Figure 1. Overexpression of constitutively active Src42A inhibits nuclear accumulation of dFOXO under conditions of starvation.

Fatbody clones from early 3^rd instar larvae are shown. Larvae were raised on yeast for 48 h and starved on PBS for 24 h. Immunostainings were performed with mouse α-GFP (green), rabbit α-dFOXO (red). The nuclear DAPI counterstain is shown in blue. (A–C) Wildtype cells show nuclear dFOXO while nuclear shuttling is blocked in cells expressing constitutively active Src42A. These cells are larger than wildtype cells, thus, they display no starvation-induced growth arrest. Genotype is y w hs-flp;; Act > CD2 > Gal4 UAS-GFP/UAS Src42A.CA. (D) Graphs show the perimeter of wildtype cells (WT) compared to cells expressing Src42A.CA (o/e) and the corrected total cell fluorescence (CTCF) calculated only for the nuclear region as a quantification of nuclear dFOXO abundance. Cells expressing consitutively active Src42A are significantly larger and have lower nuclear dFOXO levels than wildtype cells. (E–G) No significant differences in cell size or nuclear dFOXO are observed in Src42A.WT expressing cells. Genotype is y w hs-flp;; Act > CD2 > Gal4 UAS-GFP/UAS Src42A.WT. (H) Measuring the cell perimeter and calculating the nuclear fluorescence intensity shows that cells overexpressing wildtype Src42A are not significantly larger than wildtype cells and have similar nuclear dFOXO levels. (I–K) GFP-positive cells are slightly larger than wildtype cells and display nuclear dFOXO levels that are not significantly altered in comparison to wild-type cells. Genotype is y w hs-flp;; Act > CD2 > Gal4 UAS-GFP/UAS Src64B.CA. (L) Cells expressing constitutively active Src64B are significantly larger than wildtype cells but do not show a significant reduction of nuclear dFOXO signal. (M–O) Cells overexpressing the wildtype form of Src64B are of similar size like the surrounding cells and show slightly reduced starvation-induced nuclear shuttling of dFOXO. Genotype is y w hs-flp;; Act > CD2 > Gal4 UAS-GFP/UAS-Src64B.WT. (P) The graphs show that overexpression of a wildtype form of Src64B has no effect on cell perimeter, and a slight but significant one on nuclear fluorescence derived from the dFOXO antibody in fatbody cells. Scale bars represent 20 μm. Statistical significance was tested using an unpaired, two-tailed Student's t-test. Asterisks represent * = p < 0.05, ** = p < 0.01, *** = p < 0.001.