4: Kinetochores partially masked by the small chromosome arm are more likely to be incorrectly attached to microtubules.

a Illustrations show differences in the telomere’s position with respect to the kinetochore in acrocentric and metacentric bivalents when aligned in a metaphase plate. Differences in the lengths of the arms result in differences in the distance of the telomeres from the kinetochores in the two morphologies. Magenta, kinetochores; cyan, telomeres; blue, chromosomes; green, spindle axis; white, chiasmata. b Illustrations and representative Airyscan immunofluorescence images with telomere-masked kinetochores (top row) and exposed kinetochores (bottom row) in porcine oocytes. Magenta, kinetochores, (ACA); cyan, telomeres, (TRF-2); blue, chromosomes, (Hoechst). Scale bar, 0.5 μm. Representative examples of 44 immunolabelled porcine oocytes. c Illustrations and representative Airyscan immunofluorescence images from the attachment types of telomere-masked and exposed kinetochores in porcine oocytes. Arrowheads indicate the specified attachment type. Magenta, kinetochores, (ACA); cyan, telomeres, (TRF-2); blue, chromosomes, (Hoechst); green, microtubules, (a-tubulin). Scale bar, 1 μm. Representative examples of 44 immunolabelled porcine oocytes. d Quantification of the proportion of kinetochore-microtubule (Kt-Mt) attachments for telomere-masked and exposed kinetochores of acrocentric chromosomes in porcine oocytes (44 oocytes analysed). Two-sided Fisher’s exact test (end-on, p < 0.0001; latera/merotelic, p = 0.0087; unattached, p < 0.0001). e Distribution of telomere-masked and exposed kinetochores of acrocentric chromosomes on spindles with fully aligned chromosomes and spindles that have not yet aligned their chromosomes (44 oocytes analysed). Two-sided Fisher’s exact test (p = 0.0016). f Rendering of segmented acro-labelled kinetochores (yellow, TALE labelled) and non-labelled kinetochores (magenta) in a meiotic metaphase spindle (microtubules in green). The white arrow indicates the spindle direction. Scale bar, 5 μm. Representative examples of 32 immunolabelled porcine oocytes. g Magnification of the kinetochore shown in F, white square, and the corresponding confocal, 3D-STED, and segmented kinetochores in XY, XZ, and YZ views. Each tile is 2 × 2 μm. h Illustrations and representative images of different angles of kinetochores in acrocentric chromosomes. The black arrow on the left side represents the spindle axis. The numbers in each image are the dot products between the left kinetochore longest axis and spindle axis. Magenta, kinetochores; blue, chromosomes; black, spindle axis. i Box plot showing the kinetochore orientation to the spindle for the three kinetochore populations (32 oocytes analysed). Box plot shows the median (horizontal black line), mean (small black squares), 25th and 75th percentile (boxes), and outliers (whiskers). Wilcoxon two-tailed test, (acro-labelled/meta, p = 2.38e − 06; acro-labelled/meta-labelled, p = 0.002; meta/meta-labelled, p = 0.181). j Distribution of the dot-product for the kinetochore orientation to the spindle for acro-labelled (upper graph), meta (middle graph), and meta-labelled kinetochores (lower graph) (32 oocytes analysed). Number of kinetochores analysed is indicated in brackets next to each category. P-values in the graphs are indicated as follows, **p < 0.01, ****p < 0.0001, and n.s.: non-significant. The number of cells is indicated in brackets in the figure legend.