Fig. 5
circRNA-SORE is modulated by m6A RNA methylation. a Predicted m6A site in circRNA-SORE from overlapping results of a sequence-based N6-methyladenosine (m6A) modification site predictor (SRAMP) and RMBase v2.0. b RNA pulldown assays and western blotting for YTHDF1/2, METTL3, and FTO using a circRNA-SORE probe. c Flow diagram of m6A-specific immunoprecipitation (MeRIP) assays. d MeRIP assays for m6A-modified circRNA-SORE in HepG2-SR and HepG2-WT cell lines. e MeRIP assays for m6A-modified circRNA-SORE in HepG2-SR cells transfected with si-NC, si-METTL3 or si-METTL14. f MeRIP assays for m6A-modified circRNA-SORE in HepG2-SR cells transfected with si-FTO or si-NC. g MeRIP assays for m6A-modified circRNA-SORE in HepG2-SR cells transfected with MAO-SORE or MAO-NC. MAO-SORE: sequence-specific morpholino antisense oligos blocking the m6A site in circRNA-SORE. h MeRIP assays for m6A-modified circRNA-SORE in HepG2-SR cells programmable edited by CRISPR-Cas9-ALKBH5 conjugates. gRNA-3: targeting the predicted m6A site in circRNA-SORE; gRNA-NC: negative control gRNA. i MeRIP assays for m6A-modified circRNA-SORE in HepG2-SR cells over-expressing circRNA-SORE with or without the m6A site mutated. j qPCR analysis of circRNA-SORE in HepG2-SR cells transfected with si-METTL3, si-METTL14 or si-NC. k qPCR analysis of circRNA-SORE in HepG2-SR and SKhep1-SR cells transfected with MAO-SORE or MAO-NC. l qPCR analysis of circRNA-SORE in HepG2 and SKhep1 cells transfected with MAO-SORE or MAO-NC. m qPCR analysis of circRNA-SORE in HepG2-SR cells transfected with MAO-SORE or MAO-NC under actinomycin D (5 μg/mL) treatment for 0 and 6 h. n qPCR analysis of circRNA-SORE in HepG2 cells over-expressing circRNA-SORE with or without the m6A site mutated under actinomycin D (5 μg/mL) treatment for 0, 3 and 6 h. Three different independent experiments with three technical repetitions were performed. Data are expressed as the mean ± SEM. Statistical analyses used Student’s t-test, Kaplan-Meier survival analysis and the log-rank test. p < 0.05 was considered statistically significant. * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001. NS; not statistically significant