Regulatory loop between Notch signaling and v-ATPase orchestrates NB growth and self-renewal. (a) Depletion of Vha68-2 by RNAi resulted in reduced protein expression of the Notch reporter E(spl)mγ-GFP. Larval brains were stained for Mira. (b) Quantification of E(spl)mγ-GFP signal intensity in control and Vha68-2^RNAi NBs. n (control) =3, n (Vha68-2^RNAi) = 3 brain lobes. (c) Expression levels of Notch target genes in sorted NBs assessed by quantitative RT-PCR upon depletion of Vha68-2 by RNAi. n = 3 experiments. (d) Knockdown of Notch in NBs resulted in decreased NB size. Larval brains were stained for Notch^Intra and Mira. (e) Quantification of NB diameter upon Notch depletion by RNAi. Each data point represents the mean type I NB diameter per brain lobe. n (control) = 13, n (Notch^RNAi) = 10. (f) Down-regulation of Vha68-2-GFP protein expression upon Notch^RNAi. Larval brains were stained for F-actin and Mira. Arrowheads indicate NBs. (g) Quantification of Vha68-2-GFP signal intensity in control and Notch^RNAi NBs. n (control) = 14, n (Notch^RNAi) = 14 brain lobes. (h) Expression levels of v-ATPase subunits in FACS-purified NBs assessed by quantitative RT-PCR upon Notch depletion by RNAi. n = 4 experiments. Error bars represent mean ± SD. **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; unpaired two-sided Student’s t test.