Fig. 5: MTA1 is regulated by FTO in an m⁶A-dependent manner.

A The possible m⁶A sites in MTA1 transcripts. B M⁶A- RIP seq was performed in HCT116 and DLD1 cells, and the overlay track represents the m⁶A peaks with blue for input and red for IP. C Enrichment of m⁶A in different regions of MTA1 mRNA was detected by m⁶A-RIP qPCR assay in HCT116 cells (left). The m⁶A methylation of MTA1 mRNA regions in siFTO#1 versus siNC HCT116 cells by m⁶A MeRIP analysis (right). IgG was used as the negative control. The relative m⁶A enrichment was normalized to the input. The sequences are shown in Table S7. D The MTA1 mRNA decay rate was analyzed by nonlinear regression (left), and the relative mRNA expression level of MTA1 was compared between the two groups at each time point (right) in HCT116 cells. E Enrichment of m⁶A in site 4 of MTA1 mRNA was detected by m⁶A-RIP qPCR assay in CRC tumor tissues versus paired adjacent normal tissues (n = 24). F Relative MTA1 RNA expression levels in DMSO, FB23-2 (5 μM) and FB23-2 (10 μM) treated HCT116 and DLD1 cells. G Relative MTA1 RNA expression levels in empty vector (EV), FTO overexpression wild type (FTO), FTO overexpression with R316Q/R322Q mutant (FTO^R316Q/R322Q) and FTO overexpression with H231A/D233A mutant (FTO^H231A/D233A) HCT116 and DLD1 cells. H Relative m⁶A level of MTA1 mRNA regions in FTO versus FTO^H231A/D233A HCT116 cells by m⁶A MeRIP analysis. IgG was used as the negative control. Relative m⁶A level was normalized to the input. I The MTA1 mRNA decay rate in DMSO versus FB23-2 (10 μM) treated HCT116 cells (left), and the relative mRNA expression level of MTA1 was compared between the two groups at each time point (right). J The MTA1 mRNA decay rate in EV, FTO, and FTO^H231A/D233A HCT116 cells (left), and the relative MTA1 mRNA expression level in each group at the time point (right). Data in (C, D, F, G, H, I, and J) are presented as the means ± S.D. (n = 3) *P < 0.05, **P < 0.01, ***P < 0.001 (Student’s t test).