Figure 2. The InR-Tor pathway underlies the preferential neuronal growth under nutrient stress.

(A–F) ddaC neurons in the Gal4^ppk control (A and D) and animals expressing Gal4^ppk-driven InR RNAi (B and E) and Tor RNAi (C and F) in HY and LY conditions. (G) Quantification of normalized dendrite length (total dendrite length/segment width) in HY and LY conditions. HY: n = 14 for Gal4^ppk, n = 15 for InR RNAi, n = 11 for InR DN, n = 15 for Tor RNAi, n = 15 for Tor DN; LY: n = 14 for Gal4^ppk, n = 12 for InR RNAi, n = 14 for InR DN, n = 15 for Tor RNAi, n = 15 for Tor DN. Two-way ANOVA, Posthoc contrasts with a Dunnett correction. The differences between control and InR RNAi under HY and LY conditions are significantly different as indicated by a significant interaction term (p=0.003924), the same for InR DN (p=8.581e-09), Tor RNAi (p=5.876e-07) and Tor DN (p=1.062e-09). (H) The ratios of average normalized dendrite length between LY and HY. (I–K) ddaC neurons in the Gal4^R38F11 control (I) and animals expressing Gal4^R38F11-driven InR RNAi (J) and Tor RNAi (K) in HY condition. (L) Quantification of normalized dendrite length in e Gal4^R38F11-driven knockdown and overexpression. n = 23 for Gal4^R38F11, n = 17 for InR RNAi, n = 17 for InR DN, n = 16 for Tor RNAi, n = 17 for Tor DN. One-way ANOVA and Tukey’s HSD test. (M–O) pRpS6 staining (magenta) of ddaC neurons (Green) and epidermal cells in HY and LY conditions in 2-dimensional (2D) projections. The insets in (M) and (N) show pRpS6 staining at the soma (1) and primary dendrites (2) in single confocal sections, with the somas and dendrites outlined. High settings and low settings stand for high and low pRpS6 detection settings. (P) Quantification of pRpS6 intensity ratio (soma/epidermal cells and dendrites/epidermal cells) in HY and LY conditions. Soma/epi: n = 17 for HY, n = 20 for LY; dendrites/epi: n = 16 for HY, n = 20 for LY. Two-way ANOVA. The differences between HY and LY in soma and dendrites are not significantly different as indicated by a non-significant interaction term (p=0.5592). For all quantifications, ***p<0.001; *p<0.05; ns, not significant. each circle represents a neuron. Significance level is for comparison between the control and the genotype indicated under the same food condition. Black bars, mean; red bars, SD. Scale bars, 100 μm in (A–K); 10 μm in (M–O).

Figure 2—figure supplement 1. The effects of suppressing InR and Tor in ddaC neurons and epidermal cells in HY and LY conditions.

(A–F) ddaC neurons in the Gal4^ppk control (A and D) and animals expressing Gal4^ppk-driven InR DN (B and E) and Tor DN (C and F) in HY and LY conditions. (G) Larval body expression patterns of Gal4^R16D01 > GFPnls in both maximum projection and a deeper section. The arrow indicates expression in an internal organ in the head region. Gal4^R16D01 is also expressed in fewer than 20 neurons in the brain (FlyLight database). (H) Larval body expression patterns of Gal4^R38F11 > GFPnls in both maximum projection and a deeper section. The cyan and yellow arrows indicate expression in an internal organ in the head region and in the hindgut, respectively. Gal4^R38F11 is also expressed in a small number of sparse neurons in the brain (FlyLight database). (I–M) Epidermal cells in the Gal4^R16D01 control (I) and animals expressing Gal4^R16D01-driven InR RNAi (J), Tor RNAi (K), InR DN (L) and Tor DN (M) in the HY condition. Gal4^R16D01 domain is labeled by mIFP expression (magenta). All epidermal cells are labeled by Nrg-GFP (green). The blue and yellow overlays indicate the measured Gal4-expressing and wildtype (WT) epidermal cells, respectively. (N) Quantification of epidermal cell size ratio (Gal4^R16D01 cells/WT cells). Each circle represents a segment; n = 18 for Gal^R16D01, n = 20 for InR RNAi, n = 14 for InR DN, n = 18 for Tor RNAi, n = 14 for Tor DN. ***p<0.001; One-way ANOVA and Tukey’s HSD test. Significance level is for comparison between the control and the genotype indicated under the same food condition. Black bars, mean; red bars, SD. (O–Q) ddaC neurons in the Gal4^R38F11 control (O) and animals expressing Gal4^R38F11-driven InR DN (P) and Tor DN (Q) in the HY condition. (A), (D), (O) are the same as Figures 2A, D and I, respectively. Scale bars, 100 μm in (A–F) and (O–Q), 50 μm in (G–M).