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First published online September 19, 2007
doi: 10.1242/10.1242/jcs.013698

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The Dam1/DASH complex is required for the retrieval of unclustered kinetochores in fission yeast

¹ Division of Yeast Genetics, National Institute for Medical Research, The Ridgeway, Mill Hill, London, NW7 1AA, UK
² Department of Biological Sciences, University of Warwick, Coventry, CV4 7AL, UK

View larger version (31K): [in this window] [in a new window]   Fig. 1. Unclustered kinetochore pairs are efficiently retrieved to the spindle pole before sister chromatid bi-orientation. (A) ndc80-gfp cdc11-cfp and mto1 ndc80-gfp cdc11-cfp cells were fixed and stained with DAPI to reveal the position of kinetochores (green), spindle poles (red) and chromatin (blue). (B) The frequency by which unclustered kinetochores appear in mto1 ndc80-gfp cdc11-cfp cells when the nucleus is placed in the middle of the cell (region A), displaced to one side of the cell but not at the cell tip (region B) or at the cell tip (region C) was calculated. (C) Images from a movie of mto1 ndc80-gfp cdc11-cfp cells in mitosis. Spindle poles are shown in red and kinetochores are shown in green. Note that an unclustered pair of sister chromatids (white arrowhead) are drawn back to the spindle pole and then bi-oriented on the mitotic spindle. Bar, 1 µm. (D) The distance between the unclustered kinetochore pair and the spindle pole was calculated at 30-second intervals in multiple movies of ndc80-gfp cdc11-cfp cells.

View larger version (59K): [in this window] [in a new window]   Fig. 2. Chromosome retrieval occurs at the plus end of depolymerising intranuclear spindle microtubules (INAs). Images from two movies (A,B) of mto1 fta2-tdtomato nmt1-gfp-atb2 cells with unclustered kinetochores. Microtubules are shown in green and kinetochores are shown in red. The position of the unclustered kinetochores is indicated by a white arrowhead. The plus ends of intranuclear spindle microtubules are indicated by a yellow arrowhead. Bar, 1 µm.

View larger version (33K): [in this window] [in a new window]   Fig. 3. Dam1 but not Klp5 is required for the retrieval of unclustered kinetochores. The distance between the unclustered kinetochore pair and the spindle pole was calculated at 30 second intervals in multiple movies of (A) dam1 mto1 ndc80-gfp cdc11-cfp cells and klp5 mto1 ndc80-gfp cdc11-cfp cells.

View larger version (51K): [in this window] [in a new window]   Fig. 4. Unclustered kinetochores associate with the lateral surface of spindle microtubule bundles in the absence of Dam1. Images from three movies (A, B and C) of mto1 dam1 fta2-tdtomato nmt1-gfp-atb2 cells with unclustered kinetochores. Microtubules are shown in green and kinetochores are shown in red. The position of the unclustered kinetochores is indicated by a white arrowhead. The plus ends of intranuclear spindle microtubules are indicated by a yellow arrowhead. Bar, 1 µm (2 µm in C).