Fig. 3

NEDD4 is a downstream target of FTO A m6A quantification assay in cells with shFTO. B Venn diagram showing a substantial and significant overlap of different peaks of m6A-enriched genes in resistant cells, with the RIP seq of genes bound to FTO, and different genes in FTO-overexpressing cells and database from GEO (GSE80617). C Different m6A peaks are enriched in the exonic region of NEDD4, as determined by m6A-seq. D, E RIP-qPCR assay was conducted to demonstrate the FTO-regulated NEDD4 m6A modification in CFPAC-GM and Colo357-GM cells. F FTO antibody was used to identify the results of an RNA pull-down test using NEDD4 full-length RNA. G, H FTO regulates NEDD4 expression in CFPAC-GM and Colo357-GM cells. I RNA stability assay showed that FTO can maintain NEDD4 mRNA stability. J NEDD4 cDNA was fused with either the WT or the mutant of m6A consensus sequence firefly luciferase reporter. K The relative activity of the WT or Mut luciferase reporters of NEDD4 was measured in FTO-silenced CFPAC-GM and Colo357-GM cells and normalized to the negative control groups