Journal: bioRxiv

Article Title: Multiple FGFR1 mutations modulate tumorigenic mechanisms in glioneuronal tumors

doi: 10.1101/2025.05.27.654799

Figure Lengend Snippet: R661P reduces K656E and N546K mutant protein stability by rescuing lysosomal degradation. A) Receptor stability experimental design: HEK293 cells expressing WT and mutant FGFR1 are stimulated with human FGF2 (hFGF2, 25 ng/mL) to induce receptor internalization and degradation and, in parallel, with cycloheximide (100 µg/mL) to block protein synthesis. B and D) Western blot analysis of a representative time-course experiment comparing WT FGFR1, K656E (B) and N546K (D) single and double mutants showing total Flag-FGFR1 protein levels at 0, 3 and 6 hours post-treatment with FGF2 and cycloheximide (CHX). C and E) Relative amounts of WT, single and double mutant FGFR1 protein obtained by quantifying western blots from n = 3 independent experiments. Values have been normalized (FC) against each relative 0h reference values. Data is represented by mean ± SEM and significant variations in protein amounts against each specific reference values have been indicated (ANOVA test, p value * ≤ 0.05, ** ≤ 0.01, *** ≤ 0.001) in the bar plots.

Article Snippet: Next, cells were induced with 25 ng/mL of recombinant human FGF2 (RD Systems) and treated with 100 µg/mL of Cycloheximide (Merck) and 200 nM of Bafilomycin A1 (Merck).

Techniques: Mutagenesis, Expressing, Blocking Assay, Western Blot