Fig. 7.

NSC and NPC phenotypes in Df(her4;12), Df(her2;15) double deficiency mutants. (A-B″) Confocal image stacks of anti-Sox2 and anti-pH3 immunostaining in 72 hpf WT (A) and Df(her4;12), Df(her2;15) double mutants (B). (C-D″) HCR-RNA FISH for detection of ascl1a/b, neurog1 and olig2 expression. (A-D) Lateral view midline sagittal xz plane from orthogonal reconstructions. Cyan lines show dorsal view horizontal confocal planes 1 and 2 shown in A′-D′, A″-B″ (A′-D′ dorsal part of the diencephalon with the thalamus proper; A″-B″ more ventrally including the prethalamus). Yellow lines in A′-D′ show midline sagittal planes in A-D. A₁ and B₁ show magnifications of the boxed areas in A′ and B′, with arrowheads indicating Sox2^low-expressing cells. (C′,D′) Blue lines show level of frontal orthogonal reconstructions in C″ and D″ (at level of the lateral expansion of the ventricle). Arrowheads in A″ and B″ indicate the lateral expansion of ventricular Sox2^high cells. Arrowheads in C′ and D′ show olig2 in parallel to the ventricular wall and adjacent to neurog1. Arrows show the Sox2^high NSC nuclei layer covering the ventricular wall. (E-E″) Anti-Sox2 and anti-pH3 nuclei counts for Notch-dependent (N-dep) and Notch-independent (N-indep) her mutants (Fig. 4; Table S8; Materials and Methods, ‘Microscopy and cell counts’). (E) pH3/Sox2⁺ count ratios in Sox2^high and Sox2^low cells. (E′) Sox2^high and Sox2^low nuclei counts in N-dep and N-indep her mutants. (E″) pH3/Sox2⁺ nuclei count ratios in Sox2^high or Sox2^low are not significantly changed in her6,her9 double mutants or double deficiency mutants. Data are mean±s.d. Two sample, unequal variance two-tailed t-tests. Six biological replicates each except WT N-indep (four) replicates; single experiments. A-P, anterior-posterior; D-V, dorsal-ventral; ns, not significant; pth, prethalamus; tec, tectum opticum; tel, telencephalon; th, thalamus; ve, ventricle. A-P, anterior-posterior; D-V, dorsal-ventral; pth, prethalamus; tec, tectum opticum; tel, telencephalon; th, thalamus; ve, ventricle. Scale bars: 50 µm; 20 µm (A₁,B₁,C′).