Fig. 7. Katanin mislocalization does not affect the rate of exchange between polymerized and unpolymerized tubulin as assayed by fluorescence recovery after photobleaching (FRAP). Mitotic CV-1 cells expressing an integrated YFP-tubulin gene and transiently expressing either dsRed2-con80, which mislocalizes endogenous katanin from spindle poles, or dsRed2 alone were located and subjected to FRAP analysis. One half spindle was completely photobleached with the krypton/argon laser of a laser scanning confocal microscope. Recovery of fluorescence due to incorporation of unbleached YFP-tubulin was monitored by time-lapse confocal microscopy. Examples of recovery by two individual spindles are shown. I(bleached/unbleached) is the ratio of fluorescence intensity of the bleached half spindle to that of the unbleached half spindle. t-p is the time to reach 50% recovery to the apparent plateau of recovery. t is the predicted time to reach 50% recovery to an I(bleached/unbleached) ratio of 1. Averages with standard deviations were determined from 10 (dsRed2) or 13 (dsRed2-con80) spindles.

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