Extended Data Fig. 4: Mapping the chromatin distribution of Mettl3 in paused ESCs.
From: m6A RNA methylation orchestrates transcriptional dormancy during paused pluripotency
a. m6A machinery (writers Mettl3, Mettl14 and Wtap; and erasers Fto and Alkbh5) in control (Ctrl) and paused ESCs by western blot in whole cell extracts (representative of 3 biological replicates). b. Increase of Mettl3 levels in chromatin extracts upon induction of paused pluripotency, measured by cell number-normalized (CNN) western blot (representative of 3 biological replicates). c. Strategy for Mettl3 ChIP-seq in ESCs with CNN approach using human cell spiking. d. Heatmap of the top 5000 most variable Mettl3 peaks by ChIP-seq across all samples, showing higher levels in paused ESCs (n = 2 biological replicates per group). e. Density plot of the average levels of Mettl3 binding in the TSS of all genes by ChIP-seq, separated by expression and methylation status, in control and paused ESCs. Mettl3 binding is highest in the TSS of expressed genes with a methylated transcript, and in paused ESCs. Number of genes (n) as indicated. Data as mean normalized Mettl3 level (n = 2 biological replicates per group). f. Examples of gene track views showing increased average levels (fold-change > 1.5) of m6A and Mettl3, by MeRIP-seq and Mettl3 ChIP-seq, respectively.
