Fig. 3: IGF2BP3 promotes the degradation of FTO, resulting in increased total m6A levels and stabilization of CSF3 mRNA through an m6A-dependent mechanism. | Nature Communications

Fig. 3: IGF2BP3 promotes the degradation of FTO, resulting in increased total m6A levels and stabilization of CSF3 mRNA through an m6A-dependent mechanism.

From: Overcoming therapeutic resistance in oncolytic herpes virotherapy by targeting IGF2BP3-induced NETosis in malignant glioma

Fig. 3

a–d The impact of IGF2BP3 on total m6A levels a, b and m6A-related enzymes c, d. n = 3 biological independent experiments. e The interaction of IGF2BP3 with FTO mRNA. n = 3 biological independent experiments. f, g The impact of IGF2BP3 on FTO protein stability in U87MG cells. Quantification of FTO protein levels is shown g. n = 3 biological independent experiments. h, i The effect Chloroquine (50 μM) and MG132 (10 mM) on IGF2BP3-induced FTO protein stabilization in U87MG cells. The experimental design is shown h. n = 3 biological independent experiments for i. j, k The impact of chloroquine and MG132 treatments j as well as transfection with indicated HA-Ubiquitin mutants k on polyubiquitination modification of FTO protein in U87MG cells. n = 3 biological independent experiments. l–n The effect of FTO (Fto) on relative mRNA l, m and protein n levels of CSF3 (Csf3). n = 3 biological independent experiments for l, m. n = 6 or 5 biological independent samples for n. o, p, u The impact of FTO (Fto) knockdown o, p or IGF2BP3 overexpression u on the CSF3 (Csf3) mRNA stability. n = 3 biological independent experiments. q A schematic diagram of CSF3 mRNA with the predicted ‘m6A’ sites with the highest confidence at the 3’UTR highlighted in red is shown. r The m6A modification of CSF3 in U87MG cells. The enrichment of m6A was calculated by m6A-IP/input and IgG-IP/input. n = 3 biological independent experiments. s A schematic diagram of the CSF3 luciferase reporter. t, v The relative activity of the WT or MUT luciferase reporters, based on the psiCHECK2 plasmid was determined in U87MG cells. n = 3 biological independent experiments for t. n = 5 biological independent experiments for v. w The interaction between IGF2BP3 protein and CSF3 mRNA in U87MG cells. n = 3 biological independent experiments. Statistical significance was determined using two-tailed Student’s t-test in b, d, e, l–p, r, t, u, v, or one-way ANOVA in w. Data represent the mean ± SD. Source data are provided in the Source Data file.

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