127cytoplasm. This was initially suggested by the observation that both the sperm-aster
after fertilization and bipolar spindles during cleavage become centered within the
cell before astral microtubules reach the cortex. Similarly, in the abovementioned
microtubule inhibitor uncaging experiment (Wühr et al. 2010 ), spindle movement
away from the site of local microtubule depolymerization occurred before the aster
reached the cortex. As in cultured cells, interference with dynein function in zebraf-
ish blastomeres (in these experiments through injection of p150-CC1, a dominant-
negative form of the dynein partner dynactin) results in spindles that are uncentered
as well as misoriented (Wühr et al. 2010 ). A similar requirement for dynein function
was demonstrated for the centering of the sperm-aster immediately after fertiliza-
Fig. 4.3 Demonstration that asters are centered by pulling, not pushing, forces. (a) Treatment of
embryos with colcemid leads to inhibition of microtubule polymerization, and localized inactiva-
tion of colcemid with UV light results in the movement of the aster toward the site of microtubule
growth (Hamaguchi and Hiramoto 1986 ). (b) Local uncaging of the microtubule inhibiting drug
combretastatin 4A results in the movement of the aster away from the site of microtubule inhibi-
tion (Wühr et al. 2010 ). A model in which asters become centered by microtubule pushing forces
predicts the opposite effects on aster movement and is ruled out by observations
4 Vertebrate Embryonic Cleavage Pattern Determination