Fig. 6

tmi mutants exhibit defects in the segregation of dorsal determinants. (A) Females heterozygous for the maternal-effect tmi mutation consistently produce a fraction of embryos with ventralization phenotypes in 24 h embryos (V1-V4; ventralization series as in Kishimoto et al., 1997), with V4 the most severe, radially symmetric phenotype (see Fig. S7 for live images). (B-D) wnt8a RNA, originally localized to the vegetal pole of the egg, undergoes a lateral shift towards the future dorsal axis in wild-type embryos (B), a behavior absent in embryos from tmi (C) and hec (D) homozygous mutant mothers. Brackets indicate observed extent of wnt8a RNA at the cortex. (E-G) Microtubules at the vegetal cortex develop into a parallel array of microtubule bundles aligned toward the future dorsal axis (E) (Jesuthasan and Strähle, 1997; Tran et al., 2012; Ge et al., 2014). Vegetal microtubule bundling and alignment is defective in embryos from females homozygous mutant for tmi (F) and hecate (G). (H,I) Labeling of wnt8a RNA-containing particles and microtubules at the vegetal cortex of 20 mpf embryos, showing wnt8a RNA localizing to microtubules in both wild type and tmi mutants, despite different microtubule reorganization. H and I are 2D maximum projections of confocal z-stacks (see Movies 1 and 2 to view individual z-slices), and H′ and I′ show magnified views from individual z-slices. Quantification of individual z-slices (see Materials and Methods; Fig. S8) shows no statistically significant difference in the percentage of wnt8a RNA particles that co-occur with microtubules (wild type: 94.4%, n=10 ROIs, 5 embryos; tmi: 93.0%; n=10 ROIs; 5 embryos; P=0.472; unpaired Student's t-test).