Figure 5.

Significant defects in cell behavior and brain development begin at third instar when mitotic rates increase. (A and B) Central nervous system (CNS), late-stage WT, and mad2 sas-4 embryos. Elav marks neurons. (C and D) Little mitosis (PH3⁺, green) occurred in second instar WT or mad2 sas-4 brains. The OOA (identified by Ecad expression) was present in both genotypes. (E and F) Brain size (E) and mitotic index (F) at second instar were not altered in mad2 sas-4. (G–I) Apoptosis was not increased in second instar mad2 sas-4 brains. The arrow in H indicates apoptotic cells. (J) Diagram of a few NBs in second instar brains (Dpn⁺, green) that had already produced clusters of progeny (Pros⁺, red). (K and L) WT and mad2 sas-4 NBs and their progeny. (M–O) By mid–third instar, WT and single mutant brains exhibited significant proliferation (PH3⁺; M) and brain size increases (O). In contrast, mad2 sas-4 brains began to display reduced size (O) and less proliferation (N). (P–R) mad2 sas-4 brains exhibited subtle but significant increases in apoptosis (Casp3, green) at mid–third instar. Arrow in Q marks an apoptotic cell. Error bars represent means ± SD. (S) In WT mid–third instar brains, the OOA grew significantly and the medulla emerged (epithelial architecture marked by Ecad, green). (T) The OOA/medulla in mad2­ sas-4 brain also enlarged in mid–third instar but was smaller than WT and lacked normal architecture. S’ and T’ depict cross-sectional views through the OOA/medulla.