Fig. 5: FGFR2b regulates mTOR signalling and autophagy from the recycling endosomes. | Nature Communications

Fig. 5: FGFR2b regulates mTOR signalling and autophagy from the recycling endosomes.

From: Spatially resolved phosphoproteomics reveals fibroblast growth factor receptor recycling-driven regulation of autophagy and survival

Fig. 5

a Subnetwork of proteins annotated to mTOR pathway or autophagy based on KEGG analysis from Fig. 4h. Node colouring indicates whether the phosphorylated protein or the phosphorylated sites from the KEGG term ’autophagy’ were found in global, proximal phosphoproteome or both. Sites from Fig. 2d have a green border. b Immunoblot analysis (N = 3 independent biological replicates) with indicated antibodies of candidate phosphorylated proteins from the subnetwork (Fig. 5a). T47D were transfected with RAB11-APEX2 (T47D_RAB11-APEX2) stimulated with FGF10 for the indicated time points. UT, treatment with vehicle as control. Non proximal and proximal samples represent the supernatant and the pulldown following enrichment of biotinylated samples with streptavidin beads, respectively, and run against total lysates (total). c Autophagy measured using fluorescence-activated cell sorting (FACS) of T47D cells in serum, treated with vehicle (UT), FGF7 or FGF10 for 2 h. Representative images and gating are shown in Supplementary Fig. 6b–c. The number of cells counted is indicated below the graph. N = 3 independent biological replicates. p value <0.05 *, p value <0.0005 *** (one-way ANOVA with Tukey test). d Autophagy (measured by staining of autolysosomes with acridine orange) of HeLa_FGFR2bST, T47D and BT20 treated with vehicle (UT), FGF7 or FGF10. N = 3 independent biological replicates where at least six treated wells of cells were counted for each N. p value <0.001*** (one-way ANOVA with Tukey test). e Immunoblot analysis (N ≥ 3 independent biological replicates) with the indicated antibodies of the effect of serum starvation and FGF treatment on autophagic markers in T47D. LC3B 2 is the lipidated form of LC3B. Cells were treated with vehicle (UT), FGF7 or FGF10. f Immunoblot analysis (N ≥ 3 independent biological replicates) with the indicated antibodies of HeLa_FGFR2bST, T47D and BT20 treated or not with FGF7 or FGF10 for 2 h. Source data are provided as a Source Data file.

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