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. 2019 Oct 15;8:e49309. doi: 10.7554/eLife.49309

Figure 4. IIS promotes ETC activity via inhibition of GSK3 to stabilize Myc.

(A) Schematic of the conserved IIS pathway that inhibits 4E-BP/Thor and GSK3/Sgg to promote protein translation and suppress protein degradation, respectively. (B) Germaria from ovaries of ctrl; chicoRNAi; chicoRNAi, thorRNAi; and chicoRNAi, sggRNAi driven by nos-Gal4 endogenously expressing Myc-GFP and stained with anti-GFP (green) and anti-1B1 (red). Scale bars, 5 μm. (C, D) Quantification of Myc-GFP intensity (C) and COX activity (D) in germarium region 2B from ovaries with indicated genotypes, normalized to the intensity or activity values in germaria with ctrl over-expression. (C) n = 10 germaria for each genotype. (D) n = 10 germaria for each genotype. Error bars represent SEM. *p<0.05. (E) Germaria from ovaries of ctrl and sggRNAi driven by nos-Gal4 endogenously expressing Myc-GFP and stained with anti-GFP and anti-1B1. Myc protein is up-regulated in both region 2A and region 2B germ cells in the sgg RNAi ovary. Scale bars, 10 μm. (F) Quantification of relative Myc-GFP intensity in germarium region 2B from ovaries of ctrl and sggRNAi driven by nos-Gal4. Myc-GFP intensity is normalized to that of region 2B cysts with ctrl expression. n = 7 germaria for each genotype. Error bars represent SEM. *p<0.01. (G) A germarium from wt flies stained with anti-GSK3 and anti-p-GSK3. Germaria are outlined with dotted lines. Scale bars, 5 μm. (H) Upper panel: a representative image of a germarium from ovaries expressing endogenous InR-EGFP. Lower panel: visualization of the InR mRNA in germarium by FISH with fluorescently labeled DNA probes. Germaria are outlined with dotted lines. Scale bars, 5 μm.

Figure 4—source data 1. Relative COX activity and Myc-GFP intensity in the germarium.
elife-49309-fig4-data1.xlsx (14.5KB, xlsx)
DOI: 10.7554/eLife.49309.014

Figure 4.

Figure 4—figure supplement 1. Myc is post-transcriptionally regulated by the IIS-GSK3 cascade.

Figure 4—figure supplement 1.

(A) Germaria from ovaries of ctrl, chicoRNAi, and bskRNAi driven by nos-Gal4 expressing LacZ under the control of myc endogenous promoter stained with anti-β-galactosidase and anti-1B1. Scale bars, 10 μm. (B) A representative live imaging of germaria from flies expressing GFP-tagged Sgg under its endogenous promoter. Scale bar, 10 μm. (C) A germarium from wt flies endogenously expressing Myc-GFP stained with anti-GFP (green) and anti-p-GSK3 (red). Germarium regions are indicated. Myc protein highly corresponds to inhibited form of GSK3. Scale bar, 10 μm.