Fig. 2: METTL16 is a critical mediator of cuproptosis.

a m⁶A dot blot assays showing global m⁶A level of RNAs extracted from 40 pairs of GC tissues. RNAs were serially diluted and loaded equally with the concentration of 50 ng/μl. The methylene blue staining (below) was used to detect input RNA, while the intensity of dot immunoblotting (above) represented the level of m⁶A modification. Correlation between Cu concentration and m⁶A modification level in 48 pairs of GC tissues was shown on the right. P < 0.0001. b Changes in total m⁶A RNA methylation levels in HGC-27 cells treated with 1 or 5 μM CuCl₂. The m⁶A contents in GC were quantitated by RNA dot blotting. c–g METTL16 plays a crucial role in cuproptosis. Stable METTL16-knockdown HGC-27 (c, e–g) or AGS (d) cells were treated with indicated different concentrations of elesclomol-Cu (ratio=1:1) for 72 h and the growth survival rate was detected by CCK-8 assay. c The growth survival rate (left) and cell survival observed by microscope (right) in stable METTL16 knockdown HGC-27 cells under treatment with indicated different concentrations of elesclomol-Cu (ratio = 1:1) for 72 h (n = 3). P = 0.00503 (Sh1) and 0.00361 (Sh2). Scale bar, 50 μm. d The growth survival rate in stable METTL16 knockdown AGS cells under treatment with indicated different concentrations of elesclomol-Cu (ratio=1:1) for 72 h (n = 3). P = 0.00297 (Sh1) and 0.00970 (Sh2). e Stable METTL16-knockdown HGC-27 cells were treated with indicated different concentrations of disulfiram-Cu (ratio = 1:1) for 72 h (n = 3). P = 0.00990 (Sh1) and 0.00577 (Sh2). f Tetrathiomolybdate (TTM) (1 μM) supplement reversed the recovery of growth inhibition induced by shMETTL16 with different elesclomol-Cu (1:1) concentrations in HGC-27 cells (n = 3). P = 0.00198 (TTM). g, h The growth survival rate of verified METTL16-KO clones (KO-1 and KO-4) under treatment with indicated different concentrations of elesclomol-Cu (g) and disulfiram-Cu (h) for 72 h (n = 3). P = 0.00671 (g) and 0.00189 (h). Statistical data presented in this figure show mean values ± SD of three times of independent experiments. Statistical significance was determined by Two-tailed t test, *P < 0.05, **P < 0.01. Source data are provided as a Source data file.