Fig. 7. Effects of co-depleting both condensin and cohesin from Xenopus egg extracts. (A) Western blot of mock- and double-depleted (DD) extracts showing that a cohesin subunit (SMC1), and a condensin subunit (XCAP-G) have been depleted. Background bands are marked with asterisks. (B) Spindle assembly defects are additive. Spindles assembled in extracts depleted using control (Mock), cohesin (Cohesin), condensin (Condensin) or both (Cohesin+Condensin) antibodies were evaluated for spindle assembly defects. Bars represent the average of two experiments, with more than 400 microtubule structures evaluated under each condition. Error bars represent standard deviation (± s.d.). (C) Chromosome morphology in single- and double-depleted extracts. Right set of panels show the position of chromosomes stained with Hoechst-33258 dye (blue) within metaphase spindles (microtubules are red). Left set of panels show higher magnification images of the same chromosomes (blue) stained with antibodies to kinetochore checkpoint protein BubR1 (red). Note misaligned chromosomes in the absence of cohesin. Arrowhead shows chromosome distortion apparent in the absence of condensin, probably due to pulling at kinetochores by spindle microtubules. Both the alignment and distortion defects are rescued in the double depletion. Bars, 10 µm.