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Fig. 2. Bipolar spindle morphology is restored in cells co-depleted of Orbit and Klp10A. (A-D") Cells were treated with the indicated dsRNAs and stained for the distribution of microtubules (
-tubulin; red), 
-tubulin (green) and DNA (blue) 72 hours later. (A,A') Control metaphase spindles are bipolar and bi-centrosomal with a centrosome (green) attached to each end. (B) orbit RNAi commonly results in monopolar spindles with both centrosomes at the centre of a monoaster. Bipolar spindles may form in Orbit depleted cells, although these tend to be abnormally short (B'). After Klp10A knockdown, spindles form that are on average about 150% the length of controls. Half of these spindles are marked by the presence of both centrosomes at a single, well focused, spindle pole and an acentrosomal pole that is focused to varying degrees (C,C'). Double RNAi against orbit and Klp10A rescues bipolar spindle morphology. These bipolar spindles can be placed in three classes: bipolar bi-centrosomal spindles morphologically similar to controls (D), bipolar bi-centrosomal spindles significantly longer than controls (D') and bipolar monoastral spindles similar in length to controls but which have an asymmetric localisation of -tubulin. In this cell, the -tubulin diffusely stains one pole while at the other it forms a large aggregate in the centre of the aster, suggesting the presence of both centrosomes (D", see text for details). (E) Distribution of spindle lengths following each of the indicated treatments. Note the enhancement of spindle length in Klp10A single downregulated cells and the shortening of those in cells depleted of Orbit. The double knockdown of orbit and Klp10A can rescue the average spindle length and further restores the distribution of spindle lengths to values similar to that seen in the controls. All images are shown at the same magnification. Bars, 10 µm.
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