Fig. 5: METTL3 facilitates pri-miR-17-92 processing by m6A modification on the A879 locus.
a Results of m6A-RIP and DGCR8-RIP in METTL3-overexpressing (lvM3) and control (lvCTL) MKN-45 cells. b Results of m6A-RIP and DGCR8-RIP in METTL3-reducing (shM3) and control (shCTL) HGC-27 cells. Input RNA and RNA pulled down by specific (anti-m6A or anti-DGCR8) antibodies (Sab) or polyclonal immunoglobulin G (IgG) were quantified by RT-PCR, and their ratios to corresponding inputs were shown in a, b. c Schematic diagram of the wild type pri-miR-17-92 (WT), miniMIR17HG, and pri-miR-17-92 with point mutations. “GGAC” motifs were not displayed in the diagram when mutated into “GGCC”. d Fold change (control vs. METTL3-reducing HGC-27 cells) of miR-19a, 19b, 20a, and their mean values when cells were transfected with vectors encoding pri-miR-17-92-WT or those with point mutations. Statistical significance is for comparisons between cells transfected with pri-miR-17-92-WT and those with mutations. e m6A-RIP in METTL3-reducing and control HGC-27 cells transfected with pri-miR-17-92-WT, A879, or empty vector (EV). RNAs pulled down by anti-m6A or IgG were quantified by RT-PCR, and their ratios to inputs are displayed. Statistical significance is for comparisons between the m6A-RIP data from cells transfected with WT and A879C. f, g Proliferation and migration assays of HGC-27 cells overexpressing pri-miR-17-92-WT, A879C, and EV. The indicated significances were for comparisons between cells overexpressing pri-miR-17-92-WT and A879C. n = 3 for each group. Data are presented as mean ± SD. nsP > 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.
