Spindle oscillation is inhibited in nip100delta null cells arrested in S phase. nip100 is the yeast homologue of p150[glued] and encodes the large subunit of the cytoplasmic dynactin complex. In wild-type cells, the spindle moves in both directions through the bud neck; these movements coincide with lateral sliding of a cytoplasmic microtubule along the cell cortex (in movie S1, CIL# 23599). These events require dynein-dynactin function since nip100delta mutants display frequent contacts between microtubule ends and the cell cortex, but no spindle movements or microtubule sliding events were observed. Movie plays at 50x real time and is Movie S2 in Proc Natl Acad Sci. 2009. 106: 5147-5152.
S. cerevisiae (MATalpha nip100delta::hygB ura3-52 lys2-801 leu2-delta1::GFP-TUB1::LEU2 his3-delta200 trp1-delta63) expressing GFP-tubulin (GFP-TUB1) were arrested in hydroxyurea and mounted on agarose pads for microscopy. Images were captured on an Olympus Bmax-60F microscope equipped with a 1.35NA 100× UPlanApo objective, spinning disc Confocal Scanner Unit (CSU10), Picarro Cyan laser (488 nm), and a Stanford Photonics XR-Mega10 ICCD camera, by using QED software (Media Cybernetics). Image analysis was performed by using ImageJ. Timelapse images were captured at 10-s intervals for 15 min and each image in the movie represents a composite of 9 planes separated by 500 nm.
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