Figure 5. Knockdown of KATNB1 orthologs in zebrafish and Drosophila results in small brain phenotype.

katnb1 morpholino reduces zebrafish midbrain size: Confocal microscopy shows that the katnb1 morphants at (B) 9 nanogram (ng), (C) 15 ng, and (D) 30 ng have smaller midbrains (arrows) as compared with (A) control at 2 days post fertilization (dpf). The reduction in brain size is statistically significant (E). Zebrafish brain is labeled with green fluorescence by Tg(HuC:Kaede). (H) Left panel: Schematic of the Drosophila brain; box indicates brain lobe imaged. Right panel: Schematic of a single brain lobe marks the location of symmetrically dividing neuroepithelium (NE, green) and asymmetrically dividing neuroblasts (NBs, red). (I, J) Expression of kat80-IR with prospero-Gal4 results in a dramatically reduced brain size in 3^rd instar larvae. There is an overall reduction in the number of neurons and glia generated as seen by ELAV (red) and Repo (green) staining, respectively. (K, L) kat80-IR expressed under worniu-Gal4, UAS-mir::GFP, UAS-zeus::mCherry results in a significant reduction in NB number in central brain. Images in F–I are 3D projection of identical Z-sections. (F) Quantification of glial cell counts seen in panel F–G. There is significantly reduced number of glial cells in kat80-IR larvae (Error bars indicate SD, yw: 1080±110; kat80-IR: 673±116; two-tailed test, P=0.034). (G) GFP- and RFP-positive cells were quantified using 3D projections of identical Z-stacks from worniu>gal4 and worniu>kat80-IR brains, which reveal a significant reduction in central brain NBs per brain lobe (yw: 96.5±7.9; kat80-IR: 62.5±2.3; P = 0.002). (See also Figure S3)