Fig. 4

Distinct codon signature at the ribosome A- and P-sites in self-renewing ESCs. a Illustration of a ribosome binding to mRNA protecting at least nine codons (position − 5 to + 3) from degradation by the RNase digestion. Positions − 2, − 1, and 0 correspond to the E-, P-, and A-site that host the charged tRNA. b The raw codon frequency was calculated for each specific codon (e.g. A-site) and normalized to the mean frequency across all nine positions. Calculations were done separately for self-renewing and differentiating samples. c–e Codon enrichment at the E- (c), P- (d), and A-sites (e). Stop codons are highlighted in red, and proline codons are highlighted in blue. The codons are sorted according to their frequency (shown in Additional file 1: Figure S3A-C). f, g Codon enrichment of stop (f) or all other codons (g) across all ribosome-protected codons. h Log₂ fold change of normalized codon usage in differentiated versus self-renewing hESCs across all ribosome-protected codons. Significantly different codons (Welch’s t-test; FDR correction) are marked in red (enriched) and blue (reduced) and occur mainly at the P-site. i Log₂ fold change of normalized codon usage in mouse differentiated (embryoid bodies) versus self-renewing embryonic stem cells (mESCs). Blue and red dots represent the significantly changed codon shown in h