Fig. 1

METTL3 regulates the expression and stability of RNGTT. (A) Genomic view of the 3’UTR of RNGTT. m⁶A pulldown peaks (REPIC) and METTL3 motif prediction (FIMO) are displayed as tracks in IgvR. (B) Representative western blot analysis of protein expression of METTL3, RNGTT, and loading control Vinculin done in triplicate. Original blots are presented in Supplementary Fig. 1. Band intensities were normalized to vinculin and sh- was set as 1. (C) mRNA levels of RNGTT analyzed by RT-qPCR normalized to GAPDH expression by ΔΔCT. Sh- was set to 1. Error bars represent S.E.M. in biological triplicates. * Represents student’s t-test p. value < 0.05. (D) m⁶A RNA immunoprecipitation on predicted site of m⁶A modification of RNGTT mRNA analyzed by RT-qPCR. Immunoprecipitation is represented as % input relative to recovery of m⁶A positive control. sh- was set to 100%. Error bars represent S.E.M. with biological replicates represented as a dot n ≥ 3. ** represents two-tailed student’s t-test p. value < 0.01. E) RNA stability assay in A549 sh-/shMETTL3 analyzed by RT-qPCR. RNA expression is represented as RNA levels normalized to GAPDH at the indicated timepoint relative to the RNA levels at timepoint 0 h in either sh- or shMETTL3. Error bars represent S.E.M. of biological replicates n = 4. * Represents two-tailed student’s t-test of sh- vs shMETTL3 p. value < 0.05 at each indicated timepoint, ** represents p. value < 0.01.