The structural analysis of PAN RNA in METTL3 and FTO knockdown BCBL-1. (A) PAN RNA SHAPE-MaP reactivity profiles with blue trace corresponding to reactivity values obtained in METTL3 KD cells, red trace representing reactivity values from FTO KD cells. X-axis indicates nucleotide position on PAN RNA, y-axis represents SHAPE-MaP reactivity values. Nucleotides 1–14 and 1056–1077 were not covered in the analysis due to primer annealing. The green boxes indicate the local and global structural changes invoked by the methylome ablations. The position of the expression and nuclear retention (ENE) motif is indicated. (B) The secondary structure of PAN ENE motif (green) probed in wild-type, FTO and METTL3 knockdown (KD) cells. Nucleotides are colored by SHAPE reactivity as follows: gray residues display low reactivity (<0.2), pink residues representing medium reactivity (0.2–0.8), and red values representing high reactivity (>0.8). (C) Fraction of structural changes estimated for PAN RNA SHAPE profile obtained in wild-type BCBL-1 cells vs wild-type (background), FTO, and METTL3 knockdown cells. No change is marked in blue, local change is indicated in orange, while global change is colored in gray. (D) ROC curves generated for PAN RNA SHAPE profiles representing the m6A influence over PAN secondary structure. The x-axis corresponds to false positive rate (FPR); the y-axis represents true positive rate (TPR). The AUC column chart (right) indicates the level of agreement between nucleotide reactivity and pairing status. A value of 0.5 indicates that nucleotide reactivity reports no base-pairing information.
