Figure 2.

KNL-1 localizes to kinetochores throughout mitosis. (A) KNL-1 is a 1010-amino-acid novel protein with a predicted molecular mass of 113 kD. The N terminus contains four repeats of a loosely conserved sequence motif (indicated in red and aligned beneath the sequence). The C terminus of KNL-1 has a region predicted to form a coiled coil (underlined and lettered in blue). (B) Western blots of embryo extracts probed using affinity-purified antibodies raised against two overlapping regions of KNL-1 (amino acids 1-150 and amino acids 8-256). (C) Projected 3D data sets of fixed embryos at different cell cycle stages stained for microtubules and DNA (red and cyan; left column), the kinetochore marker CeCENP-C (second column), and KNL-1 (third column). The merged images on the far right show CeCENP-C (red) and KNL-1 (green) at 2.5× higher magnification. (D) Immuno-EM of KNL-1 in multicellular embryos prepared by high-pressure freezing/freeze substitution. Insets were magnified 2×. The schematic to the right illustrates the positions of the gold particles in the field of view. Bars: C, 10 μm; D, 500 nm.