Two different point mutations of Ci-Aurora prevent midbody localization.

<p><b>A</b>) Epifluorescence images from a 4D time-lapse series (time in min. is indicated). Point mutation of Ci-Aurora S60R::Venus prevents localization to the spindle pole as expected. However, midbody localization is also perturbed. Ci-Plk1::mCherry localization to the central spindle and midbody is indicated (red arrows) while Ci-Aurora S60R::Venus shows no midbody localization in the same cell. Corresponding brightfield images are shown. Scale bar = 10 µm. n = 9, 3 animals. <b>B</b>) Epifluorescence images from a 4D time-lapse series. Point mutation of Ci-Aurora S60R::Venus prevents midbody localization. Ci-TPX2::mCherry localizes to the midbody (red arrow) while Ci-Aurora S60R::mCherry is completely absent from the midbody at telophase. Corresponding brightfield images are shown. Scale bar = 10 µm. n = 9, 3 animals. <b>C</b>) Epifluorescence images from a time-lapse sequence showing that point mutation of Ci-Aurora G103N::Venus prevents midbody localization (grey circle) during telophase. Note that centrosome localization during interphase is preserved (blue arrows). HH2B::Rfp1 was used to precisely follow the mitotic cycle. Note also that AuroraG103N::Venus did not localize to the chromosomes. Corresponding brightfield images are shown. Scale bar = 10 µm. n = 23, 3 animals. D) GST-CiAur^wt, CiAur^G103N and CiAur^S60R were retained on the Sephadex beads for later interaction with radioactive ³⁵S-labeled CiTPX2. More TPX2 was found attached to Aur^wt than to either Aur^G103N or Aur^S60R. Three experiments.