Fig. 8

Chromosome, γ-Tubulin and microtubule organization in the spottyp08bdth mutant egg and zygote. A. Ectopic cytoplasmic domain formation in the spotty mutant. Instead of a regular organization of cytoplasm intermingled with YGs in the wild-type yolk cell (n = 74 embryos from 2 females), the spotty mutant (n = 65 embryos from 2 females) shows numerous, peripheral cytoplasmic domains (black arrows). B. Animal viewed confocal images of DiOC6- stained zygotes reveal that YG arrangement is affected, coinciding with peripheral patches of cytoplasm (white asterisks). Higher magnification DAPI- and γ-Tubulin-stained images show perturbed nucleus formation and γ-Tubulin assembly in spotty (n = 37 from 3 females) compared to wild-type (n = 33 from 3 females) zygotes at 25 mpf. Boxes show magnified areas below. C. DAPI-stained wild-type (n = 20 from 2 females) and mutant (n = 48 from 2 females) unactivated eggs showing chromosome organization during metaphase II of meiosis. D. α-Tubulin- and DiOC6-stained confocal z-projections showing MT and YG distribution at the cortex of wild-type (n = 34) and spotty (n = 41) eggs. Higher magnification images reveal the magnitude of such MT structures and YG organization in the mutant egg. E. α- and γ-Tubulin co-staining indicates that MTs emanate from numerous MTOCs across the mutant egg (n = 14). Image 4 was selected from a z-stack containing 31 x 1 μm-thick optical sections across the mutant egg. Boxes show magnified (B, D and E) areas. bd, blastodisc; yc, yolk cell; yg, yolk globules; cp, cytoplasmic pocket; A, animal pole; V, vegetal pole; mpf, minutes post fertilization. Scale bar: 190 μm (A), 200 μm (B, top row), 50 μm (B, bottom row), 5 μm (C), 180 μm (D, low magnification), 38 μm (D, high magnification), 180 μm (E, top row), 60 μm (E, bottom row).