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. Author manuscript; available in PMC: 2010 Mar 1.
Published in final edited form as: Dev Cell. 2009 Mar;16(3):374–385. doi: 10.1016/j.devcel.2009.01.011

Figure 2. The Ska1 complex is required for kinetochore-microtubule attachments.

Figure 2

(A) Overexpression of GFP-Ska1 results in the localization to, and bundling of, interphase microtubules. HeLa cells were transiently transfected with a GFP-Ska1 plasmid and imaged 32 hrs after transfection. (B) Depletion of Ska1 or Rama1 results in a penetrant mitotic arrest. Graph shows percent of G2/M cells based on the fluorescent activated cell sorting and staining against DNA. Visual examination of these cells confirmed that they were arresting in mitosis (not shown). (C) Depletion of Ska1 and Rama1 results in severe chromosome segregation defects. Depleted cells were imaged for microtubules (using anti-tubulin antibodies), DNA, CENP-A (using anti-GFP antibodies in a cell line stably expressing GFP-CENP-A), and Rama1 (using anti-Rama1 antibodies). 1) Class 1 phenotype with the majority of chromosomes aligned at the metaphase plate. Arrows point to a single pair of off axis chromosomes in the depleted cells. 2) Class 2 phenotype with chromosomes severely mis-aligned. 3) Class 3 phenotype with mis-aligned chromosomes and multi-polar spindles. For each condition, 600 mitotic cells were counted to quantify the percent of cells with each of the indicated phenotypes. Scale bars, 10 μm.